Synthesis of chemical building blocks from C5-C6 Sugars

Scout intake sheet

5
Challenge description

The industry faces a significant challenge changing itself from a fossil feedstock based industry towards a biobased industry. Several important steps are already taken concerning the valorisation of biomass. In the industry, there is an opportunity to valorise the hemicellulose waste stream further. Therefore RVO and BCB have set up a working group to explore the possibilities of hemicellulose. This scouting project aims at further processing the C5-C6 sugars in hemicellulose (arabinose, xylose, mannose, galactose, ribose, lyxose) to chemical building blocks with excellent synthesis properties (low oxygen level, good selectivity). Examples of such a building block are 1,5-Pentanediol and ethylene oxide. These building blocks can be used to produce new biobased products. The type of building blocks will remain open in this scouting project, aiming at exploring new conversion routes and products.

Scope
Discover Demonstrate Develop Deploy
Current known technique(s)
  • Xylitol
  • Furfural (& derivatives)
  • Bio-ethanol from C5
  • D-Xylulose
  • Xylonic acid
  • Arabinonic acid
  • New aromatics (e.g. methylphthalic acid)
Ideal outcome

An overview of C5-C6 sugar synthesis or conversion routes to chemical building blocks with low oxygen level **Minimum outcome** An overview of C5-C6 sugar synthesis or conversion routes to new chemical building blocks

Objective(s)
  • Bulk industry application
  • Good atom efficiency
  • Low oxygen level in molecule ~ O2-O3
Constraint(s)
  • Cost (C-yield; Mass & Energy balance; selectivity)
  • Complexity of process (steps)
  • Medium/conditions
Functions
Action = [convert] OR [valorize] OR [synthesize] OR [produce] OR [convert] OR [use] OR [synthesize] OR [optimize] OR [be]

Object = [C5-C6 sugar] OR [hemicellulose] OR [platform chemical] OR [alcohol] OR [furfural] OR [C6] OR [chemical] OR [production strain] OR [PHA]

Environment = [bulk] OR [chemical] OR [building block] OR [bio-based] OR [Pentose] OR [hemicellulose] OR [arabinose] OR [conversion route] OR [oxygen level] OR [C5 sugar] OR [pathway] OR [ferment] OR [synthetic] OR [synthetic route] OR [C5] OR [C5 sugar] OR [sugar] OR [PHA]
Terminology
  • biobased, synthesis, conversion, building blocks, bulk
Case Confirmation
Confirmed by
Comments

Preliminary Results

Published 8/5/19

Based on the case described above we have executed the first line of queries in IGOR^AI. The goal was to obtain a broad set of (bio)synthesis routes to chemical compounds that can be derived from C5-C6 sugars. 7 concepts are distinguished based on the results: 1. Biological routes to alcohols 2. Biological routes to organic acids 3. Other biological routes 4. Synthetic routes to organic acids 5. Synthetic routes to alcohols 6. Synthetic routes to furfural-like compounds 7. Other synthetic routes Every concept comprises multiple (bio)synthesis routes to chemical compounds (44 in total). Below the table, short descriptions, research findings and sources per (bio)synthesis routes are listed as well. You can use this information to get a better understanding of the (bio)synthesis routes. During the midway meeting, we would like to discuss the (bio)synthesis routes to chemical compounds and concepts, determine their relevance and select the top selection that needs to be deepened in the second phase of the project.

To determine which technologies are relevant to proceed to the next scouting phase you can play the technology selection game by clicking on the button below.

Concept Technology Selection
1. Biological routes to alcohols
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1.1 Ethanol

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1.2 Ethylene glycol

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1.3 ABE fermentation

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1.4 2,3-butanediol

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1.5 1,2,4-Butanetriol

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1.6 1-hexadecanol

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1.7 Xylitol

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2. Biological routes to organic acids
2.1 Glycolic acid

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2.2 3-hydroxypropionic acid

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2.3 Butyric acid

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2.4 Fumaric acid

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2.5 Lactic acid

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2.6 Malic acid

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2.7 Succinic acid

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2.8 Itaconic acid

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2.9 3-dehydroshikimic acid

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3. Other biological routes
3.1 Biohydrogen

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3.2 Polyhydroxyalkanoates

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3.3 Terpenes

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3.4 Diaminopentane

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3.5 Amino acids

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3.6 Glucose 2,6 succinate

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4. Synthetic routes to organic acids
4.1 Aldose oxidation by peroxide

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4.2 Catalytic conversion to lactic acid

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4.3 Galactose hydrolysis to levulinic acid

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4.4 Galactose oxidation to mucic acid

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4.5 Gas-solids heterogeneous partial oxidation

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4.6 Pentose oxidation to tetronic acid

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4.7 Wet oxidation

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4.8 Dehydrogenation of sugar alcohols to lactic acid

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4.9 Hydrogenolysis of xylitol

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4.10 Methyl levulinate (highly recalcitrant cellulose and part hemicellulose)

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4.11 Acid catalysed levulinic acid/ester production

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5. Synthetic routes to alcohols
5.1 Xylose catalytic dehydrogenation

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5.2 From furfural: 1,5-pentanediol

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5.3 Catalyst single step conversion of C5 and C6 sugars to ethylene glycol, propylene glycol & glycerol

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6. Synthetic routes to furfural-like compounds
6.1 Xylose dehydration to furfural

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6.2 Microwave-assisted furfural production

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6.3 Catalytic coversion of hexoses to 5HMF

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7. Other synthetic routes
7.1 Conversion route of pentoses to 3,4-dihydroxyprolines

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7.2 Xylose conversion to xylonic acid and/or xylitol

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7.3 Furfural to methylfuorate

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7.4 Furfural to cyclopentanone/ cylopentanol

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7.5 Xylose conversion in alkaline environment

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1. Biological routes to alcohols

Back

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1.1 Ethanol

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Ethanol (also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is a chemical compound, a simple alcohol, and is often abbreviated as EtOH. Ethanol is a volatile, flammable, colorless liquid with a slight characteristic odor. Ethanol is an important industrial ingredient. It has widespread use as a precursor for other organic compounds such as ethyl halides, ethyl esters, diethyl ether, acetic acid, and ethyl amines. [Wiki](https://en.wikipedia.org/wiki/Ethanol#Feedstock) **Research Findings** - Recombinant strains containing genes coding for xylose reductase (XR) and xylitol dehydrogenase (XDH) from the xyloseutilizing yeast Pichia stipitis have been reported; however, such strains ferment xylose to ethanol poorly. One reason for this may be the low capacity of xylulokinase, the third enzyme in the xylose pathway. To investigate the potential limitation of the xylulokinase step, we have overexpressed the endogenous gene for this enzyme (XKS1) in S. cerevisiae that also expresses the P. stipitis genes for XR and XDH. The metabolism of this recombinant yeast was further investigated in pure xylose bioreactor cultivation at various oxygen levels. The results clearly indicated that overexpression of XKS1 significantly enhances the specific rate of xylose utilization. In addition, the XKoverexpressing strain can more efficiently convert xylose to ethanol under all aeration conditions studied. One of the important illustrations is the significant anaerobic and aerobic xylose conversion to ethanol by the recombinant Saccharomyces; moreover, this was achieved on pure xylose as a carbon. Under microaerobic conditions, 5.4 g L−1 ethanol was produced from 47 g L−1 xylose during 100 h. Art. [#ARTNUM](#article-25157-2042082999)

1.1.1 1.1 Ethanol
Conversion of xylose to ethanol by recombinant Saccharomyces cerevisiae: importance of xylulokinase (XKS1) and oxygen availability.
The yeast Saccharomyces cerevisiae efficiently ferments hexose sugars to ethanol, but it is unable to utilize xylose, a pentose sugar abundant in lignocellulosic materials. Recombinant strains containing genes coding for xylose reductase (XR) and xylitol dehydrogenase (XDH) from the xylose-utilizing yeast Pichia stipitis have been reported; however, such strains ferment xylose to ethanol poorly. One reason for this may be the low capacity of xylulokinase, the third enzyme in the xylose pathway. To investigate the potential limitation of the xylulokinase step, we have overexpressed the endogenous gene for this enzyme (XKS1) in S. cerevisiae that also expresses the P. stipitis genes for XR and XDH. The metabolism of this recombinant yeast was further investigated in pure xylose bioreactor cultivation at various oxygen levels. The results clearly indicated that overexpression of XKS1 significantly enhances the specific rate of xylose utilization. In addition, the XK-overexpressing strain can more efficiently convert xylose to ethanol under all aeration conditions studied. One of the important illustrations is the significant anaerobic and aerobic xylose conversion to ethanol by the recombinant Saccharomyces; moreover, this was achieved on pure xylose as a carbon. Under microaerobic conditions, 5.4 g L−1 ethanol was produced from 47 g L−1 xylose during 100 h. In fed-batch cultivations using a mixture of xylose and glucose as carbon sources, the specific ethanol production rate was highest at the highest aeration rate tested and declined by almost one order of magnitude at lower aeration levels. Intracellular metabolite analyses and in vitro enzyme activities suggest the following: the control of flux in a strain that overexpresses XKS1 has shifted to the nonoxidative steps of the pentose phosphate pathway (i.e., downstream of xylose 5-phosphate), and enzymatic steps in the lower part of glycolysis and ethanol formation pathways (pyruvate kinase, pyruvate decarboxylase, and alcohol dehydrogenase) do not have a high flux control in this recombinant strain. Furthermore, the intracellular ATP levels were found to be significantly lower for the XK strain compared with either the control strain under similar conditions or glucose-grown Saccharomyces. The ATP : ADP ratios were also lower for the XK strain, especially under microaerobic conditions (0.9 vs 6.4).
7/1/01 12:00:00 AM
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1.1.2 1.1 Ethanol
Engineering xylose and arabinose metabolism in recombinant Saccharomyces cerevisiae
Utilization of all sugars in lignocellulose hydrolysates is a prerequisite for economically feasible bioethanol production. The yeast commonly used for industrial ethanol production, Saccharomyces cerevisiae, is naturally unable to utilize pentose sugars xylose and arabinose, which constitute a large fraction of many lignocellulosic materials. Xylose utilization by S. cerevisiae can be achieved by heterologous expression of a xylose utilization pathway, consisting either of xylose reductase (XR), xylitol dehydrogenase (XDH) and xylulokinase (XK), or alternatively, of xylose isomerase (XI) and XK. Xylitol formed by XR is a major by-product in xylose fermentation when using the XR-XDH pathway. In this thesis, high-level expression of both XR and XDH was shown to decrease xylitol formation. The influence of other genetic modifications was also evaluated. It was shown that the overexpression of the non-oxidative pentose phosphate pathway (PPP) genes enables efficient growth on xylose and xylose fermentation, provided that the initial xylose pathway is expressed at a high level. When comparing the two xylose utilization pathways, higher ethanol productivity was achieved using the XR-XDH pathway, whereas higher ethanol yield was achieved with the XI pathway. The industrial xylose-fermenting S. cerevisiae strain TMB 3400, which has been previously generated by mutagenesis and selection, was tested for fermentation of lignocellulose hydrolysate. TMB 3400 displayed significantly better fermentation performance compared to the laboratory strains tested, highlighting the need for robust industrial strains in lignocellulose fermentation. TMB 3400 was also characterized by proteome analysis using difference in-gel 2-D electrophoresis. Consistently with the results obtained in other studies, increased activities of XR, XDH and a PPP enzyme TKL were found. The bacterial arabinose utilization pathway was introduced into TMB 3400, which resulted in the novel glucose, xylose and arabinose co-fermenting strain TMB 3063, with ethanol, xylitol and arabitol as the main fermentation products. (Less)
1/1/06 12:00:00 AM
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1.1.3 1.1 Ethanol
Method for producing low carbon chemical ethanol from C6 sugar alcohol
The invention relates to a method for producing low carbon chemical ethanol from C6 sugar alcohol. Chinese patent disclosed number CN1683293A discloses a method for producing dihydric alcohol and andpolyhydric alcohol by cracking sorbierite. The method comprises the steps of converting the sorbierite into glycol, propanediol and glycerin, but process data and a process route as well as products do not achieve good industrial production requirements. The method mainly adopts a continuous process to hydrocrack the C6 sugar alcohol to generate a low carbon ethanol mixture and finally obtains a single (or mixed) product by separating and refining in alkaline environment, in the presence of a nickel chrome catalyst and at high temperature and high pressure. The process route has the advantages of simple process, easy operation, high yield in unit time and variety of the products; and as the catalyst has different selectivities, the composition rate of the produced products can be substantially adjusted.
9/30/09 12:00:00 AM
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1.1.4 1.1 Ethanol
Method and apparatus for conversion of cellulosic material to ethanol

1. A method for treatment of cellulosic material comprising the steps of:exposing said cellulosic material to a first temperature or pressure condition in a first reactor section having an inlet and an outlet,separating a liquid fraction from a solid fraction following the first temperature or pressure condition by pressing a solid fraction of the cellulosic material at the outlet of the first reactor section, wherein the liquid fraction is subsequently subjected to fermentation, andexposing said solid fraction to a second temperature or pressure condition in a second reactor section following said separation of a liquid fraction from the solid fraction. 2. The method of claim 1, wherein a subsequent liquid fraction is separated from said solid fraction following exposure to said second temperature or pressure condition. 3. The method of claim 2, wherein the concentration of lignin in the solid fraction following exposure to the second temperature or pressure condition and separation of said subsequent liquid fraction is higher than the lignin concentration in the cellulosic material prior to exposure to said first temperature or pressure condition. 4. The method of claim 1, wherein the severity of the second temperature or pressure condition is higher than the severity of the first temperature or pressure condition. 5. The method of claim 1, wherein the first temperature or pressure condition comprises exposing said cellulosic material to a temperature between 170° C. and 230° C. 6. The method of claim 1, wherein the cellulosic material is soaked at ambient pressures at temperatures up to 100° C. prior to exposure to the first set of temperature or pressure conditions. 7. The method of claim 6, wherein a liquid fraction is separated from the cellulosic material after said soaking and prior to exposure of the first set of temperature or pressure conditions. 8. The method of claim 1, wherein said first and second temperature or pressure conditions take place in separate zones of a single reactor. 9. The method of claim 1 wherein said first and second temperature or pressure conditions take place in separate reactors. 10. The method of claim 1, wherein said treatment method is continuous. 11. The method of claim 1, wherein said treatment occurs at increased pressure when compared to atmospheric pressure. 12. The method of claim 11, wherein said increased pressure is not released until after said second temperature or pressure condition. 13. The method of claim 1, wherein the pressing is accomplished under pressurized conditions. 14. The method of claim 13 wherein the pressing under pressurized conditions is accomplished using a screw press. 15. The method of claim 1, wherein the solid fraction is subsequently subject to enzymatic liquefaction and saccharification. 16. The method of claim 15, wherein the solid fraction is further fermented to produce ethanol following enzymatic liquefaction and saccharification. 17. The method of claim 1, wherein the solid fraction is cooled by a counter current wash stage prior to exposure to said second set of temperature or pressure conditions. 18. The method of claim 1, wherein said fermentation converts C5 sugars of the liquid fraction to ethanol. 19. The method of claim 1 wherein the feedstock is wheat straw, bagasse, corn stover, or cereal straw. 20. The method of claim 1, wherein the second temperature or pressure condition comprises a higher temperature and pressure than the temperature and pressure of the first temperature or pressure condition. 21. The method of claim 1, wherein the liquid fraction undergoes hydrolysis following said separating of a liquid fraction and prior to subjecting the liquid fraction to fermentation. 22. The method of claim 21, wherein the pH of the liquid fraction is adjusted prior to subjecting the liquid fraction to fermentation. 23. The method of claim 1, wherein the pH of the liquid fraction is adjusted prior to subjecting the liquid fraction to fermentation.
5/1/15 12:00:00 AM
Link to Patent

1.2 Ethylene glycol

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Ethylene glycol (IUPAC name: ethane-1,2-diol) is an organic compound with the formula (CH2OH)2. It is mainly used for two purposes, as a raw material in the manufacture of polyester fibers and for antifreeze formulations. It is an odorless, colorless, sweet-tasting, viscous liquid. [Wiki](https://en.wikipedia.org/wiki/Ethylene_glycol) **Research Findings** - These pathways were incorporated into Escherichia coli , and engineered strains produced ethylene glycol from various pentoses, including simultaneously from D xylose and L arabinose; one strain achieved the greatest reported titer of ethylene glycol, 40 g/L, from D xylose at a yield of 0.35 g/g. Art. [#ARTNUM](#article-25156-2214411608) - Here, we developed an alternative xylose utilization pathway that largely bypasses the PPP. In the new pathway, dxylulose is converted to dxylulose1phosphate, a novel metabolite to S. cerevisiae, which is then cleaved to glycolaldehyde and dihydroxyacetone phosphate. This synthetic pathway served as a platform for the biosynthesis of ethanol and ethylene glycol. Art. [#ARTNUM](#article-25156-2462386339) - The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative Dxylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding Dxylose dehydrogenase (XylB) and Dxylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from Dxylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2keto3deoxyxylonic acid [(S)4,5dihydroxy2oxopentanoic acid]/ (2K3DXA) was produced and Dxylonic acid accumulated to ca. 9 g/L. A significant amount of Dxylonic acid (ca. 14%) was converted to 3deoxypentonic acid (3DPA), and also, 3,4dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldoketo reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. Art. [#ARTNUM](#article-25156-2765987778)

1.2.1 1.2 Ethylene glycol
Bypassing the Pentose Phosphate Pathway: Towards Modular Utilization of Xylose.
The efficient use of hemicellulose in the plant cell wall is critical for the economic conversion of plant biomass to renewable fuels and chemicals. Previously, the yeast Saccharomyces cerevisiae has been engineered to convert the hemicellulose-derived pentose sugars xylose and arabinose to d-xylulose-5-phosphate for conversion via the pentose phosphate pathway (PPP). However, efficient pentose utilization requires PPP optimization and may interfere with its roles in NADPH and pentose production. Here, we developed an alternative xylose utilization pathway that largely bypasses the PPP. In the new pathway, d-xylulose is converted to d-xylulose-1-phosphate, a novel metabolite to S. cerevisiae, which is then cleaved to glycolaldehyde and dihydroxyacetone phosphate. This synthetic pathway served as a platform for the biosynthesis of ethanol and ethylene glycol. The use of d-xylulose-1-phosphate as an entry point for xylose metabolism opens the way for optimizing chemical conversion of pentose sugars in S. cerevisiae in a modular fashion.
6/23/16 12:00:00 AM
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1.2.2 1.2 Ethylene glycol
Efficient utilization of pentoses for bioproduction of the renewable two-carbon compounds ethylene glycol and glycolate.
Abstract The development of lignocellulose as a sustainable resource for the production of fuels and chemicals will rely on technology capable of converting the raw materials into useful compounds; some such transformations can be achieved by biological processes employing engineered microorganisms. Towards the goal of valorizing the hemicellulose fraction of lignocellulose, we designed and validated a set of pathways that enable efficient utilization of pentoses for the biosynthesis of notable two-carbon products. These pathways were incorporated into Escherichia coli , and engineered strains produced ethylene glycol from various pentoses, including simultaneously from D -xylose and L -arabinose; one strain achieved the greatest reported titer of ethylene glycol, 40 g/L, from D -xylose at a yield of 0.35 g/g. The strategy was then extended to another compound, glycolate. Using D -xylose as the substrate, an engineered strain produced 40 g/L glycolate at a yield of 0.63 g/g, which is the greatest reported yield to date.
3/1/16 12:00:00 AM
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1.2.3 1.2 Ethylene glycol
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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1.2.4 1.2 Ethylene glycol
Production of ethylene glycol or glycolic acid from D-xylose in Saccharomyces cerevisiae
The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative D-xylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding D-xylose dehydrogenase (XylB) and D-xylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from D-xylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2-keto-3-deoxyxylonic acid [(S)-4,5-dihydroxy-2-oxopentanoic acid] (2K3DXA) was produced and D-xylonic acid accumulated to ca. 9 g/L. A significant amount of D-xylonic acid (ca. 14%) was converted to 3-deoxypentonic acid (3DPA), and also, 3,4-dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldo-keto reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. The D-xylonate dehydratase activity in yeast was notably low, possibly due to inefficient Fe–S cluster synthesis in the yeast cytosol, and leading to D-xylonic acid accumulation. The dehydratase activity was significantly improved by targeting its expression to mitochondria or by altering the Fe–S cluster metabolism of the cells with FRA2 deletion.
11/1/17 12:00:00 AM
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1.3 ABE fermentation

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Acetone, Butanol and Ethanol can be produced from xylose. Acetone–butanol–ethanol (ABE) fermentation is a process that uses bacterial fermentation to produce acetone, n-Butanol, and ethanol from carbohydrates such as starch and glucose. [Wiki](https://en.wikipedia.org/wiki/Acetone%E2%80%93butanol%E2%80%93ethanol_fermentation) **Research Findings** - Here, we reported a useful metabolic engineering strategy to improve d xylose consumption by C. beijerinckii . Gene cbei2385, encoding a putative d xylose repressor XylR, was first disrupted in the C. beijerinckii NCIMB 8052, resulting in a significant increase in d xylose consumption. A d xylose protonsymporter (encoded by gene cbei0109) was identified and then overexpressed to further optimize d xylose utilization, yielding an engineered strain 8052xylRxylT ptb ( xylR inactivation plus xylT overexpression driven by ptb promoter). We investigated the strain 8052xylRxylT ptb in fermenting xylose mother liquid, an abundant byproduct from industrialscale xylose preparation from corncob and rich in d xylose, finally achieving a 35% higher A cetone, B utanol and E thanol (ABE) solvent titer (16.91 g/L) and a 38% higher yield (0.29 g/g) over those of the wildtype strain. Art. [#ARTNUM](#article-25177-1976516786)

1.3.1 1.3 ABE fermentation
Metabolic engineering of d-xylose pathway in Clostridium beijerinckii to optimize solvent production from xylose mother liquid
Abstract Clostridium beijerinckii is an attractive butanol-producing microbe for its advantage in co-fermenting hexose and pentose sugars. However, this Clostridium strain exhibits undesired efficiency in utilizing d -xylose, one of the major building blocks contained in lignocellulosic materials. Here, we reported a useful metabolic engineering strategy to improve d -xylose consumption by C. beijerinckii . Gene cbei2385, encoding a putative d -xylose repressor XylR, was first disrupted in the C. beijerinckii NCIMB 8052, resulting in a significant increase in d -xylose consumption. A d -xylose proton-symporter (encoded by gene cbei0109) was identified and then overexpressed to further optimize d -xylose utilization, yielding an engineered strain 8052xylR-xylT ptb ( xylR inactivation plus xylT overexpression driven by ptb promoter). We investigated the strain 8052xylR-xylT ptb in fermenting xylose mother liquid, an abundant by-product from industrial-scale xylose preparation from corncob and rich in d -xylose, finally achieving a 35% higher A cetone, B utanol and E thanol (ABE) solvent titer (16.91 g/L) and a 38% higher yield (0.29 g/g) over those of the wild-type strain. The strategy used in this study enables C. beijerinckii more suitable for butanol production from lignocellulosic materials.
9/1/12 12:00:00 AM
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1.3.2 1.3 ABE fermentation
Method using xylose production waste liquid to produce acetone and butanol
A method using the xylose production waste liquid to produce acetone and butanol belongs to the technical field of biochemical engineering, utilizes the xylose production waste liquid as the raw material, utilizes the microbial fermentation technology and the biotransformation sugar source to produce chemical products of acetone and butanol, and has the specific technological steps of culture medium preparation, bacterial strain activation, inoculation and fermentation, so the products of ethanol, acetone and butanol are obtained. The invention has advanced technology and a low equipment input cost; the utilization problem of the xylose production waste liquid is solved, and the additional value is high; compared with chemical synthesis, fossil resources are also saved; meanwhile, compared with the production method which takes the grain crop as the fermentation raw material, the price of the raw material is low, the method does not compete with the human being for food, and does not compete with the grain forest for land; and the comprehensive utilization of the xylose production waste liquid is realized, the comprehensive cost of the xylose production is reduced, the utilization efficiency of the biomass resource is improved, the industrial by-products are reused, the environment pollution is less, the energy consumption is low, and the social benefit is obvious.
6/11/14 12:00:00 AM
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1.3.3 1.3 ABE fermentation
Processes for biobutanol production from renewable resources
The impact of petroleum fuel emissions and more rapid diminishing petroleum reserves have increased the research for alternative biofuel sources. In this scenario, recently is rising the biorefinery concept. A biorefinery is a facility that produces fuels, power, heat, and value-added chemicals from biomass conversion. The study carried out during the present Ph.D. program aimed at investigating the butanol production process by fermentation from renewable resources. The activities, in order to pursue the biorefinery concept, were articulated according to three paths :i)Feedstock market and techno-economic feasibility assessment of butanol production; ii)Biomass Pretreatment; iii)Butanol production and characterization of the ABE fermentation process. These studies were carried out at the "Enco" Engineering Consulting Company, at the "University of Western Ontario" Canada and at the Dipartimento di Ingegneria Chimica, dei Materiali e della Produzione Industriale of the University of Naples 'Federico II'. Waste to energy: feedstocks market assessment The cost of the substrate represents about 60% of the overall production cost for a fermentation process. For this reason, feedstock available at high mass rate, with a constant availability over the year and low cost is a key issue for the success of the butanol production. A survey of potential "waste biomass" for butanol production was proposes. The maximum butanol production rate from each biomass has been estimated taking into account the feedstock availability rate, the average composition and the butanol yield. Furthermore, a study aiming at investigating the techno-economic feasibility of butanol production from lignocellulosic biomass was carried out. A potential flowsheet to produce butanol by conversion of a lignocellulosic biomass has been simulated by means of the software Aspen Plus®. The production process has been splitted into three sections: the upstream section, the fermentation section, and the butanol recovery section. Particular attention has been paid to the upstream process. The upstream units have been analysed according the approximated cost-estimation methods integrated with the simulation software Aspen Plus®. Biomass Pretreatment A new class of solvents DES (Deep Eutectic Solvent) has been investigated to obtain fermentable sugars from corncob. Corncob, a byproduct of corn grain production, is currently being used as a potential feedstock for cellulosic ethanol production in the United States, as it has a low lignin and high carbohydrate contents. DESs exhibit similar physico-chemical properties to ionic liquids, but they are environmentally friendlier and much cheaper. The pretretated corncob was characterized in term of lignin content, inhibitors concentration, crystallinity index and enzymatic digestibility. Butanol production and characterization of the ABE fermentation process The study was aimed at the assessment of the butanol production by C. acetobutylicum. Xylose and lactose are used as carbon source. Xylose is one of the mains components of the lignocellulose hydrolysates, lactose is used to mime cheese-whey, a wastewater stream released from the cheese industry. In order to optimize a continuous biofilm reactor, which is characterized by a heterogeneous cell population, the kinetics of acidogenic and solventogenic cells are investigated. Acids production by acidogenic cells and butanol production by solventogenic cells were investigated using different reactor configurations: CSTR under controlled pH and CSTR with microfiltration unit respectively. Operating conditions of the continuous tests were selected to maximize the butanol production and butanol selectivity.
4/8/15 12:00:00 AM
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1.4 2,3-butanediol

1

2,3-Butanediol (2,3-BD) is a platform chemical that can be converted into a variety of chemicals through dehydrogenation, ketalization, esterification, and dehydration. **Research Findings** - To achieve 2,3-BD production from xylose in yeast, Kim et al. constructed an engineered S. cerevisiae able to efficiently produce 2,3-BD from glucose first, and introduced the XR-XDH pathway into the 2,3-BD strain. Specifically, the 2,3-BD strain was prepared through laboratory evolution of pyruvate decarboxylase (PDC) deficient S. cerevisiae (Δpdc1, Δpdc5) with the heterologous 2,3-BD biosynthetic pathway consisting of acetolactate synthase, acetolactate decarboxylase, and butanediol dehydrogenase, during cultivations in glucose. The resulting strain, overexpressing both the 2,3-BD pathway and the XR-XDH pathway, produced 2,3-BD from xylose with 20.7 g∙L−1 titer and 0.27 g/g yield during batch culture. Additional overexpression of Tal1 enhanced the xylose consumption rate (1.47 ± 0.13 g∙L−1) and 2,3-BD productivity (0.47 ± 0.01 g∙L−1∙h−1) of the strain during xylose culture. - The ability of strain E. cloacae SG1 for utilization various pentoses and hexoses were evaluated and found that the strain can utilize both arabinose and glucose with a comparable 2,3butanediol yield. Art. [#ARTNUM](#article-25171-2282059219) - Bacillus licheniformis mutants, WX02ΔbudC and WX02ΔgldA, were elucidated for the potential to use Miscanthus as a costeffective biomass to produce optically pure 2,3BD. Both WX02ΔbudC and WX02ΔgldA could efficiently use xylose as well as mixed sugars of glucose and xylose to produce optically pure 2,3BD. Art. [#ARTNUM](#article-25171-2801861631)

1.4.1 1.4 2,3-butanediol
Evaluation of oil palm front hydrolysate as a novel substrate for 2,3-butanediol production using a novel isolate Enterobacter cloacae SG1
The present work deals the production of 2,3-butanediol, an industrially important chemical, through biological route using a novel bacterial isolate. Batch fermentation trials for the production of 2,3-butanediol were carried out using the isolated strain Enterobacter cloacae SG-1. The study resulted 14.67 g/l of 2,3-butanediol with 48.9% yield using glucose as the carbon source. In order to replace the expensive glucose in the production media, non-detoxified oil palm frond hydrolysate was used as the carbon source and it resulted 2,3-butanediol yield of 7.67 g/l. Process parameters like pH, temperature and initial sugar concentration were optimized. The ability of strain E. cloacae SG-1 for utilization various pentoses and hexoses were evaluated and found that the strain can utilize both arabinose and glucose with a comparable 2,3-butanediol yield.
12/1/16 12:00:00 AM
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1.4.2 1.4 2,3-butanediol
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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1.4.3 1.4 2,3-butanediol
Production of optically pure 2,3-butanediol from Miscanthus floridulus hydrolysate using engineered Bacillus licheniformis strains
2,3-Butanediol (2,3-BD) can be produced by fermentation of natural resources like Miscanthus. Bacillus licheniformis mutants, WX-02ΔbudC and WX-02ΔgldA, were elucidated for the potential to use Miscanthus as a cost-effective biomass to produce optically pure 2,3-BD. Both WX-02ΔbudC and WX-02ΔgldA could efficiently use xylose as well as mixed sugars of glucose and xylose to produce optically pure 2,3-BD. Batch fermentation of M. floridulus hydrolysate could produce 21.6 g/L d-2,3-BD and 23.9 g/L meso-2,3-BD in flask, and 13.8 g/L d-2,3-BD and 13.2 g/L meso-2,3-BD in bioreactor for WX-02ΔbudC and WX-02ΔgldA, respectively. Further fed-batch fermentation of hydrolysate in bioreactor showed both of two strains could produce optically pure 2,3-BD, with 32.2 g/L d-2,3-BD for WX-02ΔbudC and 48.5 g/L meso-2,3-BD for WX-02ΔgldA, respectively. Collectively, WX-02ΔbudC and WX-02ΔgldA can efficiently produce optically pure 2,3-BD with M. floridulus hydrolysate, and these two strains are candidates for industrial production of optical purity of 2,3-BD with M. floridulus hydrolysate.
5/1/18 12:00:00 AM
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1.5 1,2,4-Butanetriol

1

1,2,4-Butanetriol (BT) is a four-carbon polyol with three hydrophilic hydroxyl groups. As an important fine chemical, BT has versatile applications in many different fields and attracted considerable interest in the past few years. Nitration of racemic d,l-1,2,4-butanetriol 1 affords the energetic material d,l-1,2,4-butanetriol trinitrate 2, which is less shock sensitive, more thermally stable, and less volatile than nitroglycerin. [Wiki](https://en.wikipedia.org/wiki/1,2,4-Butanetriol) **Research Findings** - Oxidation of dxylose by Pseudomonas fragi provides dxylonic acid in 70% yield. Escherichia coli DH5α/pWN6.186A then catalyzes the conversion of dxylonic acid into d1,2,4butanetriol in 25% yield. P. fragi is also used to oxidize larabinose to a mixture of larabino1,4lactone and larabinonic acid in 54% overall yield. After hydrolysis of the lactone, larabinonic acid is converted to l1,2,4butanetriol in 35% yield using E. coli BL21(DE3)/pWN6.222A. Art. [#ARTNUM](#article-25160-2027683911) - In this study, the metabolic network of Escherichia coli was reconstructed by heterogeneously expressing a keto acid decarboxylase(mdl C) from Pseudomonas putida ATCC12633 and a Dxylose dehydrogenase(xdh) from Caulobacter crescentus CB15, and knocking out xyl A, yjh H and yag E which were the genes of xylose utilization pathway and intermediary metabolite pathway for D1,2,4butanetriol synthesis. The recombinant strain could synthesize D1,2,4butanetriol directly using Dxylose as precursor. Art. [#ARTNUM](#article-25160-2358732798) - In this study, an engineered Escherichia coli strain was constructed to produce BT from xylose, which is a major component of the lignocellulosic biomass. Through the coexpression of a xylose dehydrogenase (CCxylB) and a xylonolactonase (xylC) from Caulobacter crescentus, native E. coli xylonate dehydratase (yjhG), a 2keto acid decarboxylase from Pseudomonas putida (mdlC) and native E. coli aldehyde reductase (adhP) in E. coli BL21 star(DE3), the recombinant strain could efficiently convert xylose to BT. Art. [#ARTNUM](#article-25160-2401937566) - Here we reconstituted a cellfree system in vitro using purified enzymes to produce BT from Dxylose. The factors that influencing the efficiency of cellfree system, including enzyme concentration, reaction buffer, pH, temperature, metal ion additives and cofactors were first identified to define optimal reaction conditions and essential components for the cascade reaction. Meanwhile, a natural cofactor recycling system was found in cellfree system. Finally, we were able to convert 18 g/L xylose to 6.1 g/L BT within 40 h with a yield of 48.0%. Art. [#ARTNUM](#article-25160-2912064407)

1.5.1 1.5 1,2,4-Butanetriol
Biosynthesis of D-1,2,4-butanetriol from D-xylose by recombinant Escherichia coli
1,2,4-Butanetriol(BT) is an important organic synthetic intermediate. In this study, the metabolic network of Escherichia coli was reconstructed by heterogeneously expressing a keto acid decarboxylase(mdl C) from Pseudomonas putida ATCC12633 and a D-xylose dehydrogenase(xdh) from Caulobacter crescentus CB15, and knocking out xyl A, yjh H and yag E which were the genes of xylose utilization pathway and intermediary metabolite pathway for D-1,2,4-butanetriol synthesis. The recombinant strain could synthesize D-1,2,4-butanetriol directly using D-xylose as precursor. Culture conditions such as temperature, medium volume, p H of fermentation broth were investigated at the titer of D-1,2,4-butanetriol of 3.96 g·L-1 under suitable fermentation conditions. The relationship between glucose utilization and D-1,2,4-butanetriol synthesis was discussed. After modifying the phosphoenolpyruvate: sugar phosphotransferase system(PTS) by knocking out pts G the reconstructed E.coli could utilize glucose and xylose simultaneously, leading to a higher D-1,2,4-butanetriol productivity.
1/1/15 12:00:00 AM
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1.5.2 1.5 1,2,4-Butanetriol
Biotechnological production of 1,2,4-butanetriol: An efficient process to synthesize energetic material precursor from renewable biomass
1,2,4-Butanetriol BT) is a valuable chemical with extensive applications in many different fields. The traditional chemical routes to synthesize BT suffer from many drawbacks, e. g., harsh reaction conditions, multiple steps and poor selectivity, limiting its industrial production. In this study, an engineered Escherichia coli strain was constructed to produce BT from xylose, which is a major component of the lignocellulosic biomass. Through the coexpression of a xylose dehydrogenase (CCxylB) and a xylonolactonase (xylC) from Caulobacter crescentus, native E. coli xylonate dehydratase (yjhG), a 2-keto acid decarboxylase from Pseudomonas putida (mdlC) and native E. coli aldehyde reductase (adhP) in E. coli BL21 star(DE3), the recombinant strain could efficiently convert xylose to BT. Furthermore, the competitive pathway responsible for xylose metabolism in E. coli was blocked by disrupting two genes (xylA and EcxylB) encoding xylose isomerase and xyloluse kinase. Under fed-batch conditions, the engineered strain BL21 Delta xylAB/pE-mdlCxylBC&pA-adhPyjhG produced up to 3.92 g/L of BT from 20 g/L of xylose, corresponding to a molar yield of 27.7%. These results suggest that the engineered E. coli has a promising prospect for the large-scale production of BT.
11/1/16 12:00:00 AM
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1.5.3 1.5 1,2,4-Butanetriol
High-yield production of D-1,2,4-butanetriol from lignocellulose-derived xylose by using a synthetic enzyme cascade in a cell-free system
Abstract Approaches using metabolic engineering to produce D-1, 2, 4-butanetriol (BT) from renewable biomass in microbial systems have achieved initial success. However, due to the lack of incomplete understanding of the complex branch pathway, the efficient fermentation system for BT production was difficult to develop. Here we reconstituted a cell-free system in vitro using purified enzymes to produce BT from D-xylose. The factors that influencing the efficiency of cell-free system, including enzyme concentration, reaction buffer, pH, temperature, metal ion additives and cofactors were first identified to define optimal reaction conditions and essential components for the cascade reaction. Meanwhile, a natural cofactor recycling system was found in cell-free system. Finally, we were able to convert 18 g/L xylose to 6.1 g/L BT within 40 h with a yield of 48.0%. The feasibility of cell-free system to produce BT in corncob hydrolysates was also determined.
2/1/19 12:00:00 AM
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1.5.4 1.5 1,2,4-Butanetriol
Microbial Synthesis of the Energetic Material Precursor 1,2,4-Butanetriol
The lack of a route to precursor 1,2,4-butanetriol that is amenable to large-scale synthesis has impeded substitution of 1,2,4-butanetriol trinitrate for nitroglycerin. To identify an alternative to the current commercial synthesis of racemic d,l-1,2,4-butanetriol involving NaBH4 reduction of esterified d,l-malic acid, microbial syntheses of d- and l-1,2,4-butanetriol have been established. These microbial syntheses rely on the creation of biosynthetic pathways that do not exist in nature. Oxidation of d-xylose by Pseudomonas fragi provides d-xylonic acid in 70% yield. Escherichia coli DH5α/pWN6.186A then catalyzes the conversion of d-xylonic acid into d-1,2,4-butanetriol in 25% yield. P. fragi is also used to oxidize l-arabinose to a mixture of l-arabino-1,4-lactone and l-arabinonic acid in 54% overall yield. After hydrolysis of the lactone, l-arabinonic acid is converted to l-1,2,4-butanetriol in 35% yield using E. coli BL21(DE3)/pWN6.222A. As a catalytic route to 1,2,4-butanetriol, microbial synthesis ...
10/1/03 12:00:00 AM
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1.6 1-hexadecanol

1

Cetyl alcohol, also known as hexadecan-1-ol and palmityl alcohol, is a fatty alcohol with the formula CH₃(CH₂)₁₅OH. [Wiki](https://en.wikipedia.org/wiki/Cetyl_alcohol) **Research Findings** - Production of 1-hexadecanol from glucose in S. cerevisiae had been achieved by introducing Tyto alba fatty acyl-CoA reductase and Y. lipolytica ATP-citrate lyase and manipulating endogenous acetyl-CoA and phospholipid metabolisms. Based on this mutant strain, an engineered S. cerevisiae converting xylose into 1-hexadecanol was constructed through optimized overexpression of S. stipitis XR, XDH, and XK, followed by laboratory evolution under xylose conditions. The resulting yeast strain achieved a 1-hexadecanol titer of 0.79 ± 0.10 g∙L−1 in xylose batch and 1.2 g∙L−1 in xylose fed-batch cultures. Yields of 1-hexadecanol from the resulting S. cerevisiae strain grown on xylose were noticeably higher as compared to the case on glucose, in both batch (0.10 g/g xylose, 0.03 g/g glucose) and fed-batch (0.08 g/g xylose, <0.01 g/g glucose) cultures. Art. [#ARTNUM](#article-25188-2905366765)

1.6.1 1.6 1-hexadecanol
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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1.7 Xylitol

1

Xylitol is a sugar alcohol used as a sugar substitute. Xylitol is categorized as a polyalcohol or sugar alcohol (specifically an alditol). It has the formula CH2OH(CHOH)3CH2OH. It is a colorless or white solid that is soluble in water. [Wiki](https://en.wikipedia.org/wiki/Xylitol#Uses) Xylitol can be produced from xylose and arabinose Art. [#ARTNUM](#article-25166-1974978885) Art. [#ARTNUM](#article-25166-2120706216)

1.7.1 1.7 Xylitol
Bio-production of a Polyalcohol (Xylitol) from Lignocellulosic Resources: A Review
Lignocellulosic materials being supplied from a variety of resources at low price can be used as feedstock for chemicals and bio-products. Xylitol is a high value polyalcohol produced by the reduction of D-xylose (from hemicellulose fraction of lignocellulose) and is employed in food and pharmaceutical industries. The large number of advantageous properties, such as its low-calorie sweetening power and anticariogenicity justifies the high industrial interest for xylitol. Biotechnological production of this substance is lately becoming more attractive than the chemical method of catalytic hydrogenation due to the higher yield and because downstream processing is expected to be less costly. Studies about the bio-production of xylitol, in which microorganisms or enzymes are involved as catalysts to convert xylose into xylitol under mild conditions of pressure and temperature, have been mostly focused on establishing the operational parameters and the process options that maximize its yield and productivity in free cell system. However, some gaps in knowledge exist regarding this bioconversion process in immobilized cell system and selection or making an appropriate carrier (support) for biocatalysts in fermentation medium.
1/1/06 12:00:00 AM
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1.7.2 1.7 Xylitol
Engineering xylose and arabinose metabolism in recombinant Saccharomyces cerevisiae
Utilization of all sugars in lignocellulose hydrolysates is a prerequisite for economically feasible bioethanol production. The yeast commonly used for industrial ethanol production, Saccharomyces cerevisiae, is naturally unable to utilize pentose sugars xylose and arabinose, which constitute a large fraction of many lignocellulosic materials. Xylose utilization by S. cerevisiae can be achieved by heterologous expression of a xylose utilization pathway, consisting either of xylose reductase (XR), xylitol dehydrogenase (XDH) and xylulokinase (XK), or alternatively, of xylose isomerase (XI) and XK. Xylitol formed by XR is a major by-product in xylose fermentation when using the XR-XDH pathway. In this thesis, high-level expression of both XR and XDH was shown to decrease xylitol formation. The influence of other genetic modifications was also evaluated. It was shown that the overexpression of the non-oxidative pentose phosphate pathway (PPP) genes enables efficient growth on xylose and xylose fermentation, provided that the initial xylose pathway is expressed at a high level. When comparing the two xylose utilization pathways, higher ethanol productivity was achieved using the XR-XDH pathway, whereas higher ethanol yield was achieved with the XI pathway. The industrial xylose-fermenting S. cerevisiae strain TMB 3400, which has been previously generated by mutagenesis and selection, was tested for fermentation of lignocellulose hydrolysate. TMB 3400 displayed significantly better fermentation performance compared to the laboratory strains tested, highlighting the need for robust industrial strains in lignocellulose fermentation. TMB 3400 was also characterized by proteome analysis using difference in-gel 2-D electrophoresis. Consistently with the results obtained in other studies, increased activities of XR, XDH and a PPP enzyme TKL were found. The bacterial arabinose utilization pathway was introduced into TMB 3400, which resulted in the novel glucose, xylose and arabinose co-fermenting strain TMB 3063, with ethanol, xylitol and arabitol as the main fermentation products. (Less)
1/1/06 12:00:00 AM
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1.7.3 1.7 Xylitol
Selective reduction of xylose to xylitol from a mixture of hemicellulosic sugars.
Abstract The biocatalytic reduction of d -xylose to xylitol requires separation of the substrate from l -arabinose, another major component of hemicellulosic hydrolysate. This step is necessitated by the innate promiscuity of xylose reductases, which can efficiently reduce l -arabinose to l -arabinitol, an unwanted byproduct. Unfortunately, due to the epimeric nature of d -xylose and l -arabinose, separation can be difficult, leading to high production costs. To overcome this issue, we engineered an E. coli strain to efficiently produce xylitol from d -xylose with minimal production of l -arabinitol byproduct. By combining this strain with a previously engineered xylose reductase mutant, we were able to eliminate l -arabinitol formation and produce xylitol to near 100% purity from an equiweight mixture of d -xylose, l -arabinose, and d -glucose.
9/1/10 12:00:00 AM
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2. Biological routes to organic acids

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2.1 Glycolic acid

1

Glycolic acid (hydroacetic acid or hydroxyacetic acid); chemical formula C2H4O3 (also written as HOCH2CO2H), is the smallest α-hydroxy acid (AHA). This colorless, odorless, and hygroscopic crystalline solid is highly soluble in water. It is used in various skin-care products. [Wiki](https://en.wikipedia.org/wiki/Glycolic_acid) **Research Findings** - The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative Dxylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding Dxylose dehydrogenase (XylB) and Dxylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from Dxylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2keto3deoxyxylonic acid [(S)4,5dihydroxy2oxopentanoic acid]/ (2K3DXA) was produced and Dxylonic acid accumulated to ca. 9 g/L. A significant amount of Dxylonic acid (ca. 14%) was converted to 3deoxypentonic acid (3DPA), and also, 3,4dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldoketo reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. Art. [#ARTNUM](#article-25190-2765987778)

2.1.1 2.1 Glycolic acid
Efficient utilization of pentoses for bioproduction of the renewable two-carbon compounds ethylene glycol and glycolate.
Abstract The development of lignocellulose as a sustainable resource for the production of fuels and chemicals will rely on technology capable of converting the raw materials into useful compounds; some such transformations can be achieved by biological processes employing engineered microorganisms. Towards the goal of valorizing the hemicellulose fraction of lignocellulose, we designed and validated a set of pathways that enable efficient utilization of pentoses for the biosynthesis of notable two-carbon products. These pathways were incorporated into Escherichia coli , and engineered strains produced ethylene glycol from various pentoses, including simultaneously from D -xylose and L -arabinose; one strain achieved the greatest reported titer of ethylene glycol, 40 g/L, from D -xylose at a yield of 0.35 g/g. The strategy was then extended to another compound, glycolate. Using D -xylose as the substrate, an engineered strain produced 40 g/L glycolate at a yield of 0.63 g/g, which is the greatest reported yield to date.
3/1/16 12:00:00 AM
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2.1.2 2.1 Glycolic acid
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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2.1.3 2.1 Glycolic acid
PRODUCTION OF ACID(S) AND ALCOHOL FROM SUGARS USING YEAST
The present invention relates to method of producing glycolic acid using the Dahms pathway, as well as to a microorganism, which is able to convert D-xylose derived from biomass to 2-keto-3-deoxy pentanoic acid, 3-deoxy pentonoic acid, glycolic acid, or concomitantly to glycolic and lactic acid, and to ethylene glycol. The starting material, pentose sugar D-xylose, is a major component in lignocellulosic hydrolysates and its efficient conversion to value-added products is essential in the context of biomass utilisation and cost-effective biorefineries. Further, the present invention also relates to a glycolic acid product and to a use of said micro-organism to produce such glycolic acid.
4/7/14 12:00:00 AM
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2.1.4 2.1 Glycolic acid
Production of ethylene glycol or glycolic acid from D-xylose in Saccharomyces cerevisiae
The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative D-xylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding D-xylose dehydrogenase (XylB) and D-xylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from D-xylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2-keto-3-deoxyxylonic acid [(S)-4,5-dihydroxy-2-oxopentanoic acid] (2K3DXA) was produced and D-xylonic acid accumulated to ca. 9 g/L. A significant amount of D-xylonic acid (ca. 14%) was converted to 3-deoxypentonic acid (3DPA), and also, 3,4-dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldo-keto reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. The D-xylonate dehydratase activity in yeast was notably low, possibly due to inefficient Fe–S cluster synthesis in the yeast cytosol, and leading to D-xylonic acid accumulation. The dehydratase activity was significantly improved by targeting its expression to mitochondria or by altering the Fe–S cluster metabolism of the cells with FRA2 deletion.
11/1/17 12:00:00 AM
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2.2 3-hydroxypropionic acid

1

3-Hydroxypropionic acid (3-HP) is a non-chiral isomer of lactic acid with a β-hydroxyl group. It is a precursor for the synthesis of value-added chemicals including 1,3-propanediol, acrylic acid, and a high value-added biocompatible polymer for medical applications called poly(3-HP). **Research Findings** - We introduced the 3HP biosynthetic pathways via malonylCoA or βalanine intermediates into a xyloseconsuming yeast. Using controlled fedbatch cultivation, we obtained 7.37±0.17g 3HPL1 in 120hours with an overall yield of 29±1%Cmol 3HPCmol1 xylose. Art. [#ARTNUM](#article-25151-2219621454)

2.2.1 2.2 3-hydroxypropionic acid
A platform for Acetyl-CoA synthesis using xylose as feedstock in Saccharomyces cerevisiae
Global rise in temperature and diminishing oil reserves has stimulated a market of alternative replacements to traditional petroleum based products. An alternative is the use of a bio refinery capable of converting biomass to the normally petroleum based products. The baker yeast Saccharomyces cerevisiae is an attractive cell factory as its existing large-scale infrastructures for bioethanol production. However, it cannot utilize xylose, an otherwise unusable part of the plant biomass, which represents of utmost importance in the bio-refinery development. To also have a strain capable to produce a wide range of products, it could be used as a platform to base a bio refinery upon. Therefore, the aim of this study is to generate platform strains capable of forming acetyl-CoA, an intermediate metabolite in many of the cells metabolic reactions and also for many other industrially relevant bio-chemicals. With this goal in mind, the metabolism of S. cerevisiae was engineered. The genes encoding an isomerase-based xylose assimilation pathway (RTG, XI, XKS), and a phosphoketolase pathway (XPK, PTA), were cloned into the yeast strain CEN.PK113-5D to enable the yeast to take up and convert xylose into acetyl-CoA. The functionality of this synthetic pathway were evaluated for the production of 3-hydroxypropionic acid via introduction of ACC1** and MCR genes into the engineered strains. By characterisation of all the engineered strains on glucose growth we found increase of acetate production in strains with the phosphoketolase pathway expressed, indicating the in vivo activity of this pathway. However, expression of the xylose assimilation pathway through genome integration did not render the strains able to grow on xylose, suggesting the low efficiency of the assembled xylose assimilation pathway. To overcome this adaptive laboratory evolution is recommended.
1/1/15 12:00:00 AM
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2.2.2 2.2 3-hydroxypropionic acid
Production of 3-hydroxypropionic acid from glucose and xylose by metabolically engineered Saccharomyces cerevisiae
Biomass, the most abundant carbon source on the planet, may in the future become the primary feedstock for production of fuels and chemicals, replacing fossil feedstocks. This will, however, require development of cell factories that can convert both C6 and C5 sugars present in lignocellulosic biomass into the products of interest. We engineered Saccharomyces cerevisiae for production of 3-hydroxypropionic acid (3HP), a potential building block for acrylates, from glucose and xylose. We introduced the 3HP biosynthetic pathways via malonyl-CoA or β-alanine intermediates into a xylose-consuming yeast. Using controlled fed-batch cultivation, we obtained 7.37±0.17g 3HPL-1 in 120hours with an overall yield of 29±1%Cmol 3HPCmol-1 xylose. This study is the first demonstration of the potential of using S. cerevisiae for production of 3HP from the biomass sugar xylose.
12/1/15 12:00:00 AM
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2.2.3 2.2 3-hydroxypropionic acid
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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2.3 Butyric acid

0

Butyric acid (from Ancient Greek: βούτῡρον, meaning "butter"), also known under the systematic name butanoic acid,[5] is a carboxylic acid with the structural formula CH3CH2CH2-COOH. Salts and esters of butyric acid are known as butyrates or butanoates. Butyric acid is used in the preparation of various butyrate esters. Low-molecular-weight esters of butyric acid, such as methyl butyrate, have mostly pleasant aromas or tastes.[5] As a consequence, they are used as food and perfume additives. [Wiki](https://en.wikipedia.org/wiki/Butyric_acid#Uses) **Research Findings** - The effect of glucose and xylose on the yield of butyric acid produced by C. tyrobutyricum was investigated, separately. Cell growth of C. tyrobutyricum increased with increasing initial glucose or xylose concentration, but was not affected significantly when the concentration was above 55g /l for glucose or 35g/l for xylose. Butyric acid yield increased as the initial sugar concentration increased in both xylose and glucose, but the conversion rate from xylose or glucose to butyric acid decreased as the sugar concentration increased. The xylose to glucose ratio in their mixture did not significantly affect cell growth or butyric acid yield. Art. [#ARTNUM](#article-25159-2272686839)

2.3.1 2.3 Butyric acid
Butyric and docosahexaenoic acids production from hemicellulose
Many of the current industrial fermentation processes cannot use pentose as the carbon source. However, complete substrate utilization of sugars in lignocellulose is one of the prerequisites to render economic development of biofuels or bioproducts from biomass. In this study we proposed a new process for DHA production from renewable carbon sources by first using anaerobic bacteria, Clostridium tyrobutyricum to convert pentose into organic acids with butyric acid as the main product, and then using the organic acids to feed microaglae, Crypthecodinium cohnii to produce DHA. The effect of glucose and xylose on the yield of butyric acid produced by C. tyrobutyricum was investigated, separately. Cell growth of C. tyrobutyricum increased with increasing initial glucose or xylose concentration, but was not affected significantly when the concentration was above 55g/l for glucose or 35g/l for xylose. Butyric acid yield increased as the initial sugar concentration increased in both xylose and glucose, but the conversion rate from xylose or glucose to butyric acid decreased as the sugar concentration increased. The xylose to glucose ratio in their mixture did not significantly affect cell growth or butyric acid yield. The effect of arabinose on the yield of butyric acid produced by C. tyrobutyricum was also studied. As for butyric acid production, compared with glucose or xylose, the arabinose was in a low efficiency, with butyric acid output of 2.25g/l in 10g/l arabinose and a long lag period of about 3-4 d. However, a low concentration of arabinose could be used as a nutritional supplement to improve the fermentability of a mixture of xylose and glucose. The conversion rate of sugar to butyric acid increased as the supplement arabinose increased. In order to obtain low cost xylose, corncobs were hydrolyzed and this xylose-rich product was used to culture C. tyrobutyricum. The results showed that at end of the 9 d fermentation, the concentration of butyric acid from corncob hydrolysate reached 10.56 g/l, and the mimic medium reached 11.3 g/l. This suggests that corncob hydrolysate can be used as a carbon source for butyric acid production by C. tyrobutyricum, although some inhibitory effects were found on cell growth with corncob hydrolysate. The effect of butyric acid, lactic acid and acetic acid on the yield of DHA produced by C. cohnii was also investigated, separately. The DHA yield was highly related to both biomass and DHA content in the cell, whereas lower growth rate could bring higher DHA content. The best concentration for DHA yield seemed to be 1.2g/l in three single organic acid media. In two organic acids mixture media, acetic acid tended to be beneficial for biomass accumulation, regardless whether butyric acid or lactic acid was mixed with acetic acid, the OD could reach 1.3 or above. When butyric acid was mixed with lactic acid, the highest DHA yield was achieved, due to increased DHA content from mutual influence between butyric acid and lactic acid.
1/1/12 12:00:00 AM
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2.4 Fumaric acid

2

Fumaric acid or trans-butenedioic acid is the chemical compound with the formula HO2CCH=CHCO2H. [Wiki](https://en.wikipedia.org/wiki/Fumaric_acid) It can be a precursor for polymerization. **Research Findings** - In this study, a new strain Rhizopus arrhizus RH 7139 # was selected from the R. arrhizus RH 713 through a novel convenient and efficient selection method. Efficient production of fumaric acid (45.31 g/L) from xylose was achieved by the new strain, and the volumetric productivity was still 0.472 g/L h. Moreover, the conversion of xylose reached 73% which is close to the theoretic yield (77%). The production of fumaric acid was increased approximate by 172%, compared with the initial strain counterpart. Art. [#ARTNUM](#article-25176-2041954232) - When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Whereas the fumaric acid concentration was not significantly altered when cultivating with xylose (4.5 g/L). Art. [#ARTNUM](#article-25176-2057194747)

2.4.1 2.4 Fumaric acid
High production of fumaric acid from xylose by newly selected strain Rhizopus arrhizus RH 7-13-9#.
Abstract Fumaric acid, as an important material for polymerization, is highly expected to be produced by fermentation of lignocellulosic biomass which is composed of cellulose, hemicellulose and lignin. Xylose as the main component of hemicellulose cannot be efficiently utilized by most of the common fermentation. In this study, a new strain Rhizopus arrhizus RH 7-13-9# was selected from the R. arrhizus RH 7-13 through a novel convenient and efficient selection method. Efficient production of fumaric acid (45.31 g/L) from xylose was achieved by the new strain, and the volumetric productivity was still 0.472 g/L h. Moreover, the conversion of xylose reached 73% which is close to the theoretic yield (77%). The production of fumaric acid was increased approximate by 172%, compared with the initial strain counterpart. These results indicated that xylose, as the main component of hemicellulose, has a promising application for the production of fumaric acid on an industrial-scale.
6/1/15 12:00:00 AM
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2.4.2 2.4 Fumaric acid
Process characterization and influence of alternative carbon sources and carbon-to-nitrogen ratio on organic acid production by Aspergillus oryzae DSM1863
l-Malic acid and fumaric acid are C4 dicarboxylic organic acids and considered as promising chemical building blocks. They can be applied as food preservatives and acidulants in rust removal and as polymerization starter units. Molds of the genus Aspergillus are able to produce malic acid in large quantities from glucose and other carbon sources. In order to enhance the production potential of Aspergillus oryzae DSM 1863, production and consumption rates in an established bioreactor batch-process based on glucose were determined. At 35 °C, up to 42 g/L malic acid was produced in a 168-h batch process with fumaric acid as a by-product. In prolonged shaking flask experiments (353 h), the suitability of the alternative carbon sources xylose and glycerol at a carbon-to-nitrogen (C/N) ratio of 200:1 and the influence of different C/N ratios in glucose cultivations were tested. When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Whereas the fumaric acid concentration was not significantly altered when cultivating with xylose (4.5 g/L), it is clearly enhanced by using glycerol (9.3 g/L). When using glucose as a carbon source, an increase or decrease of the C/N ratio did not influence malic acid production but had an enormous influence on fumaric acid production. The highest fumaric acid concentrations were determined at the highest C/N ratio (300:1, 8.44 g/L) and lowest at the lowest C/N ratio (100:1, 0.7 g/L).
6/1/14 12:00:00 AM
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2.5 Lactic acid

1

Lactic acid (2-hydroxypropanoic acid) is a three‑carbon carboxylic acid used in food, cosmetic, pharmaceutical, and polymer industries. Current commercial production of lactic acid largely depends on the microbial culture because of mild operating conditions and the high chiral purity of the product. **Research Findings** - Lactobacillus brevis ATCC 367 was selected to produce lactic acid. This strain possesses a relaxed carbon catabolite repression mechanism that can use glucose and xylose simultaneously; however, lactic acid yield was only 0.52 g  g −1 from a mixture of glucose and xylose, and 5.1 g  L −1 of acetic acid and 8.3 g  L −1 of ethanol were also formed during production of lactic acid. The yield was significantly increased and ethanol production was significantly reduced if L. brevis was cocultivated with Lactobacillus plantarum ATCC 21028. L. plantarum outcompeted L. brevis in glucose consumption, meaning that L. brevis was focused on converting xylose to lactic acid and the byproduct, ethanol, was reduced due to less NADH generated in the fermentation system. Sequential cofermentation of L. brevis and L. plantarum increased lactic acid yield to 0.80 g  g −1 from poplar hydrolyzate and increased yield to 0.78  g lactic acid per g of biomass from alkalitreated corn stover with minimum byproduct formation. Art. [#ARTNUM](#article-25163-2005699602) - Production of L(+)lactic acid by R. oryzae using xylose has been reported; however, its yield and conversion rate are poor compared with that of using glucose. In this study, we report an adapted R. oryzae strain HZS6 that significantly improved efficiency of substrate utilization and enhanced production of L(+)lactic acid from corncob hydrolysate. It increased L(+)lactic acid final concentration, yield, and volumetric productivity more than twofold compared with its parental strain. Art. [#ARTNUM](#article-25163-2081748911)

2.5.1 2.5 Lactic acid
Catalytic conversion of hemicellulosic biomass to lactic acid in pH neutral aqueous phase media
Abstract The conversion of lignocellulosic biomass into value-added chemicals using non-toxic heterogeneous catalysts and water as solvent is an attractive green process. Biomass-derived lactic acid is an important renewable chemical building block for synthesizing commodity chemicals, e.g. biodegradable plastics. This paper reports that hemicellulosic biomass, xylan and xylose, can be converted to lactic acid over a ZrO 2 catalyst starting from pH neutral aqueous solutions. The effects of reaction conditions, including temperature, oxygen partial pressure, biomass loading, and catalyst loading, etc., on the conversions of hemicellulosic biomass and the corresponding yields of lactic acid have been investigated. Molar yields of lactic acid, up to 42% and 30% were produced from xylose and xylan, respectively, under the investigated reaction conditions and with the ZrO 2 catalyst. The key intermediates such as glyceraldehyde, glycolaldehyde and pyruvaldehyde were used as the reactants to probe the reaction mechanism. The role of the ZrO 2 catalyst in the retro-aldol condensation of xylose, as well as the catalyst stability, has been discussed.
1/1/15 12:00:00 AM
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2.5.2 2.5 Lactic acid
Enhanced L-(+)-Lactic Acid Production by an Adapted Strain of Rhizopus oryzae using Corncob Hydrolysate
Corncob is an economic feedstock and more than 20 million tons of corncobs are produced annually in China. Abundant xylose can be potentially converted from the large amount of hemicellulosic materials in corncobs, which makes the crop residue an attractive alternative substrate for a value-added production of a variety of bioproducts. Lactic acid can be used as a precursor for poly-lactic acid production. Although current industrial lactic acid is produced by lactic acid bacteria using enriched medium, production by Rhizopus oryzae is preferred due to its exclusive formation of the L-isomer and a simple nutrition requirement by the fungus. Production of L-(+)-lactic acid by R. oryzae using xylose has been reported; however, its yield and conversion rate are poor compared with that of using glucose. In this study, we report an adapted R. oryzae strain HZS6 that significantly improved efficiency of substrate utilization and enhanced production of L-(+)-lactic acid from corncob hydrolysate. It increased L-(+)-lactic acid final concentration, yield, and volumetric productivity more than twofold compared with its parental strain. The optimized growth and fermentation conditions for Strain HZS6 were defined.
1/1/08 12:00:00 AM
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2.5.3 2.5 Lactic acid
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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2.5.4 2.5 Lactic acid
Isolation and Characterization of Acid-Tolerant, Thermophilic Bacteria for Effective Fermentation of Biomass-Derived Sugars to Lactic Acid
Biomass-derived sugars, such as glucose, xylose, and other minor sugars, can be readily fermented to fuel ethanol and commodity chemicals by the appropriate microbes. Due to the differences in the optimum conditions for the activity of the fungal cellulases that are required for depolymerization of cellulose to fermentable sugars and the growth and fermentation characteristics of the current industrial microbes, simultaneous saccharification and fermentation (SSF) of cellulose is envisioned at conditions that are not optimal for the fungal cellulase activity, leading to a higher-than-required cost of cellulase in SSF. We have isolated bacterial strains that grew and fermented both glucose and xylose, major components of cellulose and hemicellulose, respectively, to l(+)-lactic acid at 50°C and pH 5.0, conditions that are also optimal for fungal cellulase activity. Xylose was metabolized by these new isolates through the pentose-phosphate pathway. As expected for the metabolism of xylose by the pentose-phosphate pathway, [13C]lactate accounted for more than 90% of the total 13C-labeled products from [13C]xylose. Based on fatty acid profile and 16S rRNA sequence, these isolates cluster with Bacillus coagulans, although the B. coagulans type strain, ATCC 7050, failed to utilize xylose as a carbon source. These new B. coagulans isolates have the potential to reduce the cost of SSF by minimizing the amount of fungal cellulases, a significant cost component in the use of biomass as a renewable resource, for the production of fuels and chemicals.
5/1/06 12:00:00 AM
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2.5.5 2.5 Lactic acid
Lactic acid production from biomass-derived sugars via co-fermentation of Lactobacillus brevis and Lactobacillus plantarum.
Lignocellulosic biomass is an attractive alternative resource for producing chemicals and fuels. Xylose is the dominating sugar after hydrolysis of hemicellulose in the biomass, but most microorganisms either cannot ferment xylose or have a hierarchical sugar utilization pattern in which glucose is consumed first. To overcome this barrier, Lactobacillus brevis ATCC 367 was selected to produce lactic acid. This strain possesses a relaxed carbon catabolite repression mechanism that can use glucose and xylose simultaneously; however, lactic acid yield was only 0.52 g g −1 from a mixture of glucose and xylose, and 5.1 g L −1 of acetic acid and 8.3 g L −1 of ethanol were also formed during production of lactic acid. The yield was significantly increased and ethanol production was significantly reduced if L. brevis was co-cultivated with Lactobacillus plantarum ATCC 21028. L. plantarum outcompeted L. brevis in glucose consumption, meaning that L. brevis was focused on converting xylose to lactic acid and the by-product, ethanol, was reduced due to less NADH generated in the fermentation system. Sequential co-fermentation of L. brevis and L. plantarum increased lactic acid yield to 0.80 g g −1 from poplar hydrolyzate and increased yield to 0.78 g lactic acid per g of biomass from alkali-treated corn stover with minimum by-product formation. Efficient utilization of both cellulose and hemicellulose components of the biomass will improve overall lactic acid production and enable an economical process to produce biodegradable plastics.
6/1/15 12:00:00 AM
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2.5.6 2.5 Lactic acid
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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2.5.7 2.5 Lactic acid
d-lactic acid production from renewable lignocellulosic biomass via genetically modified Lactobacillus plantarum.
d-lactic acid is of great interest because of increasing demand for biobased poly-lactic acid (PLA). Blending poly-l-lactic acid with poly-d-lactic acid greatly improves PLA's mechanical and physical properties. Corn stover and sorghum stalks treated with 1% sodium hydroxide were investigated as possible substrates for d-lactic acid production by both sequential saccharification and fermentation and simultaneous saccharification and cofermentation (SSCF). A commercial cellulase (Cellic CTec2) was used for hydrolysis of lignocellulosic biomass and an l-lactate-deficient mutant strain Lactobacillus plantarum NCIMB 8826 ldhL1 and its derivative harboring a xylose assimilation plasmid (ΔldhL1-pCU-PxylAB) were used for fermentation. The SSCF process demonstrated the advantage of avoiding feedback inhibition of released sugars from lignocellulosic biomass, thus significantly improving d-lactic acid yield and productivity. d-lactic acid (27.3 g L−1) and productivity (0.75 g L−1 h−1) was obtained from corn stover and d-lactic acid (22.0 g L−1) and productivity (0.65 g L−1 h−1) was obtained from sorghum stalks using ΔldhL1-pCU-PxylAB via the SSCF process. The recombinant strain produced a higher concentration of d-lactic acid than the mutant strain by using the xylose present in lignocellulosic biomass. Our findings demonstrate the potential of using renewable lignocellulosic biomass as an alternative to conventional feedstocks with metabolically engineered lactic acid bacteria to produce d-lactic acid. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:271–278, 2016
3/1/16 12:00:00 AM
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2.6 Malic acid

1

Malic acid is an organic compound with the molecular formula C4H6O5. It is a dicarboxylic acid that is made by all living organisms, contributes to the sour taste of fruits, and is used as a food additive. [Wiki](https://en.wikipedia.org/wiki/Malic_acid) **Research Findings** - Molds of the genus Aspergillus are able to produce malic acid in large quantities from glucose and other carbon sources. In order to enhance the production potential of Aspergillus oryzae DSM 1863, production and consumption rates in an established bioreactor batchprocess based on glucose were determined. At 35 °C, up to 42 g/L malic acid was produced in a 168h batch process with fumaric acid as a byproduct. In prolonged shaking flask experiments (353 h), the suitability of the alternative carbon sources xylose and glycerol at a carbontonitrogen (C/N) ratio of 200:1 and the influence of different C/N ratios in glucose cultivations were tested. When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Art. [#ARTNUM](#article-25195-2057194747)

2.6.1 2.6 Malic acid
Process characterization and influence of alternative carbon sources and carbon-to-nitrogen ratio on organic acid production by Aspergillus oryzae DSM1863
l-Malic acid and fumaric acid are C4 dicarboxylic organic acids and considered as promising chemical building blocks. They can be applied as food preservatives and acidulants in rust removal and as polymerization starter units. Molds of the genus Aspergillus are able to produce malic acid in large quantities from glucose and other carbon sources. In order to enhance the production potential of Aspergillus oryzae DSM 1863, production and consumption rates in an established bioreactor batch-process based on glucose were determined. At 35 °C, up to 42 g/L malic acid was produced in a 168-h batch process with fumaric acid as a by-product. In prolonged shaking flask experiments (353 h), the suitability of the alternative carbon sources xylose and glycerol at a carbon-to-nitrogen (C/N) ratio of 200:1 and the influence of different C/N ratios in glucose cultivations were tested. When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Whereas the fumaric acid concentration was not significantly altered when cultivating with xylose (4.5 g/L), it is clearly enhanced by using glycerol (9.3 g/L). When using glucose as a carbon source, an increase or decrease of the C/N ratio did not influence malic acid production but had an enormous influence on fumaric acid production. The highest fumaric acid concentrations were determined at the highest C/N ratio (300:1, 8.44 g/L) and lowest at the lowest C/N ratio (100:1, 0.7 g/L).
6/1/14 12:00:00 AM
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2.7 Succinic acid

2

Succinic acid is a dicarboxylic acid with the chemical formula (CH2)2(CO2H)2.[5] The name derives from Latin succinum, meaning amber. Succinic acid is a precursor to some polyesters and a component of some alkyd resins. [Wiki](https://en.wikipedia.org/wiki/Succinic_acid#Applications) **Research Findings** - In this study, corncob hydrolysate was used for succinic acid production. After diluted acid treatment, xylose was released from hemicellulose as the predominant monosaccharide in the hydrolysate, whereas glucose was released very little and most was retained as cellulose in the raw material. Without any detoxification, corncob hydrolysate was used directly as the carbon source in the fermentation. Actinobacillus succinogenes could utilize the sugars in the hydrolysate to produce succinic acid efficiently. Through medium optimization, yeast extract was selected as the nitrogen source and MgCO3 was used to control pH. A total of 23.64 g/l of succinic acid was produced with a yield of 0.58 g/g based on consumed sugar, indicating that the waste corncob residue can be used to produce valueadded chemicals practically. Art. [#ARTNUM](#article-25162-2029670927) - E. coli strain AFP184 was able to utilize all sugars and sugar combinations except sucrose for biomass generation and succinate production. When using xylose as a carbon source, a yield of 0.50 g g 1 was obtained. Art. [#ARTNUM](#article-25162-2031359270) - Metabolically engineered E. coli M6PM was constructed and fermentation with pure sugars revealed that it could utilize xylose and glucose efficiently. E. coli M6PM produced a final succinate concentration of 30.03 ± 0.02 g/L and a yield of 1.09 mol/mol during 72 h dualphase fermentation using elephant grass stalk hydrolysate, which resulted in 64% maximum theoretical yield of succinic acid. Art. [#ARTNUM](#article-25162-2904282411)

2.7.1 2.7 Succinic acid
Development of succinic acid production from corncob hydrolysate by Actinobacillus succinogenes
Succinic acid is one of the most important platform chemicals since it has great potential in industrial applications. In this study, corncob hydrolysate was used for succinic acid production. After diluted acid treatment, xylose was released from hemicellulose as the predominant monosaccharide in the hydrolysate, whereas glucose was released very little and most was retained as cellulose in the raw material. Without any detoxification, corncob hydrolysate was used directly as the carbon source in the fermentation. Actinobacillus succinogenes could utilize the sugars in the hydrolysate to produce succinic acid efficiently. Through medium optimization, yeast extract was selected as the nitrogen source and MgCO3 was used to control pH. A total of 23.64 g/l of succinic acid was produced with a yield of 0.58 g/g based on consumed sugar, indicating that the waste corncob residue can be used to produce value-added chemicals practically.
10/1/10 12:00:00 AM
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2.7.2 2.7 Succinic acid
Effect of Different Carbon Sources on the Production of Succinic Acid Using Metabolically Engineered Escherichia coli
Succinic acid (SA) is an important platform molecule in the synthesis of a number of commodity and specialty chemicals. In the present work, dual-phase batch fermentations with the E. coli strain AFP184 were performed using a medium suited for large-scale industrial production of SA. The ability of the strain to ferment different sugars was investigated. The sugars studied were sucrose, glucose, fructose, xylose, and equal mixtures of glucose and fructose and glucose and xylose at a total initial sugar concentration of 100 g L -1 . AFP184 was able to utilize all sugars and sugar combinations except sucrose for biomass generation and succinate production. For sucrose as a substrate no succinic acid was produced and none of the sucrose was metabolized. The succinic acid yield from glucose (0.83 g succinic acid per gram glucose consumed anaerobically) was higher than the yield from fructose (0.66 g g -1 ). When using xylose as a carbon source, a yield of 0.50 g g -1 was obtained. In the mixed-sugar fermentations no catabolite repression was detected. Mixtures of glucose and xylose resulted in higher yields (0.60 g g -1 ) than use of xylose alone. Fermenting glucose mixed with fructose gave a lower yield (0.58 g g -1 ) than fructose used as the sole carbon source. The reason is an increased pyruvate production. The pyruvate concentration decreased later in the fermentation. Final succinic acid concentrations were in the range of 25-40 g L -1 . Acetic and pyruvic acid were the only other products detected and accumulated to concentrations of 2.7-6.7 and 0-2.7 g L -1 . Production of succinic acid decreased when organic acid concentrations reached approximately 30 g L -1 . This study demonstrates that E. coli strain AFP184 is able to produce succinic acid in a low cost medium from a variety of sugars with only small amounts of byproducts formed.
4/9/07 12:00:00 AM
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2.7.3 2.7 Succinic acid
Efficient production of succinic acid from corn stalk hydrolysates by a recombinant Escherichia coli with ptsG mutation
Succinic acid is considered to be one of the key platform chemicals used in a variety of industrial applications. The exploitation of biomass to produce succinic acid requires a microbial type that can ferment the mixture of reducing sugars derived from lignocellulose. The recombinant Escherichia coli strains with homologous or cyanobacterial ppc overexpression and IdhA, pflB, ptsG mutations were constructed, and the mixed sugar fermentations were carried out with the prominent strain SD121. Then, a modeled corn stalk hydrolysates containing 30 g l(-1) glucose, 10 g l(-1) xylose and 2.5 g l(-1) arabinose was applied for succinic acid fermentation with SD121. A yield of 0.77 g succinic acid g(-1) total sugar was achieved. Fermentation of corn stalk hydrolysates with SD121 produced a final succinic acid concentration of 36.55 g l(-1) with a higher yield of 0.83 g g(-1) total sugar in anaerobic bottles. In two-stage fermentation process in bioreactor, initial aerobic growth facilitated the subsequent anaerobic succinic acid production with a final concentration of 57.81 g l(-1), and a yield of 0.87 g g(-1), total sugar. This was the first report of succinic acid production from corn stalk hydrolysates by metabolically engineered Escherichia coli. The higher succinic acid yield from corn stalk hydrolysates compared to modeled sugar mixtures, showed a great potential usage of renewable biomass as a feedstock for an economical succinic acid production using E. coli. (C) 2010 Elsevier Ltd. All rights reserved.
1/1/11 12:00:00 AM
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2.7.4 2.7 Succinic acid
Enzyme Activity Analysis and Directive Breeding of Actinobacillus succinogenes Fermenting Crop Straw Hydrolysate Containing Pentose and Hexose for Production of Succinic Acid
Objective:It is very important to obtain high yield mutant strain on the base of key enzyme activity analysis of A. succinogenes for the industrial bioconversion of succinic acid from crop straw.Methods:The pentose and hexose hydrolyzed from crop straw hydrolyzed by dilute sulfuric acid were determinated using HPLC.During the fermentation of the hydrolysate, the activities of key enzymes were detenninated and regulated.In order to decrease ethanol yield,those strains mutated by soft X-ray of synchronous radiation were screened out on the plates with high concentration of allyl alcohol.Then the alcohol dehydrogenase activity of the mutant strain was compared with that of the parent strain.Results:Determination of the hydrolysate showed that about 47 g glucose and 26 g xylose are from 200 g hydrolyzed crop straw.Pepck,Pc and Mdh are identified as the key enzymes of succinic acid metabolism and the high activity of Adh causes the cumulation of ethanol. Determination of end products from the parent strain indicated that the concentration of ethanol is the highest among those byproducts and that the yield of succinic acid and byproduct ethanol are 54.2 g/L and 8.9 g/L,respectively.Compared with the parent strain,the ethanol concentration produced by anti-allyl alcohol mutant strain S.JST01 decreases by 84%,from 8.9 to 1.4 g/L.Meanwhile the yield of succinic acid increases by 16e,from 54.2 to 63.1 g/L.Enzyme determination showed that the activity unit of alcohol dehydrogenase (Adh)decreases from 614 to 108.Conclusions:The interdiction of metabolic pathway of Adh decreases the metabolism prosuced by ethanol accordingly,thus the succinic acid flux is strengthened by the redundant carbon flux from the byproduct.Furthermore,the mutant strain S.JST01 with the end product yield of 63.1 g/L succinic acid is worth applying to industrial fermentation.
1/1/09 12:00:00 AM
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2.7.5 2.7 Succinic acid
Modular pathway engineering of Corynebacterium glutamicum to improve xylose utilization and succinate production
Abstract Xylose-negative Corynebacterium glutamicum has been engineered to utilize xylose as the sole carbon source via either the xylose isomerase (XI) pathway or the Weimberg pathway. Heterologous expression of xylose isomerase and overexpression of a gene encoding for xylulose kinase enabled efficient xylose utilization. In this study, we show that two functionally-redundant transcriptional regulators (GntR1 and GntR2) present on xylose repress the pentose phosphate pathway genes. For efficient xylose utilization, pentose phosphate pathway genes and a phosphoketolase gene were overexpressed with the XI pathway in C. glutamicum . Overexpression of the genes encoding for transaldolase (Tal), 6-phosphogluconate dehydrogenase (Gnd), or phosphoketolase (XpkA) enhanced the growth and xylose consumption rates compared to the wild-type with the XI pathway alone. However, co-expression of these genes did not have a synergetic effect on xylose utilization. For the succinate production from xylose, overexpression of the tal gene with the XI pathway in a succinate-producing strain improved xylose utilization and increased the specific succinate production rate by 2.5-fold compared to wild-type with the XI pathway alone. Thus, overexpression of the tal , gnd , or xpkA gene could be helpful for engineering C. glutamicum toward production of value-added chemicals with efficient xylose utilization.
9/1/17 12:00:00 AM
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2.7.6 2.7 Succinic acid
Processes and apparatus for producing furfural, levulinic acid, and other sugar-derived products from biomass
In some variations, the invention provides a process for producing furfural, 5-hydroxymethylfurfural, and/or levulinic acid from cellulosic biomass, comprising: fractionating the feedstock in the presence of a solvent for lignin, sulfur dioxide, and water, to produce a liquor containing hemicellulose, cellulose-rich solids, and lignin; hydrolyzing the hemicellulose contained in the liquor, to produce hemicellulosic monomers; dehydrating the hemicellulose to convert at least a portion of C5 hemicelluloses to furfural and to convert at least a portion of C6 hemicelluloses to 5-hydroxymethylfurfural; converting at least some of the 5-hydroxymethylfurfural to levulinic acid and formic acid; and recovering at least one of the furfural, the 5-hydroxymethylfurfural, or the levulinic acid. Other embodiments provide a process for dehydrating hemicellulose to convert oligomeric C5 hemicelluloses to furfural and to convert oligomeric C6 hemicelluloses to 5-hydroxymethylfurfural. The furfural may be converted to succinic acid, or to levulinic acid, for example.
11/18/13 12:00:00 AM
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2.7.7 2.7 Succinic acid
Production of Succinic Acid for Lignocellulosic Hydrolysates
The purpose of this Cooperative Research and Development Agreement (CRADA) is to add and test new metabolic activities to existing microbial catalysts for the production of succinic acid from renewables. In particular, they seek to add to the existing organism the ability to utilize xylose efficiently and simultaneously with glucose in mixtures of sugars or to add succinic acid production to another strain and to test the value of this new capability for production of succinic acid from industrial lignocellulosic hydrolyasates. The Contractors and Participant are hereinafter jointly referred to as the 'Parties'. Research to date in succinic acid fermentation, separation and genetic engineering has resulted in a potentially economical process based on the use of an Escherichia coli strain AFP111 with suitable characteristics for the production of succinic acid from glucose. Economic analysis has shown that higher value commodity chemicals can be economically produced from succinic acid based on repliminary laboratory findings and predicted catalytic parameters. The initial target markets include succinic acid itself, succinate salts, esters and other derivatives for use as deicers, solvents and acidulants. The other commodity products from the succinic acid platform include 1,4-butanediol, {gamma}-butyrolactone, 2-pyrrolidinone and N-methyl pyrrolidinone. Current economic analyses indicate that thismore » platform is competitive with existing petrochemical routes, especially for the succinic acid and derivatives. The report presents the planned CRADA objectives followed by the results. The results section has a combined biocatalysis and fermentation section and a commercialization section. This is a nonproprietary report; additional proprietary information may be made available subject to acceptance of the appropriate proprietary information agreements.« less
6/1/02 12:00:00 AM
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2.7.8 2.7 Succinic acid
Succinate Production with Metabolically Engineered Escherichia coli Using Elephant Grass Stalk (Pennisetum purpureum) Hydrolysate as Carbon Source
Succinic acid is a spectacular chemical that can be used as the precursor of various industrial products including pharmaceuticals and biochemicals. The improvement of the succinic acid market depends on strains engineering that is capable of producing succinic acid at high yield and excellent growth rate which could utilize the wide range of carbon sources such as renewable biomass. Here we use counter selection using catAsacB for pathway design and strains developments. In this investigation, metabolically engineered Escherichia coli M6PM strain was constructed for the synthesis of succinic acid using elephant grass stalk (Pennisetum purpureum) as a carbon source. Elephant grass stalk hydrolysate was prepared which comprised of 11.60 ± 0.04 g/L glucose, 27.22 ± 0.04 g/L xylose and 0.65 ± 0.04 g/L arabinose. Metabolically engineered E. coli M6PM was constructed and fermentation with pure sugars revealed that it could utilize xylose and glucose efficiently. E. coli M6PM produced a final succinate concentration of 30.03 ± 0.02 g/L and a yield of 1.09 mol/mol during 72 h dual-phase fermentation using elephant grass stalk hydrolysate, which resulted in 64% maximum theoretical yield of succinic acid. The high succinate yield from elephant grass stalk demonstrated possible application of renewable biomass as feedstock for the synthesis of succinic acid using recombinant E. coli.
12/7/18 12:00:00 AM
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2.8 Itaconic acid

2

Itaconic acid, or methylidenesuccinic acid, is an organic compound. This dicarboxylic acid is a white solid that is soluble in water, ethanol, and acetone. Historically, itaconic acid was obtained by the distillation of citric acid, but currently it is produced by fermentation. It is used in industry as a precursor of polymers used in plastics, adhesives, and coatings. [Wiki] (https://en.wikipedia.org/wiki/Itaconic_acid) **Research Findings** - One hundred A. terreus strains were evaluated for the first time for production of IA from xylose and arabinose. Twenty strains showed good production of IA from the sugars. Among these, six strains (NRRL strains 1960, 1961, 1962, 1972, 66125, and DSM 23081) were selected for further study. One of these strains NRRL 1961 produced 49.8 ± 0.3, 38.9 ± 0.8, 34.8 ± 0.9, and 33.2 ± 2.4 g IA from 80 g glucose, xylose, arabinose and their mixture (1:1:1), respectively, per L at initial pH 3.1 and 33°C. This is the first report on the production of IA from arabinose and mixed sugar of glucose, xylose, and arabinose by A. terreus. Art. [#ARTNUM](#article-25167-2609779945) - In this work, 20 A. terreus strains were evaluated for the first time for IA production from mannose and galactose in shakeflasks at initial pH of 3.1, 33 °C and 200 rpm for 7 days. Strain NRRL1971 possesses the unique ability to produce high concentrations of IA from mannose. It produced 36.4±0.2 g IA from 80 g mannose per liter with a yield of 0.46 g g1 mannose (highest titer reported so far). Art. [#ARTNUM](#article-25167-2760843080)

2.8.1 2.8 Itaconic acid
First- and second-generation biochemicals from sugars: biosynthesis of itaconic acid
Competitive prices for sugars are needed to expand the range of biochemicals that can be industrially synthesized profitably. The most commonly accepted path from these starting materials to products is the Task 42 IEA BioEnergy biorefinery classification system, which is schematically depicted in Fig. 1. The upper part of the figure shows a wide range of potential raw material sources, while the lower part shows progressive series of chemicals that can be produced from C6 or C5 sugar core backbones through chemical catalysis or microbial fermentation. Subsequent modification of these compounds through chemical catalysis gives rise to yet another group of chemicals. Figure 1 High‐level representation of pathways via the sugar platform. The aim of IEA (International Energy Agency) Bioenergy Task 42 is to initiate and actively promote information exchange on all features of biorefinery. Sugars, which serve as the core structure upon which these chemicals are built, are typically generated using starch that originates from different grains. Of the total corn produced in the world, less than 5% is used for ethanol production. Given global controversy regarding the use of food for fuel production, many biofuel companies have begun to explore the production of ethanol from lignocellulosic materials, such as straw and other agricultural waste materials. Ethanol generated from starch is known as 1G ethanol, while ethanol produced from lignocellulose is known as 2G ethanol. We have adopted the terms 1G and 2G for use with biochemicals that can be made from starch and lignocellulose, respectively. The use of lignocellulosic materials to produce biochemicals is challenging because these starting materials require intensive pretreatment (physical, chemical or biological) followed by enzymatic hydrolysis. The hydrolysis process is mediated by a set of enzymes, generically known as cellulases, which work synergistically to produce sugars. While glucose is almost the only product that results from starch hydrolyses, lignocellulose yields glucose in addition to a range of other sugars, such as xylose, arabinose, rhamnose and galactose. Regardless of whether the sugars are derived from starch or lignocellulose, fermentation of the sugars can produce downstream products than include alcohols, organic acids, alkenes, lipids and a wide range of other chemicals. This conversion can be accomplished using bacteria, fungi or yeast (genetically modified or not) using a variety of process conditions (e.g. low/high pH, aerobic/anaerobic, mesophilic/thermophilic and various nutritional regimes). The biotransformation industry modifies these and other variables to develop new processes through iterative parameter optimization with one aim: to attain the highest possible biochemical yields at rates that enable maximal recovery after downstream processing. The number of biochemicals currently produced through this manner at a commercial scale is still low. Examples of these include: ethanol, lactic acid, succinic acid, butanol, acetone, sorbitol and itaconic acid. In this issue of Microbial Biotechnology, a new pathway for production of itaconic acid is described by Geiser et al. (2015). The annual production of itaconic acid, an unsaturated dicarboxylic acid, is around 50 000 tons/year. This biochemical is used as building block for the biosynthesis of pharmaceuticals, certain resins and adhesives (Steiger et al., 2013). It should be noted that poly (itaconic acid) can be used to develop superabsorbents, anti‐scaling agents in water treatment, and can comprise components of detergents and dispersants (Klement and Buchs, 2013). Because of its industrial potential, itaconic acid was selected by US Department of Energy as one of the top 12 chemical candidates (that can be generated from biomass) to serve as a building block for the production of value added chemicals (Werpy and Petersen, 2004). The existing itaconic biosynthesis pathway was first studied in Aspergillus terreus. Early studies established that itaconic acid was produced from cis‐aconitate, a tricarboxylic acid cycle intermediate, via the action of a decarboxylase known as CadA (Okabe et al., 2009; Klement and Buchs, 2013) (Fig. 2). The microbial‐based production of itaconic acid has been reported to be as high as 45–80 g l−1 of media (Kanamasa et al., 2008; Steiger et al., 2013). In addition to Aspergillus, previous studies have shown that other fungi, such as Ustilago maydis, produce itaconic acid. Figure 2 Pathways for itaconic acid biosynthesis. cis‐Aconitate is an output chemical from the tricarboxylic acid (TCA) cycle. Blue lines represent the classic pathway described in A spergillus species (Huang et al., 2014). Red lines represent ... The new pathway revealed by Geiser et al. (2015) has been identified in U. maydis and it involves the isomerisation of cis‐ to trans‐aconitate, via a cytosolic aconitase isomerase (Adi1), followed by a decarboxylation step mediated by a novel decarboxylase (Tad1), which exhibits significant sequence similarities to lactonizing enzymes. A quick BLASTp search reveals that the isomerase described in this study and the new decarboxylase are present in a limited number of fungi with the best hit with sequences from Pseudozyma hubeiensis. The production rates reported for U. maydis are lower than those from A. terreus; however, gene regulation studies and optimization of production conditions in U. maydis are needed to reveal the biotechnological potential of the new pathway. The current limitation of the biological production of itaconic acid on an industrial scale seems to be production costs (Klement and Buchs, 2013). This limitation can be overcome through improving microbial strains, optimization of processes and the sourcing of cheaper raw materials. The newly identified pathway also provides new options for optimizing production processes, and moves us one step closer to overcoming the current affordability challenges.
1/1/16 12:00:00 AM
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2.8.2 2.8 Itaconic acid
Mannose and galactose as substrates for production of itaconic acid by Aspergillus terreus
Itaconic acid (IA), an unsaturated 5-carbon dicarboxylic acid, is a building block platform chemical that is currently produced industrially from glucose by fermentation with Aspergillus terreus. Softwood has the potential to serve as low cost source of sugars for its production. Effective utilization of all softwood derived sugars such as glucose, mannose and galactose by the fungus for production of IA will lower the cost of its production. In this work, 20 A. terreus strains were evaluated for the first time for IA production from mannose and galactose in shake-flasks at initial pH of 3.1, 33 °C and 200 rpm for 7 days. Strain NRRL1971 possesses the unique ability to produce high concentrations of IA from mannose. It produced 36.4±0.2 g IA from 80 g mannose per liter with a yield of 0.46 g g-1 mannose (highest titer reported so far). This strain has the potential to be used for IA production from softwood. The maximum (1.1±0.2 g) IA was produced by strain DSM 23081 from 80 g galactose per liter utilizing only 9.1±0.3 g. Galactose was not suitable for IA production by these strains. This is the first detailed report on the production of IA from mannose and galactose. This article is protected by copyright. All rights reserved.
12/1/17 12:00:00 AM
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2.8.3 2.8 Itaconic acid
Production of itaconic acid from pentose sugars by Aspergillus terreus
Itaconic acid (IA), an unsaturated 5-carbon dicarboxylic acid, is a building block platform chemical that is currently produced industrially from glucose by fermentation with Aspergillus terreus. However, lignocellulosic biomass has potential to serve as low-cost source of sugars for production of IA. Research needs to be performed to find a suitable A. terreus strain that can use lignocellulose-derived pentose sugars and produce IA. One hundred A. terreus strains were evaluated for the first time for production of IA from xylose and arabinose. Twenty strains showed good production of IA from the sugars. Among these, six strains (NRRL strains 1960, 1961, 1962, 1972, 66125, and DSM 23081) were selected for further study. One of these strains NRRL 1961 produced 49.8 ± 0.3, 38.9 ± 0.8, 34.8 ± 0.9, and 33.2 ± 2.4 g IA from 80 g glucose, xylose, arabinose and their mixture (1:1:1), respectively, per L at initial pH 3.1 and 33°C. This is the first report on the production of IA from arabinose and mixed sugar of glucose, xylose, and arabinose by A. terreus. The results presented in the article will be very useful in developing a process technology for production of IA from lignocellulosic feedstocks. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1059–1067, 2017
7/1/17 12:00:00 AM
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2.9 3-dehydroshikimic acid

0

3-Dehydroshikimic acid is a hydroaromatic precursor to chemicals ranging from lphenylalanine to adipic acid. The concentration and yield of 3dehydroshikimic acid microbially synthesized from various carbon sources has been examined under fedbatch fermentor conditions. Examined carbon sources included dxylose, larabinose, and dglucose. A mixture consisting of a 3:3:2 molar ratio of glucose/xylose/arabinose was also evaluated as a carbon source to model the composition of pentose streams potentially resulting from the hydrolysis of corn fiber. Escherichia coli KL3/pKL4.79B, which overexpresses feedbackinsensitive DAHP synthase, synthesizes higher concentrations and yields of 3dehydroshikimic acid when either xylose, arabinose, or the glucose/ xylose/arabinose mixture is used as a carbon source relative to when glucose alone is used as a carbon source. E. coli KL3/pKL4.124A, which overexpresses transketolase and feedbackinsensitive DAHP synthase, synthesizes higher concentrations and yields of 3dehydroshikimic acid when the glucose/xylose/arabinose mixture is used as the carbon source relative to when either xylose or glucose is used as a carbon source. Art. [#ARTNUM](#article-25222-2001007760)

2.9.1 2.9 3-dehydroshikimic acid
Microbial synthesis of 3-dehydroshikimic acid: a comparative analysis of D-xylose, L-arabinose, and D-glucose carbon sources.
3-Dehydroshikimic acid is a hydroaromatic precursor to chemicals ranging from l-phenylalanine to adipic acid. The concentration and yield of 3-dehydroshikimic acid microbially synthesized from various carbon sources has been examined under fed-batch fermentor conditions. Examined carbon sources included d-xylose, l-arabinose, and d-glucose. A mixture consisting of a 3:3:2 molar ratio of glucose/xylose/arabinose was also evaluated as a carbon source to model the composition of pentose streams potentially resulting from the hydrolysis of corn fiber. Escherichia coli KL3/pKL4.79B, which overexpresses feedback-insensitive DAHP synthase, synthesizes higher concentrations and yields of 3-dehydroshikimic acid when either xylose, arabinose, or the glucose/xylose/arabinose mixture is used as a carbon source relative to when glucose alone is used as a carbon source. E. coli KL3/pKL4.124A, which overexpresses transketolase and feedback-insensitive DAHP synthase, synthesizes higher concentrations and yields of 3-dehydroshikimic acid when the glucose/xylose/arabinose mixture is used as the carbon source relative to when either xylose or glucose is used as a carbon source. Observed high-titer, high-yielding synthesis of 3-dehydroshikimic acid from the glucose/xylose/arabinose mixture carries significant ramifications relevant to the employment of corn fiber in the microbial synthesis of value-added chemicals.
10/1/99 12:00:00 AM
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3. Other biological routes

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3.1 Biohydrogen

1

Hydrogen can be produced microbiologically. Large quantities of H2 are needed in the petroleum and chemical industries. The largest application of H2 is for the processing ("upgrading") of fossil fuels, and in the production of ammonia. The key consumers of H2 in the petrochemical plant include hydrodealkylation, hydrodesulfurization, and hydrocracking. H2 has several other important uses. H2 is used as a hydrogenating agent, particularly in increasing the level of saturation of unsaturated fats and oils (found in items such as margarine), and in the production of methanol. It is similarly the source of hydrogen in the manufacture of hydrochloric acid. H2 is also used as a reducing agent of metallic ores. [Wiki](https://en.wikipedia.org/wiki/Hydrogen#Applications) **Research findings** - This study reports that xylose could serve as the sole carbon source for a pure culture of Klebsiella oxytoca GS408 to achieve simultaneous decolorization and biohydrogen production. With 2 g liter−1 of xylose as the substrate, a maximum xylose utilization rate (URxyl) and a hydrogen molar yield (HMY) of 93.99% and 0.259 mol of H2 mol of xylose−1, respectively, were obtained. Art. [#ARTNUM](#article-25170-2593899444)

3.1.1 3.1 Biohydrogen
Anaerobic biohydrogen production from monosaccharides by a mixed microbial community culture.
Abstract Monosaccharides (e.g. glucose and fructose) are produced from the hydrolyzation of macromolecules, such as starch, cellulose, hemicellulose and lignin, which are abundant in various industrial wastewaters. The elucidation of anaerobic activated sludge microbial community utilizing monosaccharides will lay an important foundation for the industrialization of biohydrogen production. In this study, the hydrogen production by a mixed microbial culture on four monosaccharides (glucose, fructose, galactose and arabinose) was investigated in a batch cultures. The mixed microbial culture was obtained from anaerobic activated sludge in a continuous stirred-tank reactor (CSTR) after 29 days of acclimatization. The results indicated that glucose had the highest specific hydrogen production rate of 358 mL/g.g mixed liquid volatile suspended solid (MLVSS), while arabinose had the lowest hydrogen production rate of 28 mL/g.gMLVSS. Glucose also possessed the highest specific conversion rate to hydrogen of 82 mL/g glucose, while fructose had the highest specific conversion rate to liquid product of 443 mg/g fructose. Arabinose had the lowest conversion rates to both liquid products and hydrogen. Metabolic pathways and fermentation products were the major reasons for the difference in hydrogen production from these four monosaccharides. The complex fermentation pathways of arabinose reduced its hydrogen production efficiency and a long acclimation period (over 68 h) was required before the anaerobic activated sludge could effectively utilize arabinose in batch cultures.
9/1/08 12:00:00 AM
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3.1.2 3.1 Biohydrogen
Simultaneous Decolorization and Biohydrogen Production from Xylose by Klebsiella oxytoca GS-4-08 in the Presence of Azo Dyes with Sulfonate and Carboxyl Groups
Biohydrogen production from the pulp and paper effluent containing rich lignocellulosic material could be achieved by the fermentation process. Xylose, an important hemicellulose hydrolysis product, is used less efficiently as a substrate for biohydrogen production. Moreover, azo dyes are usually added to fabricate anticounterfeiting paper, which further increases the complexity of wastewater. This study reports that xylose could serve as the sole carbon source for a pure culture of Klebsiella oxytoca GS-4-08 to achieve simultaneous decolorization and biohydrogen production. With 2 g liter−1 of xylose as the substrate, a maximum xylose utilization rate (URxyl) and a hydrogen molar yield (HMY) of 93.99% and 0.259 mol of H2 mol of xylose−1, respectively, were obtained. Biohydrogen kinetics and electron equivalent (e− equiv) balance calculations indicated that methyl red (MR) penetrates and intracellularly inhibits both the pentose phosphate pathway and pyruvate fermentation pathway, while methyl orange (MO) acted independently of the glycolysis and biohydrogen pathway. The data demonstrate that biohydrogen pathways in the presence of azo dyes with sulfonate and carboxyl groups were different, but the azo dyes could be completely reduced during the biohydrogen production period in the presence of MO or MR. The feasibility of hydrogen production from industrial pulp and paper effluent by the strain if the xylose is sufficient was also proved and was not affected by toxic substances which usually exist in such wastewater, except for chlorophenol. This study offers a promising energy-recycling strategy for treating pulp and paper wastewaters, especially for those containing azo dyes.
5/15/17 12:00:00 AM
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3.1.3 3.1 Biohydrogen
Strains for xylose fermentation hydrogen production and hydrogen production method
The invention relates to strains for xylose fermentation hydrogen production and a hydrogen production method. The strains are bacillus cereus strain S1 and brevundimonas naejangsanensis strain Z1. The strains are obtained based on xylose degradation and screening and have high hydrogen production capacity. The hydrogen production method includes: using xylose as substrate and using the bacillus cereus strain S1 and the brevundimonas naejangsanensis strain Z1 to perform fermentation cultivation so as to produce hydrogen. The hydrogen production method has the advantages that the method is simple in process, high in hydrogen production efficiency and applicable to industrial production; good strain synergic effects are achieved when the strain mixture is used for fermentation hydrogen production, and high hydrogen production efficiency is achieved; strain activity and stability can be increased to a certain degree by strain immobilized fermentation, multi-batch continuous strain use can be achieved, and production cost is lowered greatly.
1/6/16 12:00:00 AM
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3.2 Polyhydroxyalkanoates

1

Polyhydroxyalkanoates or PHAs are polyesters produced in nature by numerous microorganisms, including through bacterial fermentation of sugar or lipids.[1] When produced by bacteria they serve as both a source of energy and as a carbon store. [Wiki](https://en.wikipedia.org/wiki/Polyhydroxyalkanoates) **Research Findings** - Pseudomonas cepacia was evaluated for its ability to utilize xylose, a major hemicellulosic sugar of hardwoods, for the production of the biodegradable, thermoplastic poly(Phydroxybutyrate) (PHB). This culture produced 2.6 g . LI of biomass containing 60% (w/w) PHB when grown in shake flasks on an ammoniumlimited, mineral salts medium containing 10 g. Art. [#ARTNUM](#article-25172-2461176600)

3.2.1 3.2 Polyhydroxyalkanoates
Engineering xylose metabolism for production of polyhydroxybutyrate in the non-model bacterium Burkholderia sacchari
Despite its ability to grow and produce high-value molecules using renewable carbon sources, two main factors must be improved to use Burkholderia sacchari as a chassis for bioproduction at an industrial scale: first, the lack of molecular tools to engineer this organism and second, the inherently slow growth rate and poly-3-hydroxybutyrate [P(3HB)] production using xylose. In this work, we have addressed both factors. First, we adapted a set of BglBrick plasmids and showed tunable expression in B. sacchari. Finally, we assessed growth rate and P(3HB) production through overexpression of xylose transporters, catabolic or regulatory genes. Overexpression of xylR significantly improved growth rate (55.5% improvement), polymer yield (77.27% improvement), and resulted in 71% of cell dry weight as P(3HB). These values are unprecedented for P(3HB) accumulation using xylose as a sole carbon source and highlight the importance of precise expression control for improving utilization of hemicellulosic sugars in B. sacchari.
12/1/18 12:00:00 AM
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3.2.2 3.2 Polyhydroxyalkanoates
Hemicellulose as a potential substrate for production of
Pseudomonas cepacia was evaluated for its ability to utilize xylose, a major hemicellulosic sugar of hardwoods, for the production of the biodegradable, thermoplastic poly(P-hydroxybutyrate) (PHB). This culture produced 2.6 g . L-I of biomass containing 60% (w/w) PHB when grown in shake flasks on an ammonium-limited, mineral salts medium containing 10 g . L-l of xylose. Batch fermentation data showed that growth and PHB production kinetics on xylose were similar to previously published results for the same microorganism on fructose. On xylose, the maximum specific growth rate, the maximum specific PHB production rate (based on total biomass minus PHB biomass), the overall yield of biomass produced from substrate consumed, the yield of PHB produced from substrate consumed (YPHBIS), and the percentage of PHB were 0.22 h-l, 0.072 g . g-I. h-l, 0.29 g . g-l, 0.11 g . g-I and 45% (w/w), respectively. A high maintenance energy (0.119 g of xylose . g of biomass-I. h-I) is probably responsible for the low overall yield. However, the product yield, YPHBIS, was still the highest reported for any microorganism grown on pentosic sugars. Using the YPHBIS of 0.1 1 g . g-l, it was estimated that the substrate cost (in terms of hydrolyzed hemicellulose) for PHB production would be similar to that of cane molasses and half that of bulk glucose.
1/1/95 12:00:00 AM
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3.2.3 3.2 Polyhydroxyalkanoates
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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3.2.4 3.2 Polyhydroxyalkanoates
Upgrading wheat straw to HOMO and co-polyhydroxyalkanoates
Polyhydroxyalkanoates (PHAs) are biodegradable and thus environmentally friendly thermoplastics that are synthesized by various microbial strains as intracellular storage materials. These polyesters present a broad range of properties varying from very crystalline to more elastomeric polymers and find applications from agriculture to medicine. Despite their versatility, they are still not competitive due to the high production costs, of which the C-source accounts for circa 30%. To decrease raw materials costs, lignocellulosic agro-industrial residues rich in cellulose and hemicelluloses can be used as the C-source after being processed to yield simple sugars. Wheat straw lignocellulosic hydrolysates (LCH) were prepared (biorefinery.de GmbH) by pre-treating this residual biomass using the AFEX process followed by enzymatic hydrolysis. A hydrolysate rich in glucose and xylose and with low titres of inhibitory compounds is produced that can be used as carbon source for PHA production. Burkholderia sacchari DSM 17165 was selected for its ability to use both hexoses and pentoses. Polymer production was optimized in fed-batch cultivations in stirred-tank reactors (STR). Polymer concentration, volumetric productivity and polymer cell content of respectively 84 g/L, 1.6 g L −1 h −1 and 68 % (w/w) were attained [1]. Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P3HB-co-4HB) copolymers exhibit attractive thermal and mechanical properties due to the 4HB monomer. Synthesis of this monomer was achieved upon the addition of gamma-butyrolactone (GBL) as co-substrate to fed-batch cultures. Using a DOstat feeding strategy for LCH and a continuous addition of GBL, the maximum attained P(3HB-co-4HB) productivity and 4HB molar % were 0.5 g/(L.h) and 5.0 molar %, respectively [2]. Extraction of P(3HB) from the cells usually involves the use of halogenated solvents to attain high recovery yields and purities. However, the use of these solvents causes health and environmental hazards. To lessen this drawback green solvents were tested and high recovery yields and purities were achieved. Lignocellulosic agricultural residues can thus be ugraded with high yields and productivities to value-added products using the biorefinery concept.
2/1/15 12:00:00 AM
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3.3 Terpenes

0

Terpenes, the most diverse group of natural products, have as many biochemical functionalities as their structural diversity. To overcome limitations of traditional chemical synthesis or extraction from natural sources, metabolic engineering approaches have been conducted on yeast strains carrying the MVA pathway, the eukaryotic biosynthetic pathway for terpene precursors. Astxanthin, squalene and amorphadiene have all been produced from xylose, with higher yields and productivity than glucose. **Research findings** - A native xylose-utilizing, Crabtree-positive red yeast Phaffia rhodozyma was intensively studied for bioconversion of xylose into carotenoids, which are derivatives of tetraterpenes. P. rhodozyma convert sugars into a wide variety of carotenoids mostly consisting of astaxanthin. Wildtype P. rhodozyma (NRRL Y-17268) generated 0.556 ± 0.004 mg of total carotenoids per g cell (0.439 ± 0.000 mg astaxanthin/g cell) from the hydrolysate of Eucalyptus globulus hemicellulosic fraction, which is mainly composed of xylose, supplemented with KNO3 and peptone. Through random mutagenesis using N-Methyl-N′-nitro-N-nitrosoguanidine (NTG), the yields of total carotenoid (1.62 mg/g cell on xylose, 1.69 mg/g cell on glucose) and astaxanthin (1.34 mg/g cell on xylose, 1.39 mg/g cell on glucose) were further improved. Another random mutagenesis study of P. rhodozyma using NTG described a noteworthy potential of xylose utilization in terms of carotenoid production, such as higher specific content (1.84 ± 0.32 mg/g cell on xylose, 1.24 ± 0.03 mg/g cell on glucose) and yield (0.20 ± 0.05 mg/g xylose, 0.13 ± 0.00 mg/g glucose), despite limited consumption rate and growth on xylose. Art. [#ARTNUM](#article-25189-2905366765) - this study demonstrated further in another Crabtree-positive yeast, S. cerevisiae: Overexpression of the catalytic domain of HMG-CoA reductase 1 (tHmg1), a key enzyme of the MVA pathway, in S. cerevisiae leads to accumulation of squalene, a native yeast triterpene. Art. [#ARTNUM](#article-25189-2905366765 A xylose-utilizing S. cerevisiae overexpressing tHmg1, Erg10, and amorphadiene synthase produced 6-fold more amorphadiene on xylose (66.9 ± 10.9 mg∙L−1, 1.64 ± 0.26 mg/g xylose) as compared to glucose (10.7 ± 4.4 mg∙L−1, 0.25 ± 0.10 mg/g glucose). Art. [#ARTNUM](#article-25189-2905366765)

3.3.1 3.3 Terpenes
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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3.4 Diaminopentane

0

Cadaverine is a foul-smelling diamine compound produced by the putrefaction of animal tissue. Cadaverine is a toxic[1] diamine with the formula NH2(CH2)5NH2, which is similar to putrescine. [Wiki](https://en.wikipedia.org/wiki/Cadaverine) Diaminopentane could be used as a bionylon precursor. **Research Findings** - Corynebacterium glutamicum was metabolically engineered to produce the bionylon precursor 1,5diaminopentane from the hemicellulose sugar xylose. Comparison of a basic diaminopentane producer strain on xylose and glucose feedstocks revealed a 30% reduction in diaminopentane yield and productivity on the pentose sugar. The integration of in vivo and in silico metabolic flux analysis by 13C and elementary modes identified bottlenecks in the pentose phosphate pathway and the tricarboxylic acid cycle that limited performance on xylose. By the integration of global transcriptome profiling, this could be specifically targeted to the tkt operon, genes that encode for fructose bisphosphatase (fbp) and isocitrate dehydrogenase (icd), and to genes involved in formation of lysine (lysE) and Nacetyl diaminopentane (act). This was used to create the C. glutamicum strain DAPXyl1 icdGTG Peftufbp Psodtkt Δact ΔlysE. The novel producer, designated DAPXyl2, exhibited a 54% increase in product yield to 233 mmol mol–1 and a 100% increase in productivity to 1 mmol g–1 h–1 on the xylose substrate. In a fedbatch process, the strain achieved 103 g L–1 of diaminopentane from xylose with a product yield of 32%. Art. [#ARTNUM](#article-25183-2132471240)

3.4.1 3.4 Diaminopentane
Metabolic engineering of Corynebacterium glutamicum for production of 1,5-diaminopentane from hemicellulose
In the present work, the bio-based production of 1,5-diaminopentane (cadaverine), an important building block for bio-polyamides, was extended to hemicellulose a non-food raw material. For this purpose, the metabolism of 1,5-diaminopentane-producing Corynebacterium glutamicum was engineered to the use of the C5 sugar xylose. This was realized by heterologous expression of the xylA and xylB genes from Escherichia coli, mediating the conversion of xylose into xylulose 5-phosphate (an intermediate of the pentose phosphate pathway), in a defined diaminopentane-producing C. glutamicum strain, recently obtained by systems metabolic engineering. The created mutant, C. glutamicum DAP-Xyl1, exhibited efficient production of the diamine from xylose and from mixtures of xylose and glucose. Subsequently, the novel strain was tested on industrially relevant hemicellulose fractions, mainly containing xylose and glucose as carbon source. A two-step process was developed, comprising (i) enzymatic hydrolysis of hemicellulose from dried oat spelts, and (ii) biotechnological 1,5-diaminopentane production from the obtained hydrolysates with the novel C. glutamicum strain. This now opens a future avenue towards bio-based 1,5-diaminopentane and bio-polyamides thereof from non-food raw materials.
3/1/11 12:00:00 AM
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3.4.2 3.4 Diaminopentane
Systems metabolic engineering of xylose-utilizing Corynebacterium glutamicum for production of 1,5-diaminopentane.
The sustainable production of industrial platform chemicals is one of the great challenges facing the biotechnology field. Ideally, fermentation feedstocks would rather rely on industrial waste streams than on food-based raw materials. Corynebacterium glutamicum was metabolically engineered to produce the bio-nylon precursor 1,5-diaminopentane from the hemicellulose sugar xylose. Comparison of a basic diaminopentane producer strain on xylose and glucose feedstocks revealed a 30% reduction in diaminopentane yield and productivity on the pentose sugar. The integration of in vivo and in silico metabolic flux analysis by 13C and elementary modes identified bottlenecks in the pentose phosphate pathway and the tricarboxylic acid cycle that limited performance on xylose. By the integration of global transcriptome profiling, this could be specifically targeted to the tkt operon, genes that encode for fructose bisphosphatase (fbp) and isocitrate dehydrogenase (icd), and to genes involved in formation of lysine (lysE) and N-acetyl diaminopentane (act). This was used to create the C. glutamicum strain DAP-Xyl1 icdGTG Peftufbp Psodtkt Δact ΔlysE. The novel producer, designated DAP-Xyl2, exhibited a 54% increase in product yield to 233 mmol mol–1 and a 100% increase in productivity to 1 mmol g–1 h–1 on the xylose substrate. In a fed-batch process, the strain achieved 103 g L–1 of diaminopentane from xylose with a product yield of 32%. Xylose utilization is currently one of the most relevant metabolic engineering subjects. In this regard, the current work is a milestone in industrial strain engineering of C. glutamicum. See accompanying commentary by Hiroshi Shimizu DOI: 10.1002/biot.201300097
5/1/13 12:00:00 AM
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3.5 Amino acids

2

**Research Findings** - Amino acids like l-glutamate, l-lysine, l-arginine, and l-ornithine were produced from arabinose as sole carbon source by genetically engineered C. glutamicum strains. Arabinose utilizing amino acid secreting strains were constructed by the heterologous expression of the araBAD operon from E. coli into the corresponding amino acid producing strain of C. glutamicum. l-Glutamate and l-lysine producing arabinose utilizing strains were constructed by the recombination of E. coli araBAD operon in ATCC13032 and DM1729 strains, respectively. l-Ornithine production by the recombinant C. glutamicum strain was obtained by deletion of argR for pathway de-repression and deletion of argF to block l-ornithine conversion. Art. [#ARTNUM](#article-25173-2051523778) - While xylose utilizing C. glutamicum R strains were constructed by heterologous expression of E. coli xylose isomerase gene (xylA), which converts xylose to xylulose, the second enzyme xylulokinase (xylB) is already present in C. glutamicum R. Although in their studies, they focused on organic acid production, this strain is further engineered to produce the amino acid l-alanine. l-Alanine producing C. glutamicum R strain has been created by deleting the genes associated with production of organic acids and overexpressing alanine dehydrogenase gene (alaD) from Lysinibacillus sphaericus. Thus, amino acids were successfully produced by engineered C. glutamicum by utilising pentose sugars. Art. [#ARTNUM](#article-25173-2051523778)

3.5.1 3.5 Amino acids
Corynebacterium glutamicum as a potent biocatalyst for the bioconversion of pentose sugars to value-added products
Corynebacterium glutamicum, the industrial microbe traditionally used for the production of amino acids, proved its value for the fermentative production of diverse products through genetic/metabolic engineering. A successful demonstration of the heterologous expression of arabinose and xylose utilization genes made them interesting biocatalysts for pentose fermentation, which are the main components in lignocellulosic hydrolysates. Its ability to withstand substantial amount of general growth inhibitors like furfurals, hydroxyl methyl furfurals and organic acids generated from the acid/alkali hydrolysis of lignocellulosics in growth arrested conditions and its ability to produce amino acids like glutamate and lysine in acid hydrolysates of rice straw and wheat bran, indicate the future prospective of this bacterium as a potent biocatalyst in fermentation biotechnology. However, the efforts so far on these lines have not yet been reviewed, and hence an attempt is made to look into the efficacy and prospects of C. glutamicum to utilize the normally non-fermentable pentose sugars from lignocellulosic biomass for the production of commodity chemicals.
1/1/12 12:00:00 AM
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3.6 Glucose 2,6 succinate

1

To provide a method of producing a novel sugar derivative by utilizing C6 sugars such as glucose and C5 sugars such as xylose by using microorganisms. SOLUTION: This method of producing chemical compound expressed by the following formula (1) comprises: culturing microorganisms belonging to the genus Myceligenerans in a culture medium including a utilizable carbon source; and generating the chemical compound expressed by the following formula (1) in the culture medium. **Findings** When analyzing the measurements, it was found the compound is glucose 2,6 succinate represented by the following formula (1).


4. Synthetic routes to organic acids

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4.1 Aldose oxidation by peroxide

0

Hexoses and pentoses can be nearly completely converted to **formic acid** through oxidation by hydrogen peroxide at low temperatures. Art. [#ARTNUM](#article-25213-1585797610)

4.1.1 4.1 Aldose oxidation by peroxide
Reactions of carbohydrates with hydroperoxides : Part I. Oxidation of aldoses with sodium peroxide
Abstract It was found that aqueous alkaline hydrogen peroxide solutions at 0° degrade aldohexoses almost quantitatively to 6 moles of formic acid, and aldopentoses to 5 moles. A mechanism is proposed for the stepwise degradation of aldoses to formic acid, consisting of the addition of a hydroperoxide anion to the aldehydo modification of the sugar, followed by decomposition of the adduct to formic acid and the next lower aldose. The authors found that iron salts accelerate the reaction and suggested that decomposition of the peroxide adduct may take place by a free-radical process, as well as by an ionic mechanism. Reactions of the following hexoses were measured: d -glucose, d -mannose, d -galactose, d D-allose, d -altrose, and d -talose, and the following pentoses: d -xylose, d -arabinose, d -lyxose, and d -ribose. The reaction rates for the various sugars increased in the order just cited.
2/1/73 12:00:00 AM
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4.2 Catalytic conversion to lactic acid

1

Biomassderived lactic acid is an important renewable chemical building block for synthesizing commodity chemicals, e.g. biodegradable plastics. xylose can be converted to lactic acid by subsequent retro-aldol condensation and dehydration. Art. [#ARTNUM](#article-25204-1982721169) **Research Findings** - This paper reports that hemicellulosic biomass, xylan and xylose, can be converted to lactic acid over a ZrO 2 catalyst starting from pH neutral aqueous solutions. The effects of reaction conditions, including temperature, oxygen partial pressure, biomass loading, and catalyst loading, etc., on the conversions of hemicellulosic biomass and the corresponding yields of lactic acid have been investigated. Molar yields of lactic acid, up to 42% and 30% were produced from xylose and xylan, respectively, under the investigated reaction conditions and with the ZrO 2 catalyst. Art. [#ARTNUM](#article-25204-1982721169)

4.2.1 4.2 Catalytic conversion to lactic acid
Catalytic conversion of hemicellulosic biomass to lactic acid in pH neutral aqueous phase media
Abstract The conversion of lignocellulosic biomass into value-added chemicals using non-toxic heterogeneous catalysts and water as solvent is an attractive green process. Biomass-derived lactic acid is an important renewable chemical building block for synthesizing commodity chemicals, e.g. biodegradable plastics. This paper reports that hemicellulosic biomass, xylan and xylose, can be converted to lactic acid over a ZrO 2 catalyst starting from pH neutral aqueous solutions. The effects of reaction conditions, including temperature, oxygen partial pressure, biomass loading, and catalyst loading, etc., on the conversions of hemicellulosic biomass and the corresponding yields of lactic acid have been investigated. Molar yields of lactic acid, up to 42% and 30% were produced from xylose and xylan, respectively, under the investigated reaction conditions and with the ZrO 2 catalyst. The key intermediates such as glyceraldehyde, glycolaldehyde and pyruvaldehyde were used as the reactants to probe the reaction mechanism. The role of the ZrO 2 catalyst in the retro-aldol condensation of xylose, as well as the catalyst stability, has been discussed.
1/1/15 12:00:00 AM
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4.3 Galactose hydrolysis to levulinic acid

0

Levulinic acid, or 4-oxopentanoic acid, is an organic compound with the formula CH3C(O)CH2CH2CO2H. It is classified as a keto acid. This white crystalline solid is soluble in water and polar organic solvents. It is derived from degradation of cellulose and is a potential precursor to biofuels, such as ethyl levulinate. [Wiki](https://en.wikipedia.org/wiki/Levulinic_acid) Liquidstate galactose was converted into levulinic acid via a high-temperature reaction in a batch reactor. Art. [#ARTNUM](#article-25205-2060062622)

4.3.1 4.3 Galactose hydrolysis to levulinic acid
Optimization of levulinic acid production from Gelidium amansii
The study of bioproduct production, such as bioethanol and biochemicals, from inexpensive biomass has recently attracted considerable attention. Compared to land biomass, marine biomass can be grown rapidly and is easily cultivated without the need for expensive equipment. In addition, annual CO2 absorption by marine biomass is five to seven times higher than that of wood-biomass and the carbohydrate content is higher and can easily be converted to chemicals through proper chemical processes. In the production of various biochemicals from marine biomass, levulinic acid is a highly versatile chemical with numerous industrial uses and the potential to become a commodity chemical. It can be used as a raw material for resins, plasticizers, textiles, animal feed, coatings and antifreeze. The present study was carried out to determine the optimum conditions of temperature, acid concentration and reaction time for levulinic acid production from marine biomass Gelidium amansii using two-step treatment. In the first hydrolysis step, solid-state cellulose was obtained through acid soaking and used to produce ethanol by fermentation and liquid-state galactose. In the second hydrolysis step, the liquid-state galactose was converted into levulinic acid via a high-temperature reaction in a batch reactor. The overall production conversion of G. amansii biomass to levulinic acid in the two-step acid hydrolysis was approximately 20.6%.
6/1/13 12:00:00 AM
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4.3.2 4.3 Galactose hydrolysis to levulinic acid
Production of Levulinic Acid from Gelidium amansii Using Two Step Acid Hydrolysis
현재 1,2세대 바이오매스에 비해 상대적으로 값싸며 대량생산이 가능한 3세대 바이오매스인 해조류를 이용하여 다양한 바이오화합물 생산에 관한 연구들이 주목받고 있다. 이러한 이유는 해조류는 다른 바이오매스에 비해 빨리 자라나고, 큰 장비 없이도 쉽게 수확할 수 있다는 장점뿐만 아니라 다양한 화합물로 전환할 수 있는 당이 풍부하고 공정을 통해 쉽게 전환할 수 있기 때문이다. 이러한 해조류부터 다양한 바이오화합물을 생산하는데 있어서 한 가지로 resins, plasticizers, textiles, animal feed, coatings, antifreeze의 상업화된 공정에 사용할 수 있는 레불린산(levulinic acid)이 있다. 본 연구에서는 해조류로부터 효과적으로 레불린산을 생산하는데 있어서 온도, 시간, 산의 농도의 실험조건과 2단 산 처리 공정(two step acid treatment)을 통해 생산을 최적화 하는 조건을 탐색해 보았다. 첫번째 단계로는 상대적으로 저온에서 침지 공정을 통해 고상으로는 다양한 용도로 사용될 수 있는 셀룰로오스를 회수하고, 액상으로는 갈락토오스를 회수하였다. 2번째 단계로는 고온에서 회분식 공정을 통해 갈락토오스를 레불린산으로 전환하였다. 실험 결과 2단 산 처리 공정을 통해 초기바이오매스 기준 20.6%의 레불린산 수율을 확보하였다. 【The study of bioproduct production from inexpensive biomass such as marine biomass has recently attracted considerable attention. Because, marine biomass which compared to land biomass, it can be grown rapidly and is easily cultivated without the need for expensive equipment. In addition, the carbohydrate contents are similar or higher than land biomass such as woody biomass and can be easily converted to chemicals through proper chemical processes. In the production of various biochemicals from marine biomass, levulinic acid is a highly versatile chemical with numerous industrial uses and has the potential to become a commodity chemical. It can be used as a raw material for resins, plasticizers, textiles, animal feed, coatings and antifreeze. In this study, experiments were carried out to determine the optimum conditions of temperature, acid concentration and reaction time for production of levulinic acid from marine biomass, Gelidium amansii, using two-step treatment. In the first hydrolysis step, solid-state cellulose which was used to produce ethanol by fermentation and liquid-state galactose which used to produce bioproduct such as levulinic aicd were obtained through acid soaking. In the second hydrolysis step, the liquid-state galactose was converted into levulinic acid via a high-temperature reaction in a batch reactor. As a result, the overall production yield of Gelidium amansii to levulinic acid in the two-step acid hydrolysis was approximately 20.6% on the initial biomass basis.】
8/1/13 12:00:00 AM
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4.4 Galactose oxidation to mucic acid

1

Mucic acid, C6H10O8 or HOOC-(CHOH)4-COOH (also known as galactaric or meso-galactaric acid) is an aldaric acid obtained by nitric acid oxidation of galactose or galactose-containing compounds such as lactose, dulcite, quercite, and most varieties of gum. [Wiki](https://en.wikipedia.org/wiki/Mucic_acid) - The method of the present invention can easily synthesize mucic acid in a high yield from galactose and the like under low temperature and atmospheric pressure operating conditions, can be used as an intermediate to produce bio adipic acid, the raw material of nylon 66 that is used as a material for automobile parts, and, therefore, has high industrial applicability. Art. [#ARTNUM](#article-25206-2828893247)

4.4.1 4.4 Galactose oxidation to mucic acid
Novel synthetic process of mucic acid
The present invention relates to a method of synthesizing mucic acid from galactose derived from biomass including marine resources, and more specifically, to a method of synthesizing mucic acid which utilizes galactose as a starting material and through a chemical reaction, induces an oxidation reaction to synthesize mucic acid. The method of the present invention can easily synthesize mucic acid in a high yield from galactose and the like under low temperature and atmospheric pressure operating conditions, can be used as an intermediate to produce bio adipic acid, the raw material of nylon 66 that is used as a material for automobile parts, and, therefore, has high industrial applicability.
1/25/17 12:00:00 AM
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4.5 Gas-solids heterogeneous partial oxidation

1

Heterogeneous catalysis can be done in a fluidized bed reactor, where xylose can be converted to organic acids. **Research Findings** - The current work concentrated on gas-phase catalytic heterogeneous partial oxidation. Vanadyl pyrophosphate is an active and selective oxidative catalyst for conversion of xylose to organic acids in the gas phase at reaction temperatures of about 300 °C. The operating conditions have a considerable effect on the product acid distribution and also the production rates. All the xylose reacts: maleic anhydride, acrylic acid and acrolein and carbon dioxide were the major compounds detected. Among the carboxylic acids, acrylic acid was the most desirable, but maleic acid was the most abundant. Vanadyl pyrophosphate is both active and selective for this process and in the best case, at 300 °C and 10 %vol oxygen, maleic acid, acrylic acid and acrolein yields were 25 %, 17 % and 11 %, respectively. Art. [#ARTNUM](#article-25158-1888365454)

4.5.1 4.5 Gas-solids heterogeneous partial oxidation
CONVERSION OF D-XYLOSE TO CARBOXYLIC ACIDS IN A CAPILLARY FLUIDIZED BED
La demande pour remplacer les combustibles fossiles par la biomasse renouvelable est grande. Nous recherchons des facons de convertir les glucides en produits de plus grande valeur. Peu d’etudes ont ete faites a haute temperature et presque aucune experience n’a examine l’oxydation partielle catalytique de glucides a haute temperature. Il nous faut des criteres de conception pour exploiter cette technologie de facon commerciale. Nous presentons une nouvelle methode de valoriser les sucres C5 et nous faisons une revue de l’etat actuel des reactions de sucres C5. Nous voulions developper les conditions optimales pour la production d’acides desirables et eventuellement tester de nouveaux catalyseurs pour d’autres applications. Nous avons etudie l’oxydation catalytique heterogene en phase gazeuse du xylose dans un reacteur catalytique de gaz-solide a lit fluidise capillaire a temperature relativement haute et a pression atmospherique. Nous avons oxyde le sucre dans un systeme a trois catalyseurs, soit le pyrophosphate de vanadyle, le trioxyde de molybdene-oxyde de cobalt et le molybdate de fer, pour former des acides organiques et anhydrides. Nous avons injecte une solution eau-sucre dans un reacteur a lit fluidise capillaire dont la composante principale etait un tube en quartz (ID = 8 mm) dans un four qui operait jusqu’a 1000 °C. Nous avons determine la plage de conditions operatoires avec une etude de reconnaissance et ensuite nous avons prepare un plan d’experience. Les facteurs les plus importants etaient la temperature (200–550 °C), la concentration de xylose (3 %poids, 7 %poids, 10 %poids en eau), le temps de sejour (0,1 s, 0,2 s), la pression partielle d’oxygene (0 %vol, 3 %vol, 10 %vol, 21 %vol en azote) et le catalyseur (VPP, MoO3/CoO, FeMoO). Nous avons co-alimente l’azote pour ameliorer l’atomisation. D’autres parametres qui affectent l’atomisation sont le rapport gaz-liquide (0,1–0,2 %poids), le debit de liquide (0,01–0,1 ml min-1) et le diametre de bout de la buse et du tube capillaire. Nous avions quatre categories d’atomisation : de ‘a’, ‘b’, ‘c’ et ‘d’ en ordre decroissant de performance. Nous avons utilise le type ‘a’ pour verifier l’effet des autres parametres et le type ‘b’ pour verifier l’effet de la performance d’atomisation sur le rendement. Nous avons egalement teste l’alimentation sequentielle de xylose-oxygene et l’air a une frequence de 5 min-1. Pour verifier la productivite des catalyseurs, nous avons fait des experiences de 2 h et 4 h. Le temps de sejour dans le lit catalytique etait de 0.2 s et les experiences duraient 4 h. Le gaz de fluidisation contenait 3 %vol et 10 %vol d’oxygene en azote et entrait dans le reacteur par un distributeur en verre fritte. Le debit d’entree du gaz de fluidisation variait entre 80–150 ml min-1 pour avoir une pression partielle de l’oxygene de 3 %vol et 10 %vol. Nous avons fait les experiences avec 1 g de catalyseur VPP calcine. Une pompe a seringue a alimente la solution de xylose a 0.04 ml min-1. Nous avons atomise le liquide en gouttelettes par un tube capillaire de 0.25 mm serre a l’extremite de la buse. Nous avons alimente l’azote et la solution liquide avec un rapport gaz-liquide de 0.18 % poids pour produire un jet effervescent. Les gouttelettes se sont vaporisees rapidement et le xylose a reagit pour former de l’anhydride maleique, de l’acide acrylique et de l’acroleine. Une serie de trempes ont absorbe les produits liquides (les acides) de l’effluent du reacteur en eau distillee. Nous avons echantillonne et analyse les acides accumules par HPLC hors ligne. Pour valider l’analyse par HPLC et pour identifier d’autres produits possibles, nous avons analyse les liquides par chromatographie gazeuse (GC). Les conditions d’operation ont un effet sur la distribution des produits et les taux de production. L’acide acrylique est le plus desirable et l’acide maleique est le plus abondant. Le pyrophosphate de vanadyl est actif et selectif dans ce procede. Dans le meilleur cas, a 300 °C et 10 %vol d’oxygene, les rendements de l’acide maleique, de l’acide acrylique et de l’acroleine etaient de 25 %, 17 % et 11 % respectivement. Nous avons detecte du CO2 gazeux par GCMS lors de la reaction. L’analyse thermogravimetrique des echantillons VPP a confirme qu’aucun coke ne s’est forme sur le catalyseur. L’agglomeration et la caramelisation de la poudre n’etaient problematiques que lors de reactions hors de la plage de conditions d’operation etablies plus haut. ---------- The demand for renewable biomass as a replacement for fossil fuels has never been greater. Many paths to convert carbohydrates into higher value products are under investigation. Few studies have reported data at high temperature and almost no experiments have examined high temperature catalytic partial oxidation of carbohydrates. We need generalized design criteria to exploit this technology commercially. We present a new method to valorize C5 sugars and review state of the art C5 sugar reactions. We wanted to develop optimal process conditions to produce desirable acids and eventually to test new catalysts for other applications. We studied the gas phase heterogeneous catalytic oxidation of xylose in a gas-solid catalytic capillary fluidized bed reactor at relatively high temperature and atmospheric pressure. We oxidized the sugar over three catalyst system (vanadyl pyrophosphate, molybdenum trioxide-cobalt oxide and iron molybdate) to form organic acids/anhydrides. We injected a water-sugar solution into a capillary fluidized bed reactor whose main component is a quartz tube (ID = 8 mm) in a furnace that operates at up to 1000 °C. We determine the range of possible operating conditions with a scouting study and then made an experimental design. The most important factors were temperature (200-550 °C), xylose concentration (3 %wt, 7 %wt, 10 %wt in water), residence time (0.1 s, 0.2 s), oxygen partial pressure (0 %vol, 3 %vol, 10 %vol, 21 %vol in nitrogen), and catalyst (VPP, MoO3/CoO, FeMoO). Co-feeding nitrogen improved atomization. Parameters that affected atomization are gas-to-liquid ratio (0.1-0.2 %wt), liquid flow rate (0.01-0.1 ml min-1), and nozzle tip and capillary tube diameter. We had four categories of spray performance: ‘a’, ‘b’, ‘c’ and ‘d’ in decreasing order of performance. We used a mixed design of experiments including two factors: four temperatures (300 °C, 350 °C, 400 °C, 450 °C) and two oxygen partial pressures (3 %vol, 10 %vol) with VPP catalyst. We used type ‘a’ atomization to verify the effect of other parameters and type ‘b’ to verify the effect of atomization performance on yield. We also tested sequentially feeding of xylose-oxygen followed by air at a frequency of 5 min-1. The experimental plan included 2 h and 4 h runs to test catalyst stability. Residence time inside the catalytic bed was 0.2 s and experiments lasted 4 h. The fluidization gas contained 3 %vol and 10 %vol oxygen in nitrogen and entered the reactor through a fritted glass distributor. The inlet fluidizing gas stream varied between 80-150 ml/min to meet 3 %vol and 10 %vol oxygen. We carried out most experiments with 1 g of calcined VPP catalyst. We metered the 3 %wt xylose solution at 0.04 ml min-1 with a syringe pump. We atomized the liquid into small drops through a 0.25 mm capillary tube constricted at the end nozzle. We fed nitrogen and the liquid solution with the gas-to-liquid ratio of 0.18 %wt to produce an effervescent spray. The droplets vapourized rapidly and the xylose reacted to form maleic anhydride, acrylic acid and acrolein. We absorbed the liquid products (acids) from the reactor effluent in distilled water in a series of quenches. We sampled and analyzed the accumulated acids offline with high performance liquid chromatography (HPLC). We validated the HPLC analysis with gas chromatography (GC) and tried to identify other possible products. Operating conditions have a considerable effect on product distribution and production rates. Acrylic acid was the most desirable and maleic acid the most abundant. Vanadyl pyrophosphate is both active and selective for this process and in the best case, at 300 °C and 10 %vol oxygen, maleic acid, acrylic acid and acrolein yields were 25 %, 17 % and 11 %, respectively. We also detected gaseous carbon dioxide with GCMS during the reaction. Thermogravimetric analysis for the VPP samples we withdrew at the end of the reaction confirmed that no coke formed on the catalyst. Powder agglomeration and caramelization were only problematic when the reactor operated outside the range established during the scouting experiments.
6/1/14 12:00:00 AM
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4.6 Pentose oxidation to tetronic acid

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Tetronic acid is a chemical compound, classified as a γ-lactone, with the molecular formula C4H4O3. It interconverts between keto and enol tautomers. In organic synthesis, it is used as a precursor for other substituted and ring-fused furans and butenolides.[4][5] It is also forms the structural core of a class of pesticides, known as tetronic acid insecticides, which includes spirodiclofen and spiromesifen. [Wiki](https://en.wikipedia.org/wiki/Tetronic_acid) In alkaline solution pentoses can be oxidized to tetronic acid. Art. [#ARTNUM](#article-25214-2091071099)

4.6.1 4.6 Pentose oxidation to tetronic acid
Oxidation of Pentoses in Alkaline Solution
The reaction of pentoses with oxygen in dilute aqueous potassium hydroxide at 25.00 ± 0.05° has been studied by determining the initial rate of oxygen uptake and the rate of disappearance of reducing sugar. Pentose reactivity decreased through the series: D-xylose (2.0), D-ribose (1.4), L-arabinose (1.2), D-lyxose (1.0). The products formed were identified by paper chromatography, isolation, and n.m.r. spectroscopy. In all cases less than one molar equivalent of tetronic acid is produced. The results are discussed with reference to recent studies on the behavior of sugars in alkaline solution. The reaction of pentoses with oxygen in 0.5 M sodium carbonate at 25.00 ± 0.05° was also studied. The order of pentose reactivity in this system was changed to D-ribose (5.7), D-lyxose (2.2), D-xylose (1.9), L-arabinose (1.0).
5/1/71 12:00:00 AM
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4.7 Wet oxidation

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When xylose is oxidated in a wet environment, **formic acid** is produced. Other products formed include acetic acid, methanol, acetaldehyde, acetone, and a series of hydroxylated acids. Art. [#ARTNUM](#article-25212-2057780645)

4.7.1 4.7 Wet oxidation
Wet oxidation of model carbohydrate compounds
Abstract The major product formed by wet oxidation of a series of model compounds: d -xylose, d -glucose, d -glucitol, cellulose, and dextran, was formic acid. Its yield varied according to the structure of the carbohydrate, oxygen pressure, temperature, and the presence or absence of ferric sulfate. Acetic acid was also formed; its yield was much less dependent on the structure of the carbohydrate. Other products formed include methanol, acetaldehyde, acetone, and a series of hydroxylated acids.
5/1/84 12:00:00 AM
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4.8 Dehydrogenation of sugar alcohols to lactic acid

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Sugar alcohols, such as xylitol and arabinol, which can be produced chemically or biologically from xylose and arabinose, can be oxidated to give **lactic acid**. Art. [#ARTNUM](#article-25215-2129489681)

4.8.1 4.8 Dehydrogenation of sugar alcohols to lactic acid
Selective catalytic oxidation of sugar alcohols to lactic acid
Sorbitol and xylitol obtained from biomass are considered promising potential sources of both carbon building blocks and energy. We report the efficient and selective conversion of sorbitol, xylitol and other polyols into lactic acid as the major product through homogeneous iridium-NHC catalyzed dehydrogenative processes. The proposed reaction mechanism involves base-driven hydrolysis of simple sugars which accounts for the catalyst selectivity observed. In addition, catalyst deactivation pathways are explored and rational catalyst optimization is attempted through fine tuning of the complex.
1/1/15 12:00:00 AM
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4.9 Hydrogenolysis of xylitol

1

Xylitol, a product from xylose, can be converted to **ethylene glycol** and **propylene glycol** through hydrogenolysis. Art. [#ARTNUM](#article-25216-1990883223); [#ARTNUM](#article-25216-2067211519)

4.9.1 4.9 Hydrogenolysis of xylitol
Selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol on supported Ru catalysts
The selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol was carried out on different catalysts in the presence of Ca(OH)2. The catalysts included Ru supported on activated carbon (C) and, for comparison, on metal oxides, Al2O3, TiO2, ZrO2 and Mg2AlOx as well as C-supported other noble metals, Rh, Pd and Pt, with similar particle sizes (1.6–2.0 nm). The kinetic effects of H2 pressures (0–10 MPa), temperatures (433–513 K) and solid bases including Ca(OH)2, Mg(OH)2 and CaCO3 were examined on Ru/C. Ru/C exhibited superior activities and glycol selectivities than Ru on TiO2, ZrO2, Al2O3 and Mg2AlOx, and Pt was found to be the most active metal. Such effects of the metals and supports are attributed apparently to their different dehydrogenation/hydrogenation activities and surface acid-basicities, which consequently influenced the xylitol reaction pathways. The large dependencies of the activities and selectivities on the H2 pressures, reaction temperatures, and pH values showed their effects on the relative rates for the hydrogenation and base-catalyzed reactions involved in xylitol hydrogenolysis, reflecting the bifunctional nature of the xylitol reaction pathways. These results led to the proposition that xylitol hydrogenolysis to ethylene glycol and propylene glycol apparently involves kinetically relevant dehydrogenation of xylitol to xylose on the metal surfaces, and subsequent base-catalyzed retro-aldol condensation of xylose to form glycolaldehyde and glyceraldehyde, followed by direct glycolaldehyde hydrogenation to ethylene glycol and by sequential glyceraldehyde dehydration and hydrogenation to propylene glycol. Clearly, the relative rates between the hydrogenation of the aldehyde intermediates and their competitive reactions with the bases dictate the selectivities to the two glycols. This study provides directions towards efficient synthesis of the two glycols from not only xylitol, but also other lignocellulose-derived polyols, which can be achieved, for example, by optimizing the reaction parameters, as already shown by the observed effects of the catalysts, pH values, and H2 pressures.
1/1/11 12:00:00 AM
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4.9.2 4.9 Hydrogenolysis of xylitol
Selective hydrogenolysis of xylitol to ethylene glycol and propylene glycol over copper catalysts
Abstract Cu SiO 2 catalysts were prepared by homogeneous deposition–precipitation with a wide range of Cu contents (8.8–100 wt%) and Cu particle sizes (2.1–111.1 nm). These catalysts were evaluated in the selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol under the promotion of Ca(OH) 2 base. Their catalytic activity and selectivity to the two glycols depended strongly on the Cu particle sizes, which increased with the particle sizes and reached the maximum values at around 20–35 nm. Such size effects are apparently attributed to the effects on the dehydrogenation and hydrogenation activities of the Cu catalysts, and consequently on the xylitol hydrogenolysis pathways, reflecting the structural requirement for the xylitol hydrogenolysis. The effects of the reaction parameters including H 2 pressure (0–8.0 MPa), temperature (433–493 K) and pH values (7.0–12.4, adjusted by changing the amount of Ca(OH) 2 ) were examined. These effects confirmed the reaction pathways previously proposed for the xylitol hydrogenolysis to the two glycols, involving the dehydrogenation of xylitol to xylose on Cu as the rate-determining step, followed by the retro-aldol condensation of xylose with Ca(OH) 2 to glycolaldehyde and glyceraldehyde, and their subsequent hydrogenation to ultimately form glycols in competition with their side reactions to glycolic acid and lactic acid in the presence of Ca(OH) 2 . Upon optimizing the reaction conditions (473 K, 6.0 MPa H 2 and sufficient Ca(OH) 2 ), nearly 100% xylitol conversion and 54.4% combined selectivity to ethylene glycol and propylene glycol were obtained on Cu SiO 2 with Cu size of 35.7 nm, comparable to those on the previously reported Ni- and Ru- based catalysts. Clearly, this study provides directions for the design of more efficient Cu catalysts and the optimization of the reaction parameters toward the efficient polyol hydrogenolysis into glycols.
4/1/14 12:00:00 AM
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4.10 Methyl levulinate (highly recalcitrant cellulose and part hemicellulose)

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**Research findings** - To overcome these problems, our study focuses on directional microwave-assisted liquefaction of lignocellulosic biomass into MLA by an acid-catalyzed reaction in subcritical methanol. In this paper, we studied the conversion of lignocellulosic biomass with highly recalcitrant cellulose and part hemicellulose to products with microwave irradiation and alcohol. Art. [#ARTNUM](#article-30818-2788839178)

4.10.1 4.10 Methyl levulinate (highly recalcitrant cellulose and part hemicellulose)
Efficient Synergetic Combination of H-USY and SnO2 for Direct Conversion of Glucose into Ethyl Levulinate (Biofuel Additive)
Ethyl levulinate (EL), a biofuel additive for petroleum and biodiesel can also be used as a 100% fuel to replace petroleum diesel with the existing diesel engine. The major problem to make the EL process economical is the lack of a proper conversion technology to convert C6 sugars such as glucose with higher yield of EL as well as process which can tolerate higher glucose concentration to increase productivity. The present study highlighted the catalytic synthesis of EL from glucose over synergetic combination of zeolite H-USY and Lewis acidic catalysts such as Sn-beta, TiO2, ZrO2, and SnO2. Because of the strong Lewis acidic nature and the subsequent enhancement in the isomerization rate from glucosides to fructosides, the synergetic combination of H-USY with SnO2 showed higher EL yield than the combination with other Lewis acidic catalysts. So far, the highest EL yield of 81% from glucose (50 g/L) at 180 °C in 3 h was achieved over the optimal combination of 95% H-USY and 5% SnO2 having strong/weak acid...
2/25/19 12:00:00 AM
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4.10.2 4.10 Methyl levulinate (highly recalcitrant cellulose and part hemicellulose)
Selective catalytic conversion of waste lignocellulosic biomass for renewable value-added chemicals via directional microwave-assisted liquefaction
Selective catalytic conversion of biomass waste for producing methyl levulinate (MLA) via directional microwave-assisted liquefaction was investigated. The goal of the study was to develop a directional liquefaction process using dielectric heating with microwave energy. The methanolysis of biomass into methyl levulinate was studied in the presence of several acid catalysts. The C6 sugar substrates in biomass were successfully converted into methyl levulinate under the optimized conditions (180 °C, 40 min) with a yield of 29.39 wt%. 5-Hydroxymethyl furfural, glucose, fructose, cellobiose, corn starch, and microcrystalline cellulose were selected as models for directional microwave-assisted liquefaction. Therefore, the possible reaction pathway of biomass to methyl levulinate could be investigated. The selective catalytic conversion of biomass was found to be highly efficient for the generation of MLA (reaching a maximum yield of approximately 30 wt%), higher than the levulinic acid yield (14 wt%) in aqueous solution under the same reaction conditions. The results suggested that directional microwave-assisted liquefaction is an effective method that can produce a high value-added fuel additive (methyl levulinate) from lignocellulosic biomass under designated reaction processes.
1/1/18 12:00:00 AM
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4.11 Acid catalysed levulinic acid/ester production

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Direct conversion of biomassderived xylose and furfural into levulinic acid, a platform molecule, via acidcatalysis has been accomplished for the first time in dimethoxymethane/methanol. Dimethoxymethane acted as an electrophile to transform furfural into 5hydroxymethylfurfural (HMF). Methanol suppressed both the polymerisation of the sugars/furans and the Aldol condensation of levulinic acid/ester. Art. [#ARTNUM](#article-31252-2587934676)

4.11.1 4.11 Acid catalysed levulinic acid/ester production
One-pot conversion of biomass-derived xylose and furfural into levulinate esters via acid catalysis
Direct conversion of biomass-derived xylose and furfural into levulinic acid, a platform molecule, via acid-catalysis has been accomplished for the first time in dimethoxymethane/methanol. Dimethoxymethane acted as an electrophile to transform furfural into 5-hydroxymethylfurfural (HMF). Methanol suppressed both the polymerisation of the sugars/furans and the Aldol condensation of levulinic acid/ester.
1/1/17 12:00:00 AM
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5. Synthetic routes to alcohols

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5.1 Xylose catalytic dehydrogenation

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The present invention discloses a method for preparing **1,2-pentanediol** by using xylose as a raw material through catalytic hydrogenation. According to the method, a supported Rh or Pd based hydrogenation catalyst is used, water is adopted as a solvent, and xylose is converted at a temperature of 80-180 DEG C in a 0.5-10 MPa hydrogen atmosphere to generate 1,2-pentanediol, wherein xylose is a biomass resource with the wide source. According to the present invention, the xylose is adopted as the raw material so as to well solve the problem of the difficult source of the C5 component; and the used xylose aqueous solution can be the hydrolyzate of corn cobs, bagasse, cotton seed hulls and the like, such that the production cost can be further reduced, and the maximum yield can achieve 46%. Art. [#ARTNUM](#article-30807-2871865350)

5.1.1 5.1 Xylose catalytic dehydrogenation
Method for preparing 1,2-pentanediol from xylose
The present invention discloses a method for preparing 1,2-pentanediol by using xylose as a raw material through catalytic hydrogenation. According to the method, a supported Rh or Pd based hydrogenation catalyst is used, water is adopted as a solvent, and xylose is converted at a temperature of 80-180 DEG C in a 0.5-10 MPa hydrogen atmosphere to generate 1,2-pentanediol, wherein xylose is a biomass resource with the wide source. According to the present invention, the xylose is adopted as the raw material so as to well solve the problem of the difficult source of the C5 component; and the used xylose aqueous solution can be the hydrolyzate of corn cobs, bagasse, cotton seed hulls and the like, such that the production cost can be further reduced, and the maximum yield can achieve 46%.
6/1/16 12:00:00 AM
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5.2 From furfural: 1,5-pentanediol

1

Furfural can be produced from xylose through dehydrogenation as described in [Tech 6.1](#technology-25200) A new process for the production of 1,5-pentanediol (1,5-PDO) from biomass-derived furfural is studied. In this process, furfural is converted to 1,5-PDO in a high overall yield (80%) over inexpensive catalysts via multiple steps involving hydrogenation, dehydration, hydration, and hydrogenation subsequently. Art. [#ARTNUM](#article-30811-2768416581)

5.2.1 5.2 From furfural: 1,5-pentanediol
Conversion of Furfural to 1,5-Pentanediol: Process Synthesis and Analysis
A new process for the production of 1,5-pentanediol (1,5-PDO) from biomass-derived furfural is studied. In this process, furfural is converted to 1,5-PDO in a high overall yield (80%) over inexpensive catalysts via multiple steps involving hydrogenation, dehydration, hydration, and hydrogenation subsequently. To effectively recycle H2 as well as recover 1,5-PDO, detailed separation subsystems have been designed and integrated with reaction subsystems. Furthermore, a pioneer plant analysis is performed to estimate the risk on the cost growth and plant performance shortfalls. The integrated process leads to a minimum selling price of $1973 ton–1 for 1,5-PDO, which suggests that it could be a promising approach for converting biomass into oxygenated commodity chemicals, which are difficult to produce from petroleum-derived feedstocks. The sensitivity analysis also identifies that the most important economic parameters for the process include the furfural feedstock price and plant size.
6/5/17 12:00:00 AM
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5.2.2 5.2 From furfural: 1,5-pentanediol
Improving economics of lignocellulosic biofuels: An integrated strategy for coproducing 1,5-pentanediol and ethanol
Abstract A biorefinery strategy for the coproduction of ethanol and 1,5-pentanediol (1,5-PDO), which can be used as polyester and polyurethane component, from lignocellulosic biomass is proposed. This strategy integrates biomass fractionation with simultaneous conversion of hemicellulose and cellulose constituents into 1,5-PDO and ethanol, respectively. An experimentally-based process model is developed to determine the economic potential of the integrated strategy. The coproduction strategy becomes competitive with the ethanol-only strategy when 1,5-PDO can be sold at $1140/ton, which is well below the market price of 1,5-PDO. The most important process parameters include biomass loading for biomass fractionation, enzyme loading for enzymatic hydrolysis and fermentation, and overall achievable yields from C5 sugars to 1,5-PDO.
11/1/17 12:00:00 AM
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5.2.3 5.2 From furfural: 1,5-pentanediol
New catalytic strategies for α,ω-diols production from lignocellulosic biomass
Catalytic strategies for the synthesis of 1,5-pentanediol (PDO) with 69% yield from hemicellulose and the synthesis of 1,6-hexanediol (HDO) with 28% yield from cellulose are presented. Fractionation of lignocellulosic biomass (white birch wood chips) in gamma-valerolactone (GVL)/H2O generates a pure cellulose solid and a liquid stream containing hemicellulose and lignin, which is further dehydrated to furfural with 85% yield. Furfural is converted to PDO with sequential dehydration, hydration, ring-opening tautomerization, and hydrogenation reactions. Acid-catalyzed cellulose dehydration in tetrahydrofuran (THF)/H2O produces a mixture of levoglucosenone (LGO) and 5-hydroxymethylfurfural (HMF), which are converted with hydrogen to tetrahydrofuran-dimethanol (THFDM). HDO is then obtained from hydrogenolysis of THFDM. Techno-economic analysis demonstrates that this approach can produce HDO and PDO at a minimum selling price of $4090 per ton.
1/1/17 12:00:00 AM
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5.3 Catalyst single step conversion of C5 and C6 sugars to ethylene glycol, propylene glycol & glycerol

0

**Patent findings** - Also disclosed herein is a multifunctional catalyst capable of, in a single step, converting C5 and C6 sugars to ethylene glycol, propylene glycol, and glycerol. In one aspect, the, C5 and C6 sugars are hemicellulose or cellulose derived C5 and C6 sugars from biomass. [Link](#internalLink-2264)

5.3.1 5.3 Catalyst single step conversion of C5 and C6 sugars to ethylene glycol, propylene glycol & glycerol
Propylene Glycol: An Industrially Important C3 Platform Chemical
Global warming and the depletion of fossil fuels have created major pressure for scientists to seek alternative sources of energy and organic carbon. Biomass is a potential source from which many platform molecules can be derived. The development of sustainable technologies for producing these platform chemicals from renewable resources is very challenging. Propylene glycol belongs to the bulk chemicals that are produced from biomass. The biotechnological production of propylene glycol has received a lot of attention in recent years. This chapter aims to provide a general overview of propylene glycol and its commercial uses. Different technologies and methods of production are also presented.
1/1/16 12:00:00 AM
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5.3.2 5.3 Catalyst single step conversion of C5 and C6 sugars to ethylene glycol, propylene glycol & glycerol
Compositions and methods related to the production of acrylonitrile

1. A method comprising the step of:a) separating at least a portion of ethylene glycol and propylene glycol from a first product comprising ethylene glycol, propylene glycol, and glycerol, thereby producing a second product comprising glycerol; andb) contacting the second product comprising glycerol with a first catalyst composition, thereby producing a third product comprising acrolein and hydroxyacetone,wherein the first catalyst composition comprises a first catalyst comprising the formula: M2xM3yOz wherein M2 is a metal with acid sites promoting dehydration,wherein M3 is an amphoteric catalyst support, with acid and base sites,wherein x is a molar ratio from about 0.25 to about 4,wherein y is a molar ratio from about 0.25 to about 4,wherein z is the total amount of oxygen bound to M2, and M3, and corresponds to the sum of the oxidation states of M2, and M3. 2. The method of claim 1, wherein the method further comprises the steps of:c) separating at least a portion of the hydroxyacetone from the third product, thereby forming a fourth product comprising acrolein; andd) following step c) converting at least a portion of the acrolein in the fourth product to acrylonitrile. 3. The method of claim 1, wherein the method further comprises prior to step a), in a single step, converting C5 and/or C6 sugars to the first product comprising ethylene glycol, propylene glycol, and glycerol in the presence of a multifunctional catalyst. 4. The method of claim 1, wherein the method further comprises contacting at least a portion of the separated propylene glycol with a second catalyst composition, thereby producing propanal, wherein the second catalyst composition comprises a second catalyst having the formula: M4M5aM6bOz wherein M4 is a metal with acid sites promoting dehydration,wherein M5 is an amphoteric catalyst support, with acid and base sites, promoting selective dehydration in conjunction with M6 when present,wherein M6 is a metal promoting C--O cleavage,wherein a is a molar ratio from about 0.25 to about 4,wherein b is a molar ratio from 0 to about 4,wherein z is the total amount of oxygen bound to M4, M5, and M6, and corresponds to the sum of the oxidation states of M4, M5, and M6. 5. The method of claim 1, wherein M2 is selected from the group consisting of W, Fe, P, and, a zeolite. 6. The method of claim 1, wherein M3 is selected from the group consisting of Zr, Al, Si, Mg, Ti, La, and Ce. 7. The method of claim 1, wherein M2 is W. 8. The method of claim 1, wherein M3 is Zr. 9. The method of claim 1, wherein the first catalyst has the formula WO3ZrO2 or WO3SiO2. 10. The method of claim 3, wherein the C5 and/or C6 sugars is C5 and/or C6 hemicellulose and cellulose derived sugars. 11. The method of claim 3, wherein the multifunctional catalyst comprises one or more metals selected from the group consisting of Cu, Zn, Sn, Ni, Pt, Pd, Ru, and Re, and a support. 12. The method of claim 11, wherein the support is selected from the group consisting of Al2O3, SiO2, carbon, TiO2, and MgO. 13. The method of claim 1, wherein at least 60 wt % of the propylene glycol is separated from the first product. 14. The method of claim 1, wherein the third product comprises at least 50 wt % of acrolein. 15. The method of claim 1, wherein the second product comprises at least 2 times more glycerol than propylene glycol by weight. 16. The method of claim 1, wherein the first catalyst further comprises M1, wherein M1 is a metal promoting C--O cleavage,wherein M2 and M3 promotes selective dehydration in conjunction with M1,wherein z is the total amount of oxygen bound to M1, M2, and M3, and corresponds to the sum of the oxidation states of M1, M2, and M3. 17. The method of claim 16, wherein M1 is selected from the group consisting of Cu, Zn, and Sn. 18. The method of claim 16, wherein M1 is Cu. 19. The method of claim 4, whereinM4 is selected from the group consisting of W, Fe, P, and a zeolite,M5 is selected from the group consisting of Zr, Al, Si, Mg, Ti, La, and Ce, andM6 is selected from the group consisting of Cu, Zn, and Sn. 20. The method of claim 19, wherein the first catalyst has the formula WO3ZrO2 or WO3SiO2.
8/24/16 12:00:00 AM
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6. Synthetic routes to furfural-like compounds

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6.1 Xylose dehydration to furfural

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Xylose can be dehydrated to give furfural in different ways. Furfural is a versatile biomassderived platform compound used for the synthesis of several strategic chemicals. **Research findings:** - This study mainly focused on the dehydration of xylose to furfural with green catalysis. The reaction was under atmospheric pressure, with the 2 2 4 / ZrO SO − as the catalyst plus an inorganic salt (NaCl) as promoter. The results showed that the optimal catalytic reaction conditions were the following conditions: 30 mL of DMSO and 2g xylose, 1 g NaCl, 2.5g 2 2 4 / ZrO SO − and heating for 4 h. It resulted in the maximum yield of 47.7 %. Art. [#ARTNUM](#article-25200-2038521783) - The sonochemically synthesized Zn doped CuO nanoparticles (NPs) were used for the production of furfural. The catalytic activity of the Zn doped CuO NPs was examined, as a model, during the dehydration reaction of xylose to furfural. In addition to that, we have also compared the catalytic activity of the Zn doped CuO NP with ZnO NPs, ZnO bulk, CuO NPs, CuO bulk, etc. This nanoscale catalyst (Zn doped CuO NP) has a large surface area, which enhances its catalytic activity and enables it to completely convert the xylose to furfural at 150 °C within 12 h without any trace of byproducts, as confirmed by HPLC, 13 C NMR and 1 H NMR. HPLC analysis demonstrated that the yield of furfural is up to 86 mol %, compared to the 45 mol % obtained with ZnO NPs, ZnO bulk, CuO NPs, CuO bulk, etc. as catalysts.[#ARTNUM](#article-25200-2922412353) - The C5 xylose sugar syrup can be used to produce additives for animal feeding, furfurol, furfurylic alcohol Art. [#ARTNUM](#article-25200-2243839604) **Notes:** - Furfural can be used as a builiding block for several alcohols/acids.

6.1.1 6.1 Xylose dehydration to furfural
Conversion of Xylose Solution into Furfural at Atmospheric Pressure over the Solid Acid
This study mainly focused on the dehydration of xylose to furfural with green catalysis. The reaction was under atmospheric pressure, with the 2 2 4 / ZrO SO − as the catalyst plus an inorganic salt (NaCl) as promoter. DMSO was used as a renewable extraction solvent in a biphasic system. The optimal catalytic reaction conditions were determined by single factor design. The results showed that the optimal catalytic reaction conditions were the following conditions: 30 mL of DMSO and 2g xylose, 1 g NaCl, 2.5g 2 2 4 / ZrO SO − and heating for 4 h. It resulted in the maximum yield of 47.7 %. The experimental results showed that the preparation of furfural from xylose solution system with the solid acid catalyst, has the huge development potential. The addition of an inorganic salt (NaCl) to the aqueous phase was shown to improve the yield of furfural. Introduction Rapidly growing worldwide energy demand has triggered a renewed interest in producing fuels from biomass to add to worldwide energy supplies[1].Because of the important applications in organic chemical industry, the synthesis of plastics, pharmaceutical, pesticides and so on, furfural and its production process have been a hot spot of research in the field of Biomass energy. Furfural, as one of the most promising green platform chemical compound, there is no synthetic route available for furfural production; therefore furfural is exclusively produced from renewable biomass resources by acid-catalyzed dehydration of pentoses (such as cornstalks and corncobs, oat and peanut husks, and other agricultural surpluses)[2].Conventional mineral acids, such as sulfuric acid, hydrochloric acid, phosphoric acid or super phosphate, are generally used as catalysts for the conversion of D-xylose to furfural. At 200-250°C,over the catalyst of sulfuric acid, with the raw material of hemicellulose to produce furfural is commercially adopted method in the industry now. However, these mineral acids often lead to serious corrosion, safety problems, and difficulty in catalyst separation from the reaction products, excessive waste disposal, extensive side reactions, and loss of furfural yield due to long residence times[3].So the improvement of the chemical technology for the production of furfural therefore remains of great interest for the growth of furan-based chemical industries. Solid acids find a wide range of catalytic applications in oil and chemical conversion processes. Solid acid catalysis has attracted much interest both in fundamental and applied research because of its potential to generate significant economic and environmental benefits. Solid acids are widely used catalysts to promote the cracking of hydrocarbons. Therefore, it is hoped that solid acids can be used as catalysts to reduce the viscosity of heavy oil via mild cracking at the lower temperatures. Solid superacid refers to the acid with a strength corresponding to H+. Zirconia promoted by sulfate groups or metal oxides are commonly used as solid superacids[4]. In this paper, D-xylose is catalytically converted to furfural, respectively by zirconia promoted by sulfate groups. We found that the sulfonic acid-functionalized ordered zirconia oxide is an effective catalyst for the dehydration of D-xylose to furfural, using the dimethylsulfoxide (DMSO) as the solvent. International Conference on Materials, Environmental and Biological Engineering (MEBE 2015) © 2015. The authors Published by Atlantis Press 488 Experimental Catalyst preparation. All starting materials were purchased from commercial sources and used as received. Synthesis of 2 2 4 / ZrO SO − . A 16.16g sample of ZrOCl2·8H2O (Wako Pure Chemical) was dissolved in 100 ml of distilled water followed by addition of NH3 solution dropwise with stirring; the final pH of the solution was adjusted to 9. Gelation occurred on standing at room temperature about 72h. After that the solids were filtered and washed with water until a neutral filtrate and absence of chlorine ion was detected by AgNO3 tests. The solids were oven-dried at 65°Cfor 24 h. Then grind into powder. And then the powder was stirred in an aqueous solution of 1.0 M H2SO4 for 24 hour. The catalyst was then separated from the suspension by means of vacuum filtration. After repeating this procedure three times, the solids were oven-dried at 65°Cfor 24 h. Then grind into powder. Then heated to 550°C in air overnight using a 3°C/minute heating ramp and calcined for 5 hours. Catalytic dehydration conversion reaction of xylose to furfural. All reactions were performed under nitrogen and carried out in a biphasic reaction system system containing a reactive aqueous layer (mixture of water and a solid acid) and an extracting organic layer (using dimethylsulfoxide as the extraction solvent). All experiments were conducted using a glass flask, equipped with an agitator and a temperature-controlled electrical heating bath. In a typical experiment, 2g xylose, 1 g NaCl , 2.5g 2 2 4 / ZrO SO − , 30 mL of DMSO were poured into the reactor. The reactor was placed in a heating jacket until boiling for the time specified. After reaction, the reactor was cooled to room temperature by flowing air. The products in the DMSO or aqueous phases were analyzed using UV−visible spectrum. Authentic samples of xylose and furfural were used as standards, and calibration curves were used for quantification[5]. The two curves were showed below: 1 0.0204 0.00734X Y : Fur − = (1) 0450 . 0 0013 . 0 : yl − = X Y X (2) Y: absorbance, X: Concentration(ug/mL) Results and discussion Through analysis, there is no xylose in the organic phase, the conversion of xylose and the furfural yield was calculated as given below. % 100 o × × − = xylose m aqueous V xylose C xylose m nversion C (3)
1/1/15 12:00:00 AM
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6.1.2 6.1 Xylose dehydration to furfural
Dehydration of Biomass-derived Carbohydrates in a Biphasic Reactor using Cation Exchange Resin Catalyst for the Production of Hydroxymethylfurfural and Furfural
Biomass is a source of carbohydrates, which is composed of two different types of sugars hexose (glucose, galactose and mannose) and pentose (xylose and arabinose). These sugars can be dehydrated to produce platform chemicals like 5 - HMF (5 - hydroxymethylfur fural) and furfural. These are most promising chemicals because they are used in the production of fine chemicals, polymers and also starting materials for new products as well as for replacement of oil - derived chemicals. This work focuses on the productio n of 5 - HMF and Furfural by dehydration of fructose and xylose respectively. Dehydration of carbohydrates in aqueous phase can produce unwanted products because of side reactions such as rehydration, hydrolysis and condensation. To eliminate unwanted side r eactions an organic phase and inorganic salt were added to the system. Organic phase consisting of solvents which can extract the product from aqueous phase as it formed to eliminate the degradation of product and addition of salt to the reaction system wi ll increase the partition coefficient. Dehydration reaction was performed in a batch reactor over a cation exchange resin catalyst. Effect of reaction parameters such as temperature, time, and feed compositions were optimized to increase the yield. Effect of salt was studied and concluded that product concentration depends on salt concentration because of salting - out effect. Difference between using glucose and fructose to produce 5 - HMF was also studied and concluded that using fructose product concentrati on increased compared to the glucose. The reason behind this is dehydration of glucose can produce ene - diol intermediate which consists of unstable ring structure that can produce unwanted by products. So, glucose was initially isomerized to fructose and t hen dehydrated to produce 5 - HMF.
1/1/16 12:00:00 AM
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6.1.3 6.1 Xylose dehydration to furfural
Exploitment of niobium oxide effective acidity for xylose dehydration to furfural
Abstract Xylose together with other pentose and hexose sugars can be dehydrated to produce interesting platform chemical compounds, like 5-hydroxyfurfural (HMF) and furfural. This study continues our investigation on the niobium oxide based catalysts in connection with the research of adequate solvents systems to minimize catalyst deactivation and increase catalyst stability and durability during the dehydration of sugars. Silica-zirconia supported niobia samples (10 wt.% of Nb) prepared by impregnation or sol–gel in comparison with pure niobic acid are here presented for xylose dehydration. The reactions have been studied at different temperatures (130–180 °C) in batch or fixed bed continuous catalytic reactors in various solvents. Green solvents soluble in the aqueous solution of xylose or biphasic systems have been taken into account: water, water–isopropanol mixtures, water-γ-valerolactone, and water-cyclopentylmethyl ether. The surface acidities of the catalysts have been measured in cyclohexane ( intrinsic acidity) and also in water to determine the effective catalyst acidity. The continuous tests and batch-recycling tests showed that the supported Nb-catalysts, even if initially less active, are more stable than niobic acid. The presence of isopropanol in water improves both the activity and stability of the catalysts in comparison with water and the use of cyclopentylmethyl ether gave the most interesting selectivity to furfural preserving the catalyst stability.
10/1/15 12:00:00 AM
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6.1.4 6.1 Xylose dehydration to furfural
Selective Production of Furfural from the Dehydration of Xylose using Zn doped CuO Catalyst
Abstract Furfural is a versatile biomass-derived platform compound used for the synthesis of several strategic chemicals. The sonochemically synthesized Zn doped CuO nanoparticles (NPs) were used for the production of furfural. The catalytic activity of the Zn doped CuO NPs was examined, as a model, during the dehydration reaction of xylose to furfural. In addition to that, we have also compared the catalytic activity of the Zn doped CuO NP with ZnO NPs, ZnO bulk, CuO NPs, CuO bulk, etc. This nanoscale catalyst (Zn doped CuO NP) has a large surface area, which enhances its catalytic activity and enables it to completely convert the xylose to furfural at 150 °C within 12 h without any trace of by-products, as confirmed by HPLC, 13 C NMR and 1 H NMR. HPLC analysis demonstrated that the yield of furfural is up to 86 mol %, compared to the 45 mol % obtained with ZnO NPs, ZnO bulk, CuO NPs, CuO bulk, etc. as catalysts.
3/1/19 12:00:00 AM
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6.1.5 6.1 Xylose dehydration to furfural
SnCl4-catalyzed isomerization/dehydration of xylose and glucose to furanics in water
A number of Lewis acid catalysts were screened for their effectiveness in converting both xylose and glucose in aqueous media to furfural and 5-HMF, respectively. While other catalysts were found to be more active, SnCl4 was identified as the most selective Lewis acid. Hydrolysis of SnCl4 was observed at various concentrations and temperatures resulting in the production of Bronsted acidic protons in a 3.5 : 1 ratio to Sn4+ at all SnCl4 concentrations above 60 °C. As a consequence, there was no need to add a Bronsted acid in order to promote the dehydration of either xylose or glucose. SnCl4-promoted isomerization/dehydration of xylose and glucose at 140 °C in water resulted in conversions of 55% and 33%, respectively, after 2 h of reaction, and furfural and 5-HMF selectivities of up to 58% and 27%, respectively. Significant conversion of sugars to humins was observed in both cases, and in the case of glucose, degradation of 5-HMF to levulinic and formic acids was also noted. The effects of secondary reactions could be greatly suppressed by extraction of the furanic product as it was produced. Using n-butanol as the extracting agent, xylose and glucose conversions of 90% and 75%, respectively, were observed after 5 h of reaction, and the selectivities to furfural and 5-HMF increased to 85% and 69%, respectively. Small additional increases in the furfural and 5-HMF selectivities were obtained by adding LiCl to the aqueous phase without much effect on the conversion of either sugar. In this case, the selectivities to furfural and 5-HMF were 88% and 72%, respectively, after 5 h of reaction at 140 °C.
1/1/15 12:00:00 AM
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6.1.6 6.1 Xylose dehydration to furfural
Studies of Brønsted/Lewis Acid-Catalyzed Dehydration of Xylose to Furfural and Simultaneous Separation of Furfural by Pervaporation
Author(s): Wang, Alex | Advisor(s): Balsara, Nitash P; Bell, Alexis T | Abstract: A major component of lignocellulosic biomass is hemicellulose, a polysaccharide composed of monomeric sugars, principally xylose. Xylose can be dehydrated, most often in aqueous solution, using Bronsted acid catalysts to form furfural, which can be further reacted to produce fuels, lubricants, polymers, solvents, and pharmaceutical precursors. Furfural production can also be enhanced by using Lewis acid catalysts, which promote the formation of xylulose, an isomer of xylose which more readily dehydrates to form furfural. With either type of catalyst, side reactions consume furfural to produce a group of soluble and insoluble products known as humins. Humins formation has been stymied by extracting furfural as it is produced. This is done on the industrial scale with steam stripping, but researchers have also explored the use of liquid-liquid extraction (LLE) by an organic solvent (typically 2:1 organic:aqueous volume ratio) for the same purpose. Both extraction methods increase furfural yield, but dilute the product phase, which raises the cost of furfural production. A new method, e.g. pervaporation, must be developed to increase furfural yield and concentrate furfural in the product simultaneously.Pervaporation is a membrane-based process in which a liquid mixture is placed in contact with the feed side of the membrane while a vapor is located on the permeate side. A vacuum is used to reduce the partial pressure, and therefore the fugacity, of components in the permeate, which provides the driving force for mass transfer. Pervaporation is most often used to separate water from concentrated ethanol solutions, but may also be used to remove organics selectively, e.g. furfural, from aqueous solutions. Membranes used for such applications are typically made of polydimethylsiloxane (PDMS), but researchers have also used the PDMS-containing triblock copolymer poly(styrene-block-dimethylsiloxane¬-block-styrene) (SDS). Pervaporation with a furfural-selective membrane may be used to extract furfural as it is produced and concentrate it, rather than dilute it as steam stripping and LLE do.The objective of this investigation was to assess pervaporation as a method to extract furfural during its production. This was done by designing and constructing membrane reactors, comparing them to LLE-assisted reactors through experiments and simulations, and studying how Lewis acid catalysts can improve reaction and pervaporation compatibility and lead to the formation of novel products.The feasibility of pervaporation as a means for in situ furfural extraction was studied in comparison to LLE and a reaction without extraction during batch-mode furfural production. Both LLE and pervaporation with a commercial PDMS membrane were found to improve furfural yield over the reaction without extraction, but pervaporation with PDMS yielded a product phase that was 6.6x as concentrated as that obtained with LLE. Additionally, switching the PDMS membrane with an SDS membrane resulted in similar furfural yields, but the product with SDS was 10x as concentrated as the LLE product. Furthermore, the amount of furfural extracted was qualitatively different for LLE- and pervaporation-assisted reactions: LLE was limited to 85%, the equilibrium distribution of furfural among the organic and aqueous, whereas the amount of furfural extracted by pervaporation increased monotonically over time, reaching as high as 67% during experiments. The reaction/pervaporation system was simulated in order to identify the full extent of the benefits of reaction with pervaporation. In the simulations, water lost from the reactor due to removal by pervaporation was replenished at the equivalent rate. The simulations revealed that as the reaction approached complete xylose conversion, both the PDMS and SDS membranes led to product concentrations greater than was possible with LLE, while extracting nearly all (g98%) of the furfural formed. Ultimately, pervaporation with the SDS membrane could produce a product phase with 33% greater furfural yield than that achievable by LLE.The membrane-reactor design was revised to permit continuous, pervaporation-assisted reaction in both batch- and continuous-mode operation, with both reaction and pervaporation occurring at the same temperature. Batch-mode reactions were fed water, while continuous-mode reactions were fed an aqueous xylose solution. The reactions took place at a relatively low temperature of 90 °C, catalyzed by chromium (III) chloride (CrCl3), which contributed both Bronsted and Lewis acidity. Batch-mode reactions with varying rates of pervaporation revealed that furfural extraction had no effect on furfural yield under these conditions, but a moderate pervaporation rate did lead to an order-of-magnitude increase in furfural concentration relative to that obtained without pervaporation. Pervaporation was also found to retain all of the CrCl3 inside the reactor, demonstrating a simple way to separate product from homogeneous catalyst. This enabled continuous furfural production with only an initial charge of catalyst, in which an aqueous xylose solution was fed to the reactor while a furfural/water vapor was permeated from the reactor. The furfural permeability of the SDS membrane decreased over time during the course of reactions carried out at 90 °C due likely to interactions of soluble humins with the membrane. Experiments with the cross-linked PDMS membrane demonstrated that cross-linking of the membrane can inhibit this behavior and result in a much more stable furfural permeability. Additionally, cross-linking could lead to greater membrane thermal stability, permitting the pervaporation-assisted reaction at higher temperatures, which would benefit the chemistry by allowing extraction to have an impact on furfural yield. Pervaporation-assisted furfural production with CrCl3 and sulfuric acid at 130 °C was then simulated. Reaction rate constants were measured at this temperature but in the absence of pervaporation. Pervaporation data collected at lower temperatures were extrapolated to represent a hypothetical membrane that could operate at 130 °C. Simulations of batch-mode reactions demonstrated that increasing the membrane-area-to-reactor-volume ratio, a, would lead to higher furfural yield and more furfural extracted, but also reduce the permeate furfural concentration, demonstrating a tradeoff between furfural production and concentration. Simulations of continuous-mode reactions showed that furfural concentration and selectivity were maximized at an intermediate value of a = 0.17 cm-1. Conversely, furfural production rate increased nearly linearly with a, indicating that the optimal value of a depends on process economics and not just technical considerations.The Lewis acidity of CrCl3 was beneficial for reducing reaction temperature within the membrane stability limits, but Lewis acids, such as the Sn-containing zeolite Sn-BEA, have been shown to convert sugars (i.e., xylose and glucose) at a rate greater than the rate of formation of identified products, suggesting that additional, unidentified products are formed. Through extensive analytical chemistry, these products were determined to be hydroxyl-rich carboxylic acids and furanone esters that form by structural isomerization of the sugars, followed by dehydration, and constitute as much as 45% of the yield. These side-produced acids and esters may find use as monomers for the synthesis of biodegradable polyesters, which are often used for sutures, bone prostheses, and controlled drug delivery. This work demonstrates that Lewis acid catalysts are not only useful for bridging the gap between pervaporation-membrane limits and furfural production temperatures, but also for the formation of additional value-added chemicals.
1/1/17 12:00:00 AM
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6.1.7 6.1 Xylose dehydration to furfural
The First Biomass Refinery for the Production of Pulp, Lignins and C5 Sugars at Industrial Scale from Annual Plants and Hard Wood
The industrial process developed by CIMV is a world premiere for the manufacture of whitened paper pulp, sulphur free linear lignin and xylose syrup from annual fibre plants and hard wood. This new technology allows the separation without degradation of the three main components of the vegetable matter which are cellulose, hemicelluloses and lignins. It is a biomass refinery working on the model of an oil refinery. The organic acids, used for the biopolymers separation, are recycled by evaporation from the organic solution. The remaining syrup is treated with water to precipitate lignins which are separated from the remaining sugar solution. The lignins show a linear structure which allows a high reactivity in particular with a lot of monomers towards new polymers and new formaldehyde free adhesive formulations. The C5 sugar syrup can be used to produce additives for animal feeding, furfurol, furfurylic alcohol etc... The development at an industrial scale takes its course. The first CIMV factory has been designed to treat 150.000 tons of dry straw and will open in October / November 2009 in Vitry Le Francois in the north east of France.
1/1/08 12:00:00 AM
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6.2 Microwave-assisted furfural production

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By means of catalytic reactions in an aqueous medium, it was determined that Dxylose can efficiently be converted into furfural by the application of microwave as a green synthetic methodology. The highest yields of furfural were obtained at a HCl concentration of 4 mg/mL. When the reaction was performed at 200 °C, an optimum yield of 64% of furfural was observed after 10 min of reaction time, with 95% of the Dxylose being converted. Art. [#ARTNUM](#article-25199-1851052610)

6.2.1 6.2 Microwave-assisted furfural production
Efficient microwave-assisted production of furfural from C5 sugars in aqueous media catalysed by Brönsted acidic ionic liquids
Small amounts of SO3H-functionalised room temperature synthesized ionic liquids efficiently dehydrate aqueous xylose to furfural under microwave heating at mild reaction conditions. The RT-ionic liquid catalysts were also found to be effective catalysts for the two step one-pot simultaneous hydrolysis and dehydration of a lignocellulosic waste biorefinery-derived syrup enriched in C5 sugar oligomers.
1/1/12 12:00:00 AM
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6.2.2 6.2 Microwave-assisted furfural production
Microwave-Assisted Green Production of Furfural from D-xylose of Sugarcane Bagasse
D-xylose is a component of sugarcane bagasse that can be used as a renewable resource for the production of a variety of chemicals. By means of catalytic reactions in an aqueous medium, it was determined that D-xylose can efficiently be converted into furfural by the application of microwave as a green synthetic methodology. The highest yields of furfural were obtained at a HCl concentration of 4 mg/mL. When the reaction was performed at 200 °C, an optimum yield of 64% of furfural was observed after 10 min of reaction time, with 95% of the D-xylose being converted.
10/26/15 12:00:00 AM
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6.3 Catalytic coversion of hexoses to 5HMF

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Galactose and mannose can be converted to 5-HMF by dehydration, with different catalytic methods. Hydroxymethylfurfural (HMF), also 5-(hydroxymethyl)furfural, is an organic compound formed by the dehydration of certain sugars. It is also produced industrially on a modest scale as a carbon-neutral feedstock for the production of fuels and other chemicals. **Research findings** - In this work, Cr, Al and Sn/MCM41 catalysts were prepared by a simple impregnation method and characterized. The conversion of mannose into 5HMF was evaluated in dimethyl sulfoxide (DMSO) solvent. It was found that the assynthesized Sn/MCM41 catalyst showed a superior activity to Cr/MCM41 and Al/MCM41. Mannose could be effectively converted into 5HMF with a yield of ∼45% and ∼88% conversion at 150 °C after 60 min, which were comparable to reported results over heterogeneous catalysts. Art. [#ARTNUM](#article-25201-2891887239) **Notes** 5-HMF can act as building block to: - 2,5-di(hydroxymethyl)furan (DHMF) & 2,5-di(hydroxymethyl)-tetrahydrofuran (DHMTHF). Art. [#ARTNUM](#article-25201-2302279843) - 2,5-dimethylfuran, 2,5-dihydromethylfuran, 2,5-dihydromethyl-tetrahydrofuran, 5-thoxymethylfurfural, 1,6-hexanediol, longchain alkanes, 3-(hydroxymethyl)-cyclopentanone, pxylene, 2,5-diformylfuran, 2,5-furandicarboxylic acid and maleic anhydride. Art. [#ARTNUM](#article-25201-2810881574)

6.3.1 6.3 Catalytic coversion of hexoses to 5HMF
Catalytic conversion of 5-hydroxymethylfurfural to some value-added derivatives
5-Hydroxymethylfurfural (HMF) is a platform chemical derived from C6 sugars, which can be transformed into various important chemicals and fuels because of the presence of CO, C–O and furan ring functional groups. In this review, the selective tailoring of these groups in HMF to form 2,5-dimethylfuran, 2,5-dihydromethylfuran, 2,5-dihydromethyltetrahydrofuran, 5-ethoxymethylfurfural, 1,6-hexanediol, long-chain alkanes, 3-(hydroxy-methyl)cyclopentanone, p-xylene, 2,5-diformylfuran, 2,5-furandicarboxylic acid and maleic anhydride will be described to gain more insight into the transformation of HMF under various conditions. The focus of this review is on the mechanisms of the catalytic processes and potential design strategies for future catalysts. The activation of the functional groups and the key challenges involved in the precise design of bifunctional catalysts are highlighted. Some examples of “one-pot” transformations of fructose into various chemicals using the HMF platform are also presented.
1/1/18 12:00:00 AM
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6.3.2 6.3 Catalytic coversion of hexoses to 5HMF
Catalytic dehydration of C6 carbohydrates for the production of hydroxymethylfurfural (HMF) as a versatile platform chemical
Interest in utilizing biorenewable feedstocks to produce fuels and chemicals has risen greatly in the past decade due to the economic, political and environmental concerns associated with diminishing petroleum reserves. A fundamental challenge lying ahead in the development of efficient processes to utilize biomass feedstock is that, unlike their petroleum counterparts, biomass contains an excess amount of oxygen. Therefore, catalytic strategies such as dehydration and hydrogenolysis amongst others have been extensively studied as platform technologies for deoxygenation. In this review, we primarily discuss the catalytic dehydration of C6 carbohydrates to 5-hydroxymethylfurfural, which has attracted much attention due to the versatility of using furanic compounds as an important platform intermediate to synthesize various chemicals. The emphasis is on the fundamental mechanistic chemistry so as to provide insights for further catalyst/catalytic system design. After separately discussing fructose and glucose dehydration, this review summarizes recent progress with bi-functional catalyst systems for tandem glucose/fructose isomerization and subsequent fructose dehydration, thereby realizing highly selective HMF production directly from the more abundant and cheaper C6 sugar feedstock, glucose.
1/1/14 12:00:00 AM
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6.3.3 6.3 Catalytic coversion of hexoses to 5HMF
Efficient synthesis of 5-hydroxymethylfurfural from mannose with a reusable MCM-41-supported tin catalyst
The synthesis of furan compounds by renewable carbohydrates has emerged as an important path toward biomass utilization. 5-Hydroxymethylfurfural (5-HMF) and furfural are two typical furan products from cellulosic and hemicellulosic materials, of which the primary components (hexoses and pentoses) undergo selective dehydration. It is worth noting that hemicellulose is rich in not only pentoses but also hexoses, indicating the feasibility of producing 5-HMF and furfural simultaneously. However, research on the production of 5-HMF from hemicellulose-derived hexoses remains lacking. Mannose, a major hexose component of hemicellulose, has been previously found to be a suitable feedstock for 5-HMF production in the presence of homogeneous catalysts, such as metal salts. In this work, Cr-, Al- and Sn/MCM-41 catalysts were prepared by a simple impregnation method and characterized. The conversion of mannose into 5-HMF was evaluated in dimethyl sulfoxide (DMSO) solvent. It was found that the as-synthesized Sn/MCM-41 catalyst showed a superior activity to Cr/MCM-41 and Al/MCM-41. Mannose could be effectively converted into 5-HMF with a yield of ∼45% and ∼88% conversion at 150 °C after 60 min, which were comparable to reported results over heterogeneous catalysts. The as-synthesized Sn/MCM-41 catalyst was also efficient for the conversion of glucose and fructose with reasonable 5-HMF yields. The Sn/MCM-41 catalyst could be reused for eight consecutive cycles without significant loss of catalytic activity.
1/1/18 12:00:00 AM
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6.3.4 6.3 Catalytic coversion of hexoses to 5HMF
METHOD FOR SYNTHESISING 2,5-DI(HYDROXYMETHYL)FURAN AND 2,5-DI(HYDROXYMETHYL)TETRAHYDROFURAN BY SELECTIVE HYDROGENATION OF FURAN-2,5-DIALDEHYDE
A method for selective hydrogenation of furan-2,5-dialdehyde (DFF) into 2,5-di(hydroxymethyl)furan (DHMF) and into 2,5-di(hydroxymethyl)tetrahydrofuran (DHMTHF). In relation to the prior art, which uses C6 sugars or 5-hydroxymethyl furaldehyde (5-HMF) as raw materials, the method can be performed at low temperatures (lower than 120° C., preferably 80° C.), while consuming low amounts of catalyst relative to the initial reagent (in particular less than 5%, preferably less than 2% relative to the weight of the reagent). The heterogeneous catalyst used can also be recycled from one reaction to another. Finally, the choice of experimental conditions enables the selective formation of DHMF or DHMTHF.
9/26/13 12:00:00 AM
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6.3.5 6.3 Catalytic coversion of hexoses to 5HMF
Method for preparing 5-hydroxymethyl furfural (5-HMF) through mannose conversion
The invention discloses a method for preparing 5-hydroxymethyl furfural (5-HMF) through catalytic conversion of mannose in a liquid-phase system, and relates to the preparation of fine chemicals with high additional values through efficient conversion of carbohydrate substances, belonging to the field of conversion and utilization of biomass resources. In the invention, different types of metal salts and inorganic acids are used as a catalyst respectively, and mannose is effectively converted into 5-HMF in an organic solvent or a water-organic solvent mixed system. In the invention, the adopted catalyst and solvent are relatively cheap, the operation is simple, the reaction conditions are mild, the mannose can be quickly and directly converted into 5-HMF, and the yield of 5-HMF is relatively high. The information provided by the invention can provide useful reference to efficient conversion of other kinds of hexose and lignocellulose.
9/28/16 12:00:00 AM
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6.3.6 6.3 Catalytic coversion of hexoses to 5HMF
Method for preparing 5-hydroxymethyl furfural by degrading galactose by using double metal chloride
The invention discloses a method for preparing 5-hydroxymethyl furfural by degrading galactose by using double metal chloride, belonging to the field of biomass synthesized renewable energy source and chemical product preparation. The method comprises the following step: by adopting double metal chloride as a catalyst, degrading galactose to 5-hydroxymethyl furfural in a binary solvent system. As galactose is adopted as a raw material, and 5-hydroxymethyl furfural (HMF) is prepared through direct catalytic transformation under gentle conditions, the method has the advantages of rapidness, simplicity, convenience, simple process, gentle reaction and high efficiency, and a valuable novel method is provided for transforming galactose into a bulk chemical. The catalytic activity of metal chloride is higher than that of a single metal catalyst which is high in price, and is an excellent system for catalytic degradation of carbohydrate such as sugar.
3/25/15 12:00:00 AM
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6.3.7 6.3 Catalytic coversion of hexoses to 5HMF
Method for preparing 5-hydroxymethylfurfural (5-HMF) by converting galactose
The invention relates to a method for preparing 5-hydroxymethylfurfural (5-HMF) by converting galactose, relating to a method for preparing biomass 5-HMF and belonging to the field of preparation of high-added-value chemicals and alternative energy sources from renewable biomass resources. According to the method, galactose is converted into 5-HMF in a water-organic solvent two-element mixture system by using a metal salt as a catalyst. By using the galactose as the raw material, the galactose can be directly and quickly converted into the 5-HMF under mild conditions. The method is simple in process and easy to operate, has the advantages of mild conditions and efficient reaction performance, provides a favorable reference for galactose conversion, and provides useful information for the development of new energy technology by using saccharide biomass as the raw material in future. The catalytic reaction system in the method is a favorable system capable of selectively converting hexose raw materials.
6/15/16 12:00:00 AM
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6.3.8 6.3 Catalytic coversion of hexoses to 5HMF
Synthesis of Renewable Fine‐Chemical Building Blocks by Reductive Coupling between Furfural Derivatives and Terpenes
A new chemical industry, in which renewable resources such as lignocellulose or its constituents cellulose, lignin, and hemicellulose are the new raw materials, is slowly emerging. These raw materials can be transformed into a number of platform chemicals, such as 2-furfural, 5-hydroxymethyl-2-furfural, levulinic acid, succinic acid, and a host of other compounds. These are the new bulk chemicals. Whereas most research in this emerging field concentrates on the conversion of biomass into platform chemicals and their conversion into monomers for polymers, very few researchers have taken the next step towards fine chemicals. Obviously, making larger, higher functionalized molecules from these C2 to C6 platform chemicals entails the use of C C bond-forming reactions. In order to extend the low-carbon-footprint chemistry that is connected with biomass conversion it is important to focus on catalyzed C C bond forming reactions that avoid the use of stoichiometric leaving groups. Chemistry developed by Krische and co-workers using reductive coupling of alkenes, dienes, and alkynes to electrophiles undeniably matches this important criterion. This methodology is by now well-developed and has a wide scope. A range of coupling reactions between aldehydes or primary alcohols and different unsaturated compounds such as dienes, allenes, allyl acetate, and many other substrates using a metal catalyst and without the use of stoichiometric reagents has been described. Many other researchers have now contributed to this important field. We only found one example where this type of chemistry is applied to furfural. Furfural is readily obtainable by treatment of C5 sugars with acids at elevated temperatures. In practice, it is often made from agricultural waste such as oat hulls, which contain C5sugar-rich hemicellulose. Its worldwide production volume is already in excess of 400000 ta . Furfural has a range of applications. To build a new product tree based on this readily available platform chemical, we evaluated its reductive coupling to renewable terpenes. Thus, we report herein the coupling of 5substituted furfurals with isoprene and myrcene, using the conditions developed by Krische et al. In a first example furfural (1a) was coupled with isoprene 2 to form b,g-unsaturated ketone 3a in good yield (Table 1, entry 1). The catalyst used is the ruthenium dihydride complex
9/1/13 12:00:00 AM
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7. Other synthetic routes

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7.1 Conversion route of pentoses to 3,4-dihydroxyprolines

0

The synthesis of two naturallyoccurring isomers of 3,4dihydroxyproline is reported. l 2,3 cis 3,4 trans 3,4Dihydroxyproline was synthesized from l arabinose in 10 steps and 31% overall yield. The same series of reactions was employed to convert l xylose to l 2,3 trans 3,4 trans 3,4dihydroxyproline. Art. [#ARTNUM](#article-25192-1963983380)

7.1.1 7.1 Conversion route of pentoses to 3,4-dihydroxyprolines
The conversion of pentoses to 3,4-dihydroxyprolines
Abstract The synthesis of two naturally-occurring isomers of 3,4-dihydroxyproline is reported. l -2,3- cis -3,4- trans -3,4-Dihydroxyproline was synthesized from l -arabinose in 10 steps and 31% overall yield. The same series of reactions was employed to convert l -xylose to l -2,3- trans -3,4- trans -3,4-dihydroxyproline. Orthogonally protected versions of these amino acids were produced on gram scale, en route to the free amino acids, and these will serve as versatile intermediates in peptide synthesis. This synthetic strategy involved Nα -Fmoc protection and protection of the C3 and C4 secondary alcohols as methoxyethoxymethyl (MEM) ethers.
10/1/05 12:00:00 AM
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7.2 Xylose conversion to xylonic acid and/or xylitol

0

Xylose can be converted to xylitol and xylonic acid in several ways: **Research findings** - Xylose can be reduced to xylitol and oxidized to xylonic acid through electrocatalytic conversion. Art. [#ARTNUM](#article-25221-2060588946) - Xylose can be oxidized to xylonic acid and hydrogenated to xylitol by solid-based metal catalysts. Art. [#ARTNUM](#article-25221-1973098454)

7.2.1 7.2 Xylose conversion to xylonic acid and/or xylitol
Simultaneous electrolytic production of xylitol and xylonic acid from xylose
A new electrocatalytic process for simultaneous electrolytic production of xylitol and xylonic acid or the corresponding xylonate salts directly from α-d-xylose in alkaline solutions of neutral salt as supporting electrolyte has been developed and presented. An electrocatalytically active cathode coating with essentially hydridic features has been employed to enhance Faradaic yields in xylitol production at low current densities by heterogeneous reaction of hydrogenation with H-adatoms, and the entire mechanism has been proved by experimental evidence. An anionic selective anodic coating for titanium substrate was also used to suppress oxygen evolution and optimize the xylonic acid production by direct oxidation with anodically generated bromine, and to carry out the electrode process at almost equilibrium potential. The kinetic effect of specifically adsorbable anions for decreasing cathodic current yields in xylitol production and the resulting necessity for membrane separation of catholyte from anolyte, with proper optimization of the supporting neutral salt composition, has also been emphasized.
4/1/91 12:00:00 AM
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7.2.2 7.2 Xylose conversion to xylonic acid and/or xylitol
Solid base supported metal catalysts for the oxidation and hydrogenation of sugars
Abstract Pt impregnated on γ-Al 2 O 3 (acidic support) and hydrotalcite (basic support) catalysts were synthesized, characterized and used in the oxidation and hydrogenation reactions of C5 and C6 sugars. In the absence of homogeneous base, 83% yield for gluconic acid; an oxidation product of glucose can be achieved over Pt/hydrotalcite (HT) catalyst at 50 °C under atmospheric oxygen pressure. Similarly, 57% yield for xylonic acid, an oxidation product of xylose is also possible over Pt/HT catalyst. Hydrogenation of glucose conducted using Pt/γ-Al 2 O 3  + HT catalytic system showed 68% sugar alcohols (sorbitol + mannitol) formation. The 82% yield for C5 sugar alcohols (xylitol + arabitol) was obtained by subjecting xylose to hydrogenation over Pt/γ-Al 2 O 3  + HT at 60 °C. UV analysis helped to establish the fact that under alkaline conditions sugars prefer to remain in open chain form in the solution and thus exposes CHO group which further undergoes oxidation and hydrogenation reactions to yield acids and alcohols.
7/1/14 12:00:00 AM
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7.3 Furfural to methylfuorate

0

Furfural can be produced from xylose through dehydrogenation as shown in technology [Tech 6.1](#technology-25200). Gold supported catalysts have been successfully proven to be highly active and selective in the furfural oxidative esterification to methyl-2-furoate under mild conditions by employing oxygen as benign oxidant. Particular attention has been given to the studies in which the reaction occurs even without base as co-catalyst, which would lead to a more green and economically advantageous process. The Au catalysts are also stable and quite easily recovered and represent a feasible and promising route to efficiently convert furfural to methyl-2-furoate to be scaled up at industrial level. Art. [#ARTNUM](#article-30812-2492518672)

7.3.1 7.3 Furfural to methylfuorate
Biomass Derived Chemicals: Furfural Oxidative Esterification to Methyl-2-furoate over Gold Catalysts
The use of heterogeneous catalysis to upgrade biomass wastes coming from lignocellulose into higher value-added chemicals is one of the most explored subjects in the prospective vision of bio-refinery. In this frame, a lot of interest has been driven towards biomass-derived building block molecules, such as furfural. Gold supported catalysts have been successfully proven to be highly active and selective in the furfural oxidative esterification to methyl-2-furoate under mild conditions by employing oxygen as benign oxidant. Particular attention has been given to the studies in which the reaction occurs even without base as co-catalyst, which would lead to a more green and economically advantageous process. The Au catalysts are also stable and quite easily recovered and represent a feasible and promising route to efficiently convert furfural to methyl-2-furoate to be scaled up at industrial level.
7/20/16 12:00:00 AM
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7.4 Furfural to cyclopentanone/ cylopentanol

0

Furfural can be produced from xylose through dehydrogenation as shown in technology [Tech 6.1](#technology-25200). Cu–Co catalysts, prepared by a co-precipitation method (CP) and an oxalate sol–gel method (OG), can selectively convert furfural (FFA) to cyclopentanone (CPO) or cyclopentanol (CPL), respectively. At lower hydrogen pressure (2 MPa) and lower Cu loadings (2% for OG, 5% for CP), we obtained the highest yield of 67% CPO and 68% CPL, respectively. Art. [#ARTNUM](#article-30813-1977262992)

7.4.1 7.4 Furfural to cyclopentanone/ cylopentanol
Selective conversion of furfural to cyclopentanone or cyclopentanol using different preparation methods of Cu–Co catalysts
Cu–Co catalysts, prepared by a co-precipitation method (CP) and an oxalate sol–gel method (OG), can selectively convert furfural (FFA) to cyclopentanone (CPO) or cyclopentanol (CPL), respectively. The conversion of FFA to CPO or CPL by Cu–Co catalysts were studied in aqueous solutions. We found that the product distribution was influenced by the catalyst support, Cu loading, calcination temperature, hydrogen pressure, the number of times the catalyst was reused and the preparation method of the catalyst. The surface morphology, surface area and composition of the catalysts were studied by XRD, XPS, BET, ICP-AES and TEM characterization. We found that there was a strong interaction between Cu and Co. Cu0, Cu2O and Co0 were the main active catalyst phases on the surfaces of the catalysts, but the amounts were different in the different catalysts. Cu0, Co0 and Cu2O were the active hydrogenation species, and Cu2O also played the role of an electrophile or Lewis acid to polarize the CO bond via lone pair electrons on the oxygen atom. According to XRD and XPS, the main phases on the surface of the CP catalysts were Cu0 and Cu2O. The hydrogenation activity of the CP catalyst was relatively weak and the main product was CPO. In contrast, the hydrogenation activity of the OG catalyst was high and the main product was the fully hydrogenated product CPL due to the main active phases of Co0 and Cu2O on the surface of the OG catalyst. At lower hydrogen pressure (2 MPa) and lower Cu loadings (2% for OG, 5% for CP), we obtained the highest yield of 67% CPO and 68% CPL, respectively.
1/1/15 12:00:00 AM
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7.5 Xylose conversion in alkaline environment

0

**Research findings** Treatment of D-xylose and D-glucose with 0.63M sodium hydroxide at 96° in an atmosphere of nitrogen yielded, in addition to acidic, aliphatic degradation-products, the following cyclic enols and phenolic compounds: - 2-hydroxy-3-methyl-2-cyclopenten-1-one (1), - 2-hydroxy-3,4-dimethyl-2-cyclopenten-1-one (2), - pyrocatechol (3), - 3-methyl-1,2-benzenediol (4), - 4-methyl-1,2-benzenediol (5), - 3,4-di-methyl-1,2-benzenediol (6), - 2-methyl-1,4-benzenediol (7), - 2,5-dihydroxyacetophe-none (8), - 3-hydroxy-5-methylacetophenone (9), - 3,4-dihydroxyacetophenone (10), - 3,4-hydroxybenzaldehyde (11), - 2,3,4-trihydroxy-5-methylacetophenone (12), and; - 2,3-dihydroxy-6-methylacetophenone (13). Art. [#ARTNUM](#article-25224-2172295485)

7.5.1 7.5 Xylose conversion in alkaline environment
Reactions of D-xylose and D-glucose in alkaline, aqueous solutions☆
Abstract Treatment of D -xylose and D -glucose with 0.63 M sodium hydroxide at 96° in an atmosphere of nitrogen yielded, in addition to acidic, aliphatic degradation-products, the following cyclic enols and phenolic compounds: 2-hydroxy-3-methyl-2-cyclopenten-1-one ( 1 ), 2-hydroxy-3,4-dimethyl-2-cyclopenten-1-one ( 2 ), pyrocatechol ( 3 ), 3-methyl-1,2-benzenediol ( 4 ), 4-methyl-1,2-benzenediol ( 5 ), 3,4-di-methyl-1,2-benzenediol ( 6 ), 2-methyl-1,4-benzenediol ( 7 ), 2,5-dihydroxyacetophe-none ( 8 ), 3-hydroxy-5-methylacetophenone ( 9 ), 3,4-dihydroxyacetophenone ( 10 ), 3,4-hydroxybenzaldehyde ( 11 ), 2,3,4-trihydroxy-5-methylacetophenone ( 12 ), and 2,3-dihydroxy-6-methylacetophenone ( 13 ).
5/1/76 12:00:00 AM
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Final Results

Published 10/2/19

After the midway results meeting, 9 techniques have been reviewed and deepened. The results are organised based on the concept and presented per techniques comprising a description, findings, suppliers (if applicable), images, videos, useful links and a reference list. The technology requirements are measured and shown in the [requirements table](#requirements-table). By using the concept links below, you can quickly navigate to the concepts and their techniques descriptions.

Table of concepts:

  1. 1. Biological routes to alcohols
  2. 2. Biological routes to organic acids
  3. 3. Other biological routes

Technology Radar
Requirements Table

1. Biological routes to alcohols

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1.1 Ethylene glycol

1

Ethylene glycol (IUPAC name: ethane-1,2-diol) is an organic compound with the formula (CH2OH)2. It is mainly used for two purposes, as a raw material in the manufacture of polyester fibers and for antifreeze formulations. It is an odorless, colorless, sweet-tasting, viscous liquid. [Wiki](https://en.wikipedia.org/wiki/Ethylene_glycol) **Process** * E. coli engineered pathway (Penthose phosphate + D-arabinose degradation): yield 0.35 g/g (0.84 mol/mol) xylose at a titer of 40 g/L: * Batch bioreactor fermentations were carried out in a 3-L Bioflo culture vessel (New Brunswick, CT, USA) with a 2-L working volume. These fermentations utilized minimal medium (MM2) consisting of 2.0 g/L NH4Cl, 5.0 g/L (NH4)2SO4, 2.0 g/L KH2PO4, 0.5 g/L NaCl, 2 mL/L 1 M MgSO4, 1 mL/L mineral solution, 0.1 mL/L 4 mM Na2MoO4, and specified sugar; additionally, silicone antifoaming B emulsion was used to prevent foaming. Aerobic conditions were maintained by sparging air at 0.5 or 1 L/min, and the pH was maintained at 7.0 with 6 N NaOH. For all cultures, 50 mg/L spectinomycin, 34 mg/L chloramphenicol, and 50 mg/L kanamycin were added as appropriate. To induce gene expression, 0.1 mM isopropyl β-D-1-thiogalactopyranoside (IPTG) (Zymo Research, Irvine, CA, USA) and/or 250 μg/L anhydrotetracycline (aTc) were added. * For fed-batch bioreactor production of EG from D-xylose, an initial culture of strain EG-X was grown in LB at 37 °C, 250 rpm o/n. This culture was used to inoculate to 1% v/v two 50-mL flask cultures of MM1 with 15 g/L D-glucose. After growing o/n at 37 °C, 250 rpm, these seed cultures were combined and used to inoculate to 5% v/v a bioreactor containing MM2 with approximately 35 g/L D-xylose. Temperature was maintained at 37 °C, and dissolved oxygen content was maintained at 30% by altering agitation from 400 to 650 rpm. At 24 h, we initiated pumping of a feed solution into the bioreactor through a reactor port. The feed solution consisted of 600 g/L D-xylose, 10 g/L (NH4)2SO4, 5.0 g/L MgSO4, 0.1 mM IPTG, and spectinomycin. The flow rate was varied from 0.05 mL/min to 0.15 mL/min so as to maintain a concentration of D-xylose between 0 and 10 g/L. Art. [#ARTNUM](#article-25156-2214411608) * E. coli engineered pathway (xylulose 1P pathway): 0.91 mol/mol, 20 g/L 0.37 g/L/h. Growth of the cultures was realized in M9 minimal medium that contained (d)-glucose or (d)-xylose at concentrations of 20 or 10 g/L, respectively, together with 18 g/L Na2HPO4 · 12H2O, 3 g/L KH2PO4, 0.5 g/L NaCl, 2/L g NH4Cl, 0.5 g/L MgSO4 · 7H2O, 0.015 g/L CaCl2 · 2H2O, 0.010 g/L FeCl3, 0.006 g/L Thiamine HCl, 0.4 mg/L NaEDTA · 2H2O, 1.8 mg/L CoCl2 · 6H2O, 1.8 mg/L ZnCl2SO4 · 7H2O, 0.4 mg/L Na2MoO4 · 2H2O, 0.1 mg/L H3BO3, 1.2 mg/L MnSO4 · H2O, 1.2 mg/L CuCl2 · 2H2O. 3-(N-morpholino)propanesulfonic acid (MOPS) solution at pH 7 was used to buffer M9 minimal medium. 0.5 l bioreactors (MiniBio, Applikon) that contained 250 mL medium at an OD600 of \~1.2. The composition of the fermentation medium was similar to the mineral medium used in the shake flask experiments, except that it contained 55 g/L (d)-xylose, 2 g/L Na2HPO4 · 12H2O, 0.8 g/L KH2PO4, 6 g/L (NH4)2HPO4, 0.4 g/L (NH4)2SO4, 1 g/L tryptone (Biokar), 0.5 g/L yeast extract (Biokar), 0.4 mL/L polypropylene glycol as antifoaming agent, 1 mM IPTG, and no MOPS. The pH of the cultures was kept at 7.0 by the addition of 10 M KOH, and reactors were aerated with air at 1 vvm. Dissolved oxygen tension was regulated by adjusting the appropriate agitation speed (300–1,200 rpm, Rushton rotor, 28 mm diameter), and was either kept at 40 % to impose fully aerobic conditions, or at 2 % to impose micro-aerobic conditions. [\[Source\]](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4559361/#!po=77.0833) * S. cerevisiae: engineered pathway (Xylose isomerase and ketohexokinase): Washed seed cultures were inoculate at an initial OD600 of 10–20 in 150 mL serum flasks containing 40 mL of media. The flasks were closed with butyl rubber stoppers, sealed with aluminum crimps, and purged with nitrogen gas to obtain strict anaerobic fermentations. The fermentation media contained oMM and 0.1 M 2-(N-morpholino)ethanesulfonic acid (MES), pH 6.0 supplemented with 40 g/L xylose, and/or 80 g/L cellobiose. The flasks were incubated at 30°C, 220 rpm. Art. [#ARTNUM](#article-25156-2462386339) * S. serevisiae: engineered pathway (oxidative D-xylose): 14 mg/L: Cultivations were carried out in 250-mL Erlenmeyer flasks, at 250 rpm, 30 °C. The media were supplemented with 10 g/L D-glucose, 20 g/L D-xylose and 10 g/L CaCO3. Art. [#ARTNUM](#article-25156-2765987778) **Titers up to 70 g/L with 1.38 g/L/h productivity and 0.4 g/g yield have been reported.** [\[Source\]](http://english.cas.cn/newsroom/research_news/201805/t20180504_192186.shtml) Current production: * from petrochemicals (Dow, Shell etc.) * from Biomass by catalysis: Greencol Bio-MEG from bio-ethanol, from industrial sugars by avantium, from industrial sugars or ethanol by M&G * Biological: no known industrial-scale process,

1.1.1 Ethylene glycol
Biosynthesis of ethylene glycol from d-xylose in recombinant Escherichia coli
ABSTRACTEthylene glycol (EG) is an important chemical used as antifreeze and a raw material in polyester synthesis. The EG biosynthetic pathway from D-xylose with D-xylonate as key intermediate has some advantages, but showed low EG production. Here, we reconstructed and optimized this pathway in Escherichia coli. In view of the greater intracellular prevalence of NADH, an aldehyde reductase FucO using NADH was employed to convert glycoaldehyde into EG, in replacement of NADPH-dependent reductase YqhD. To suppress the accumulation of by-products acetate and glycolate, two genes arcA and aldA were knocked out. The resultant strain Q2843 produced 72 g/L EG under fed-batch fermentation conditions, with the yield of 0.40 g/g D-xylose and EG productivity of 1.38 g/L/h. The use of NADH-dependent enzyme FucO and by-product elimination significantly improved the performance of EG producing strain, which represented the highest titer, yield and productivity of EG reported so far.
1/1/18 12:00:00 AM
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1.1.2 Ethylene glycol
Biotechnological production of glycolic acid and ethylene glycol: current state and perspectives
Glycolic acid (GA) and ethylene glycol (EG) are versatile two-carbon organic chemicals used in multiple daily applications. GA and EG are currently produced by chemical synthesis, but their biotechnological production from renewable resources has received a substantial interest. Several different metabolic pathways by using genetically modified microorganisms, such as Escherichia coli, Corynebacterium glutamicum and yeast have been established for their production. As a result, the yield of GA and EG produced from sugars has been significantly improved. Here, we describe the recent advancement in metabolic engineering efforts focusing on metabolic pathways and engineering strategies used for GA and EG production.
2/1/19 12:00:00 AM
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1.1.3 Ethylene glycol
Bypassing the Pentose Phosphate Pathway: Towards Modular Utilization of Xylose.
The efficient use of hemicellulose in the plant cell wall is critical for the economic conversion of plant biomass to renewable fuels and chemicals. Previously, the yeast Saccharomyces cerevisiae has been engineered to convert the hemicellulose-derived pentose sugars xylose and arabinose to d-xylulose-5-phosphate for conversion via the pentose phosphate pathway (PPP). However, efficient pentose utilization requires PPP optimization and may interfere with its roles in NADPH and pentose production. Here, we developed an alternative xylose utilization pathway that largely bypasses the PPP. In the new pathway, d-xylulose is converted to d-xylulose-1-phosphate, a novel metabolite to S. cerevisiae, which is then cleaved to glycolaldehyde and dihydroxyacetone phosphate. This synthetic pathway served as a platform for the biosynthesis of ethanol and ethylene glycol. The use of d-xylulose-1-phosphate as an entry point for xylose metabolism opens the way for optimizing chemical conversion of pentose sugars in S. cerevisiae in a modular fashion.
6/23/16 12:00:00 AM
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1.1.4 Ethylene glycol
Efficient utilization of pentoses for bioproduction of the renewable two-carbon compounds ethylene glycol and glycolate.
Abstract The development of lignocellulose as a sustainable resource for the production of fuels and chemicals will rely on technology capable of converting the raw materials into useful compounds; some such transformations can be achieved by biological processes employing engineered microorganisms. Towards the goal of valorizing the hemicellulose fraction of lignocellulose, we designed and validated a set of pathways that enable efficient utilization of pentoses for the biosynthesis of notable two-carbon products. These pathways were incorporated into Escherichia coli , and engineered strains produced ethylene glycol from various pentoses, including simultaneously from D -xylose and L -arabinose; one strain achieved the greatest reported titer of ethylene glycol, 40 g/L, from D -xylose at a yield of 0.35 g/g. The strategy was then extended to another compound, glycolate. Using D -xylose as the substrate, an engineered strain produced 40 g/L glycolate at a yield of 0.63 g/g, which is the greatest reported yield to date.
3/1/16 12:00:00 AM
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1.1.5 Ethylene glycol
Enhanced yield of ethylene glycol production from d-xylose by pathway optimization in Escherichia coli
Abstract The microbial production of renewable ethylene glycol (EG) has been gaining attention recently due to its growing importance in chemical and polymer industries. EG has been successfully produced biosynthetically from d -xylose through several novel pathways. The first report on EG biosynthesis employed the Dahms pathway in Escherichia coli wherein 71% of the theoretical yield was achieved. This report further improved the EG yield by implementing metabolic engineering strategies. First, d -xylonic acid accumulation was reduced by employing a weak promoter which provided a tighter control over Xdh expression. Second, EG yield was further improved by expressing the YjgB, which was identified as the most suitable aldehyde reductase endogenous to E. coli . Finally, cellular growth, d -xylose consumption, and EG yield were further increased by blocking a competing reaction. The final strain (WTXB) was able to reach up to 98% of the theoretical yield (25% higher as compared to the first study), the highest reported value for EG production from d -xylose.
2/1/17 12:00:00 AM
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1.1.6 Ethylene glycol
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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1.1.7 Ethylene glycol
Production of ethylene glycol from xylose by metabolically engineered Escherichia coli
Ethylene glycol (EG) is an important chemical used for several industrial applications including poly(ethylene terephthalate) synthesis. In this study, Escherichia coli was metabolically engineered to efficiently produce EG from xylose. To biosynthesize EG, the Dahms pathway was introduced by expressing xylBC genes from Caulobacter crescentus (xylBC). Various E. coli strains and glycolaldehyde reductases were screened to find E. coli W3110 strain and glycolaldehyde reductase (yqhD) as optimal combination for EG production. In silico genome-scale metabolic simulation suggested that increasing the native xylose pathway flux, in the presence of the overexpressed Dahms pathway, is beneficial for EG production. This was achieved by reducing the Dahms pathway flux by employing a synthetic small regulatory RNA targeting xylB. Fed-batch culture of the final engineered E. coli strain produced 108.2g/L of EG in a xylose minimal medium. The yield on xylose and EG productivity were 0.36g/g (0.87mol/mol) and 2.25g/L/h, respectively.
12/1/18 12:00:00 AM
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1.1.8 Ethylene glycol
Production of ethylene glycol or glycolic acid from D-xylose in Saccharomyces cerevisiae
The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative D-xylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding D-xylose dehydrogenase (XylB) and D-xylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from D-xylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2-keto-3-deoxyxylonic acid [(S)-4,5-dihydroxy-2-oxopentanoic acid] (2K3DXA) was produced and D-xylonic acid accumulated to ca. 9 g/L. A significant amount of D-xylonic acid (ca. 14%) was converted to 3-deoxypentonic acid (3DPA), and also, 3,4-dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldo-keto reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. The D-xylonate dehydratase activity in yeast was notably low, possibly due to inefficient Fe–S cluster synthesis in the yeast cytosol, and leading to D-xylonic acid accumulation. The dehydratase activity was significantly improved by targeting its expression to mitochondria or by altering the Fe–S cluster metabolism of the cells with FRA2 deletion.
11/1/17 12:00:00 AM
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1.1.9 Ethylene glycol
Recombinant bacterium capable of efficiently converting xylose to produce ethylene glycol and application of recombinant bacteria
The invention discloses a recombinant bacterium capable of efficiently converting xylose to produce ethylene glycol and an application of the recombinant bacterium and belongs to the technical field of genetic engineering. The provided recombinant bacterium is used for overexpressing a xylose dehydrogenase gene, a xylosic acid lactonase gene, a xylosic acid anhydrase gene, a 3-deoxy-D-glycerin ketopentos threonine aldolase gene and a glycolic aldehyde dehydrogenase gene. A metabolic pathway for synthesizing ethylene glycol from the beginning by employing xylose as a raw material is built in the recombinant bacterium. Meanwhile, the invention further provides a construction method and application method of the recombinant bacterium. Synthesis of the ethylene glycol by firstly using the pathway in a type strain, namely escherichia coli employing the xylose as a substrate is achieved through E.coli host bacterium, and a novel technique is provided for production of the ethylene glycol.
11/24/17 12:00:00 AM
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1.1.10 Ethylene glycol
Simultaneous biosynthesis of (R)-acetoin and ethylene glycol from D-xylose through in vitro metabolic engineering
Abstract ( R )-acetoin is a four-carbon platform compound used as the precursor for synthesizing novel optically active materials. Ethylene glycol (EG) is a large-volume two-carbon commodity chemical used as the anti-freezing agent and building-block molecule for various polymers. Currently established microbial fermentation processes for converting monosaccharides to either ( R )-acetoin or EG are plagued by the formation of undesirable by-products. We show here that a cell-free bioreaction scheme can generate enantiomerically pure acetoin and EG as co-products from biomass-derived D -xylose. The seven-step, ATP-free system included in situ cofactor regeneration and recruited enzymes from Escherichia coli W3110, Bacillus subtilis shaijiu 32 and Caulobacter crescentus CB 2. Optimized in vitro biocatalytic conditions generated 3.2 mM ( R )-acetoin with stereoisomeric purity of 99.5% from 10 mM D -xylose at 30 °C and pH 7.5 after 24 h, with an initial ( R )-acetoin productivity of 1.0 mM/h. Concomitantly, EG was produced at 5.5 mM, with an initial productivity of 1.7 mM/h. This in vitro biocatalytic platform illustrates the potential for production of multiple value-added biomolecules from biomass-based sugars with no ATP requirement.
12/1/18 12:00:00 AM
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1.2 2,3-butanediol

1

2,3-Butanediol is the organic compound with the formula (CH3CHOH)2. It is classified as a vic-diol (glycol). It exists as three stereoisomers, a chiral pair and the meso isomer. All are colourless liquids. Applications include precursors to various plastics and pesticides. [\[Wiki\]](https://en.wikipedia.org/wiki/2,3-Butanediol) 2,3-Butanediol (2,3-BD) is a platform chemical that can be converted into a variety of chemicals through dehydrogenation, ketalization, esterification, and dehydration. Chemicals such as butadiene, acetoin, diacetyls, tetramethyls, butanediesters. **Process and conditions:** * S. cerevisiae engineered: 96.8 g/L 2,3-BDO and 0.58 g/L-h productivity from xylose. Batch fermentation was carried out in a 250 mL flask containing 50 mL of YP medium (10 g/L of yeast extract and 20 g/L of Bacto peptone) with 40 g/L or 80 g/L xylose and 1.5 g/L ethanol at 30 °C and 80 rpm for microaerobic condition. Fed-batch fermentation was performed in a 1 L-bench-top bioreactor (Fermentec, Korea) containing 500 mL of YP medium with 80 g/L xylose at 30 °C. During fed-batch cultivation, acidity was controlled at pH 5.5 by adding 2 N HCl and 2 N NaOH. Agitation speed and aeration were maintained at 200 rpm and 0.5 vvm, respectively. Art. [#ARTNUM](#article-25171-2622828466) * Raoultella ornithinolytica B6, using all kinds of sugars: titer: 112.19 g/L), yield 0.38 g/g, and productivity 1.35 g/L/h. Art. [#ARTNUM](#article-25171-2535981131) * engineered Bacillus licheniformis: 32.2 g/L d-2,3-BD for WX-02ΔbudC and 48.5 g/L meso-2,3-BD for WX-02ΔgldA. (per liter) 33 g corn steep liquor, 9.00 g (NH4)2SO4, 1.00 g K2HPO4, 1.50 g MgSO4, 0.50 g NaCl, 0.12 g ZnCl2, 1 mg FeCl3, and 1 mg MnSO4, with a pH value of 7.0. Xylose was added into basic medium to a concentration of 80 g/L, then used for culture of B. licheniformis WX-02ΔbudC and WX-02ΔgldA, respectively. The cultures were shaken at 37 °C and 110 rpm on a rotary shaker. Miscanthus floridulus hydrolysate medium was prepared on the basic medium after addition with M. floridulus hydrolysate containing 80 g/L total reducing sugars (48.2 g/L glucose and 31.8 g/L xylose, glucose:xylose = 3.1:2). * Batch fermentation was also conducted in a 5-L stirred bioreactor with 2 L M. floridulus hydrolysate medium. The pH value was maintained at 6.0 by automatic addition of 4 M H3PO4 or 6 M KOH using a program-controlled peristaltic pump. The agitation speed was set up at 400 rpm for 24 h, then adjusted to 200 rpm until the end. The aeration rate of 1.0 vvm was changed to 0.5 vvm after the exponential growth phase. * Fed-batch fermentation was conducted in a 5-L stirred bioreactor with 2 L M. floridulus hydrolysate medium as described above. The pH value was maintained at 6.0 by automatic addition of 4 M H3PO4 or 6 M KOH using a program-controlled peristaltic pump. The agitation speed was set up at 400 rpm for 24 h, then adjusted to 200 rpm until the end. The aeration rate of 1.0 vvm was changed to 0.5 vvm after the exponential growth phase. When sugars concentration was lower than 20 g/L, M. floridulus hydrolysate was fed into the bioreactor to maintain the total reducing sugars concentration at 40 g/L. **Current production:** * Synthetic from petrochemicals * Syntheitic from bio-based * Microbial from glucose/sucrose, sometimes hydrolysates/lignocellulose products is being developed and marketed (research started in WWII, as well as pilots: discontinued then). **Purification from fermentation broths:** Purification has been researched. One cost-effective option is through hybrid extraction distillation. [#ARTNUM](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5785907/)

1.2.1 2,3-butanediol
2,3-Butanediol Production Using Xylose Mother Liquor
Four 2,3-butanediol producing strains were compared to utilize xylose,in which Klebsiella pneumoniae CICC 10011 was the best,with which as the starting strain xylose mother liquor was used to produce 2,3-butanediol.Firstly,single-factor experiments were carried out to study the effects of xylose mother liquor and nitrogen source.The fermentation medium was then optimized by orthogonal design.The optimized composition was as follows: xylose mother liquor 90 g/L,CSLP 12 g/L,K2HPO4 7 g/L,KH2PO4 2 g/L,(NH4)2SO4 2 g/L,sodium citrate 3 g/L,MgSO4·7H2O 0.1 g/L,FeSO4·7H2O 0.005 g/L,MnSO4·7H2O 0.005 g/L,ZnSO4·7H2O 0.01 g/L.Under above conditions,final concentration of 2,3-butanediol reached 35.7 g/L,7.5 g/L higher than that under the initial conditions,the yield was 92 % of the theory.
1/1/12 12:00:00 AM
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1.2.2 2,3-butanediol
Elimination of Carbon Catabolite Repression in Bacillus subtilis for the Improvement of 2,3-Butanediol Production
2,3-butanediol is a vital platform compound, extensively used as liquid fuel and chemical raw material. In this study, Bacillus subtilis was engineered to utilize glucose and xylose for 2,3-butanediol production. Initially, the gene araE from B. subtilis, encoding the xylose transport protein AraE, was overexpressed under the control of the constitutive Pspac promoter. Subsequently, the xylose isomerase and xylulose kinase from Escherichia coli, encoded by the genes of xylA and xylB respectively, were introduced into B. subtilis genome. In mineral medium, the engineered strain BSUL02 is able to utilize d-glucose and D-xylose simultaneously to produce 2,3-butanediol. Under the fermentation conditions tested in this work, the recombinant strain BSUL02 could produce 3.1 g/L 2,3-butanediol from 10 g/L d-glucose and 5 g/L D-xylose, which sheds new light on a metabolic engineering strategy for commercial exploitation of lignocellulose to produce important building-block chemicals.
1/1/14 12:00:00 AM
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1.2.3 2,3-butanediol
Enhanced production of 2,3-butanediol from xylose by combinatorial engineering of xylose metabolic pathway and cofactor regeneration in pyruvate decarboxylase-deficient Saccharomyces cerevisiae
Abstract The aim of this study was to produce 2,3-butanediol (2,3-BDO) from xylose efficiently by modulation of the xylose metabolic pathway in engineered Saccharomyces cerevisiae . Expression of the Scheffersomyces stipitis transaldolase and NADH-preferring xylose reductase in S. cerevisiae improved xylose consumption rate by a 2.1-fold and 2,3-BDO productivity by a 1.8-fold. Expression of the Lactococcus lactis noxE gene encoding NADH oxidase also increased 2,3-BDO yield by decreasing glycerol accumulation. Additionally, the disadvantage of C 2 -dependent growth of pyruvate decarboxylase-deficient (Pdc − ) S. cerevisiae was overcome by expression of the Candida tropicalis PDC1 gene. A fed-batch fermentation of the BD5X-TXmNP strain resulted in 96.8 g/L 2,3-BDO and 0.58 g/L-h productivity from xylose, which were 15.6- and 2-fold increases compared with the corresponding values of the BD5X strain. It was concluded that facilitation of the xylose metabolic pathway, oxidation of NADH and relief of C 2 -dependency synergistically triggered 2,3-BDO production from xylose in Pdc − S. cerevisiae .
12/1/17 12:00:00 AM
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1.2.4 2,3-butanediol
Evaluation of oil palm front hydrolysate as a novel substrate for 2,3-butanediol production using a novel isolate Enterobacter cloacae SG1
The present work deals the production of 2,3-butanediol, an industrially important chemical, through biological route using a novel bacterial isolate. Batch fermentation trials for the production of 2,3-butanediol were carried out using the isolated strain Enterobacter cloacae SG-1. The study resulted 14.67 g/l of 2,3-butanediol with 48.9% yield using glucose as the carbon source. In order to replace the expensive glucose in the production media, non-detoxified oil palm frond hydrolysate was used as the carbon source and it resulted 2,3-butanediol yield of 7.67 g/l. Process parameters like pH, temperature and initial sugar concentration were optimized. The ability of strain E. cloacae SG-1 for utilization various pentoses and hexoses were evaluated and found that the strain can utilize both arabinose and glucose with a comparable 2,3-butanediol yield.
12/1/16 12:00:00 AM
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1.2.5 2,3-butanediol
High Production of 2,3-Butanediol (2,3-BD) by Raoultella ornithinolytica B6 via Optimizing Fermentation Conditions and Overexpressing 2,3-BD Synthesis Genes
Biological production of 2,3-butandiol (2,3-BD) has received great attention as an alternative to the petroleum-based 2,3-BD production. In this study, a high production of 2,3-BD in fed-batch fermentation was investigated with a newly isolated bacterium designated as Raoultella ornithinolytica B6. The isolate produced 2,3-BD as the main product using hexoses (glucose, galactose, and fructose), pentose (xylose) and disaccharide (sucrose). The effects of temperature, pH-control schemes, and agitation speeds on 2,3-BD production were explored to optimize the fermentation conditions. Notably, cell growth and 2,3-BD production by R. ornithinolytica B6 were higher at 25°C than at 30°C. When three pH control schemes (no pH control, pH control at 7, and pH control at 5.5 after the pH was decreased to 5.5 during fermentation) were tested, the best 2,3-BD titer and productivity along with reduced by-product formation were achieved with pH control at 5.5. Among different agitation speeds (300, 400, and 500 rpm), the optimum agitation speed was 400 rpm with 2,3-BD titer of 68.27 g/L, but acetic acid was accumulated up to 23.32 g/L. Further enhancement of the 2,3-BD titer (112.19 g/L), yield (0.38 g/g), and productivity (1.35 g/L/h) as well as a significant reduction of acetic acid accumulation (9.71 g/L) was achieved by the overexpression of homologous budABC genes, the 2,3-BD-synthesis genes involved in the conversion of pyruvate to 2,3-BD. This is the first report presenting a high 2,3-BD production by R.ornithinolytica which has attracted little attention with respect to 2,3-BD production, extending the microbial spectrum of 2,3-BD producers.
10/19/16 12:00:00 AM
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1.2.6 2,3-butanediol
Production of 2,3-butanediol from corncob molasses, a waste by-product in xylitol production.
Corncob molasses, a waste by-product in xylitol production, contains high concentrations of mixed sugars. In the present study, corncob molasses was used to produce 2,3-butanediol (BD) using Klebsiella pneumoniae SDM. This was the first report on the use of corncob molasses to produce bulk chemicals. Our results indicated that K. pneumoniae SDM can utilize various sugars contained in the corncob molasses in a preferential manner: glucose > arabinose > xylose. It was shown that high sugars concentration had an inhibitory effect on the cells growth and BD production. The maximum concentration of BD was 78.9 g/l after 61 h of fed-batch fermentation, giving a BD productivity of 1.3 g/l h and a yield of 81.4%. The present study suggests that the low-cost corncob molasses could be used as an alternative substrate for the production of BD by K. pneumoniae SDM, as well as a potential carbon source for production of other high-value chemicals.
7/1/10 12:00:00 AM
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1.2.7 2,3-butanediol
Production of 2,3-butanediol from xylose by engineered Saccharomyces cerevisiae
Abstract 2,3-Butanediol (2,3-BD) production from xylose that is abundant in lignocellulosic hydrolyzate would make the production of 2,3-BD more sustainable and economical. Saccharomyces cerevisiae can produce only trace amounts of 2,3-BD, but also cannot ferment xylose. Therefore, it is necessary to introduce both 2,3-BD production and xylose assimilation pathways into S. cerevisiae for producing 2,3-BD from xylose. A pyruvate decarboxylase (Pdc)-deficient mutant (SOS4) was used as a host in order to increase carbon flux toward 2,3-BD instead of ethanol. The XYL1 , XYL2 , and XYL3 genes coding for xylose assimilating enzymes derived from Scheffersomyces stipitis were introduced into the SOS4 strain to enable xylose utilization. Additionally, the alsS and alsD genes from Bacillus subtilis and endogenous BDH1 gene were overexpressed to increase 2,3-BD production from xylose. As a result, the resulting strain (BD4X) produced 20.7 g/L of 2,3-BD from xylose with a yield of 0.27 g 2,3-BD/g xylose. The titer of 2,3-BD from xylose increased up to 43.6 g/L under a fed-batch fermentation. The BD4X strain produced ( R , R )-2,3-BD dominantly (>97% of the total 2,3-BD) with trace amounts of meso -2,3-BD. These results suggest that S. cerevisiae might be a promising host for producing 2,3-BD from lignocellulosic biomass for industrial applications.
12/1/14 12:00:00 AM
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1.2.8 2,3-butanediol
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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1.2.9 2,3-butanediol
Production of optically pure 2,3-butanediol from Miscanthus floridulus hydrolysate using engineered Bacillus licheniformis strains
2,3-Butanediol (2,3-BD) can be produced by fermentation of natural resources like Miscanthus. Bacillus licheniformis mutants, WX-02ΔbudC and WX-02ΔgldA, were elucidated for the potential to use Miscanthus as a cost-effective biomass to produce optically pure 2,3-BD. Both WX-02ΔbudC and WX-02ΔgldA could efficiently use xylose as well as mixed sugars of glucose and xylose to produce optically pure 2,3-BD. Batch fermentation of M. floridulus hydrolysate could produce 21.6 g/L d-2,3-BD and 23.9 g/L meso-2,3-BD in flask, and 13.8 g/L d-2,3-BD and 13.2 g/L meso-2,3-BD in bioreactor for WX-02ΔbudC and WX-02ΔgldA, respectively. Further fed-batch fermentation of hydrolysate in bioreactor showed both of two strains could produce optically pure 2,3-BD, with 32.2 g/L d-2,3-BD for WX-02ΔbudC and 48.5 g/L meso-2,3-BD for WX-02ΔgldA, respectively. Collectively, WX-02ΔbudC and WX-02ΔgldA can efficiently produce optically pure 2,3-BD with M. floridulus hydrolysate, and these two strains are candidates for industrial production of optical purity of 2,3-BD with M. floridulus hydrolysate.
5/1/18 12:00:00 AM
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1.3 Xylitol

1

Xylitol is a sugar alcohol used as a sugar substitute. Xylitol is categorized as a polyalcohol or sugar alcohol (specifically an alditol). It has the formula CH2OH(CHOH)3CH2OH. It is a colorless or white solid that is soluble in water. [Wiki](https://en.wikipedia.org/wiki/Xylitol#Uses) Xylitol can be produced from xylose and arabinose Art. [#ARTNUM](#article-25166-1974978885) Art. [#ARTNUM](#article-25166-2120706216) **Applications:** Xylitol can also be converted to lactic acid, ethanol, ethylene and propylene glycol. Art. [#ARTNUM](#article-25166-1990883223); [#ARTNUM](#article-25166-2067211519); [#ARTNUM](#article-25166-2129489681) **Process:** * K. marxianus YZJ017 engineered: 99.29 and 149.60 g/L xylitol were produced from 99.55 and 151.91 g/L xylose with productivity of 4.14 and 3.40 g/L/h respectively, Using fed-batch fermentation through repeatedly adding non-sterilized substrate directly, YZJ074 could produce 312.05 g/L xylitol. YPXG (yeast extract 10 g/L, bacteriological peptone 20 g/L, xylose 150 g/L and glycerol 60 g/L) was used as fermentation media. 1-L modular benchtop fermenter (BioFlo 110, New Brunswick Scientific, Edison, New Jersey, USA) containing 0.5 L of fermentation medium, The fermentation temperature was maintained at 42 or 45 °C. Art. [#ARTNUM](#article-25166-2078444927) * Candida tropicalis: Approximately 95 g/L of pure xylitol could be obtained from the medium containing 400 g/L of waste xylose mother liquor (WXML) at a yield of 0.75 g/g xylose consumed. Batch biotransformation of WXML in bioreactors was carried out in a BIOSTAT® Aplus 5 L microbial bioreactor (Sartorius) containing 3 L of YCN (contained 5 g/L yeast extract (AngelYeast Co. Ltd, China), 5 g/L corn syrup powder, 2 g/L (NH4)2HPO4 and 0.5 g/L MgSO4·7H2O (pH 6.0)) medium supplemented with 400 g/L WXML. Seed cultures from the 30 h culture of yeast and the 24 h culture of B. subtilis were inoculated into the bioreactor with 5.0 % (v/v) inoculum. The medium pH was controlled at 6.0 using 25 % NaOH solution and 15 g/L of glucose was supplemented following the exhaustion of initial glucose from WXML. At the stage of micro-aerobic biotransformation, the conditions were set as temperature 30 °C, aeration 0.3 VVM and agitation 200 rpm. At the completion of biotransformation, the conditions were set as temperature 39 °C, aeration 1 VVM and agitation 300 rpm to allow for aerobic biotransformation. In the case of scaled-up experiments, the batch biotransformation was performed in 150 L and 30 m3 bioreactors following the same conditions as in the 5 L bioreactor with some modifications: the filling volume was 100 L and 20 m3, respectively; at the stage of micro-aerobic biotransformation, the aeration was set at 0.2 VVM and agitation at 60 rpm, and at the stage of aerobic transformation the parameters were 0.5 VVM and 100 rpm. Art. [#ARTNUM](#article-25166-2554952832) **Current production:** * Synthetic: chemical reduction of xylan and xylose. (uneconomical and difficult purification). Art. [#ARTNUM](#article-25166-2945082312) * Biological: mainly purification can be a bottleneck. Much research is done on this topic, there are no commercial systems.

1.3.1 Xylitol
Bio-production of a Polyalcohol (Xylitol) from Lignocellulosic Resources: A Review
Lignocellulosic materials being supplied from a variety of resources at low price can be used as feedstock for chemicals and bio-products. Xylitol is a high value polyalcohol produced by the reduction of D-xylose (from hemicellulose fraction of lignocellulose) and is employed in food and pharmaceutical industries. The large number of advantageous properties, such as its low-calorie sweetening power and anticariogenicity justifies the high industrial interest for xylitol. Biotechnological production of this substance is lately becoming more attractive than the chemical method of catalytic hydrogenation due to the higher yield and because downstream processing is expected to be less costly. Studies about the bio-production of xylitol, in which microorganisms or enzymes are involved as catalysts to convert xylose into xylitol under mild conditions of pressure and temperature, have been mostly focused on establishing the operational parameters and the process options that maximize its yield and productivity in free cell system. However, some gaps in knowledge exist regarding this bioconversion process in immobilized cell system and selection or making an appropriate carrier (support) for biocatalysts in fermentation medium.
1/1/06 12:00:00 AM
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1.3.2 Xylitol
Biosynthetic strategies to produce xylitol: an economical venture
Xylitol is a natural five-carbon sugar alcohol with potential for use in food and pharmaceutical industries owing to its insulin-independent metabolic regulation, tooth rehardening, anti-carcinogenic, and anti-inflammatory, as well as osteoporosis and ear infections preventing activities. Chemical and biosynthetic routes using D-xylose, glucose, or biomass hydrolysate as raw materials can produce xylitol. Among these methods, microbial production of xylitol has received significant attention due to its wide substrate availability, easy to operate, and eco-friendly nature, in contrast with high-energy consuming and environmental-polluting chemical method. Though great advances have been made in recent years for the biosynthesis of xylitol from xylose, glucose, and biomass hydrolysate, and the yield and productivity of xylitol are substantially improved by metabolic engineering and optimizing key metabolic pathway parameters, it is still far away from industrial-scale biosynthesis of xylitol. In contrary, the chemical synthesis of xylitol from xylose remains the dominant route. Economic and highly efficient xylitol biosynthetic strategies from an abundantly available raw material (i.e., glucose) by engineered microorganisms are on the hard way to forwarding. However, synthetic biology appears as a novel and promising approach to develop a super yeast strain for industrial production of xylitol from glucose. After a brief overview of chemical-based xylitol production, we critically analyzed and comprehensively summarized the major metabolic strategies used for the enhanced biosynthesis of xylitol in this review. Towards the end, the study is wrapped up with current challenges, concluding remarks, and future prospects for designing an industrial yeast strain for xylitol biosynthesis from glucose.
5/17/19 12:00:00 AM
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1.3.3 Xylitol
Efficient xylitol production from cornstalk hydrolysate using engineered Escherichia coli whole cells
Economic transformation of lignocellulose hydrolysate into valued-added products is of particular importance for energy and environmental issues. In this study, xylose reductase and glucose dehydrogenase were cloned into plasmid pETDuet-1 and then simultaneously expressed in Escherichia coli BL21(DE3), which was used as whole-cell catalyst for the first time to convert xylose into xylitol coupled with gluconate production. When tested with reconstituted xylose and glucose solution, 0.1 g/mL cells could convert 1 M xylose and 1 M glucose completely and produced 145.81 g/L xylitol with a yield of 0.97 (g/g) and 184.85 g/L gluconic acid with a yield of 1.03 (g/g) in 24 hours. Subsequently, the engineered cells were applied in real cornstalk hydrolysate, which generated 30.88 g/L xylitol and 50.89 g/L gluconic acid. The cells were used without penetration treatment and CaCO3 was used to effectively regulate the pH during the production, which further saved costs.
12/19/18 12:00:00 AM
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1.3.4 Xylitol
Engineering xylose and arabinose metabolism in recombinant Saccharomyces cerevisiae
Utilization of all sugars in lignocellulose hydrolysates is a prerequisite for economically feasible bioethanol production. The yeast commonly used for industrial ethanol production, Saccharomyces cerevisiae, is naturally unable to utilize pentose sugars xylose and arabinose, which constitute a large fraction of many lignocellulosic materials. Xylose utilization by S. cerevisiae can be achieved by heterologous expression of a xylose utilization pathway, consisting either of xylose reductase (XR), xylitol dehydrogenase (XDH) and xylulokinase (XK), or alternatively, of xylose isomerase (XI) and XK. Xylitol formed by XR is a major by-product in xylose fermentation when using the XR-XDH pathway. In this thesis, high-level expression of both XR and XDH was shown to decrease xylitol formation. The influence of other genetic modifications was also evaluated. It was shown that the overexpression of the non-oxidative pentose phosphate pathway (PPP) genes enables efficient growth on xylose and xylose fermentation, provided that the initial xylose pathway is expressed at a high level. When comparing the two xylose utilization pathways, higher ethanol productivity was achieved using the XR-XDH pathway, whereas higher ethanol yield was achieved with the XI pathway. The industrial xylose-fermenting S. cerevisiae strain TMB 3400, which has been previously generated by mutagenesis and selection, was tested for fermentation of lignocellulose hydrolysate. TMB 3400 displayed significantly better fermentation performance compared to the laboratory strains tested, highlighting the need for robust industrial strains in lignocellulose fermentation. TMB 3400 was also characterized by proteome analysis using difference in-gel 2-D electrophoresis. Consistently with the results obtained in other studies, increased activities of XR, XDH and a PPP enzyme TKL were found. The bacterial arabinose utilization pathway was introduced into TMB 3400, which resulted in the novel glucose, xylose and arabinose co-fermenting strain TMB 3063, with ethanol, xylitol and arabitol as the main fermentation products. (Less)
1/1/06 12:00:00 AM
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1.3.5 Xylitol
Improving xylitol production at elevated temperature with engineered Kluyveromyces marxianus through over-expressing transporters
Abstract Three transporter genes including Kluyveromyces marxianus aquaglyceroporin gene ( KmFPS1 ), Candida intermedia glucose/xylose facilitator gene ( CiGXF1) or glucose/xylose symporter gene ( CiGXS1 ) were over-expressed in K. marxianus YZJ017 to improve xylitol production at elevated temperatures. The xylitol production of YZJ074 that harbored CiGXF1 was improved to 147.62 g/L in Erlenmeyer flask at 42 °C. In fermenter, 99.29 and 149.60 g/L xylitol were produced from 99.55 and 151.91 g/L xylose with productivity of 4.14 and 3.40 g/L/h respectively at 42 °C. Even at 45 °C, YZJ074 could produce 101.30 g/L xylitol from 101.41 g/L xylose with productivity of 2.81 g/L/h. Using fed-batch fermentation through repeatedly adding non-sterilized substrate directly, YZJ074 could produce 312.05 g/L xylitol which is the highest yield reported to date. The engineered strains YZJ074 which can produce xylitol at elevated temperatures is an excellent foundation for xylitol bioconversion.
1/1/15 12:00:00 AM
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1.3.6 Xylitol
Selective catalytic oxidation of sugar alcohols to lactic acid
Sorbitol and xylitol obtained from biomass are considered promising potential sources of both carbon building blocks and energy. We report the efficient and selective conversion of sorbitol, xylitol and other polyols into lactic acid as the major product through homogeneous iridium-NHC catalyzed dehydrogenative processes. The proposed reaction mechanism involves base-driven hydrolysis of simple sugars which accounts for the catalyst selectivity observed. In addition, catalyst deactivation pathways are explored and rational catalyst optimization is attempted through fine tuning of the complex.
1/1/15 12:00:00 AM
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1.3.7 Xylitol
Selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol on supported Ru catalysts
The selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol was carried out on different catalysts in the presence of Ca(OH)2. The catalysts included Ru supported on activated carbon (C) and, for comparison, on metal oxides, Al2O3, TiO2, ZrO2 and Mg2AlOx as well as C-supported other noble metals, Rh, Pd and Pt, with similar particle sizes (1.6–2.0 nm). The kinetic effects of H2 pressures (0–10 MPa), temperatures (433–513 K) and solid bases including Ca(OH)2, Mg(OH)2 and CaCO3 were examined on Ru/C. Ru/C exhibited superior activities and glycol selectivities than Ru on TiO2, ZrO2, Al2O3 and Mg2AlOx, and Pt was found to be the most active metal. Such effects of the metals and supports are attributed apparently to their different dehydrogenation/hydrogenation activities and surface acid-basicities, which consequently influenced the xylitol reaction pathways. The large dependencies of the activities and selectivities on the H2 pressures, reaction temperatures, and pH values showed their effects on the relative rates for the hydrogenation and base-catalyzed reactions involved in xylitol hydrogenolysis, reflecting the bifunctional nature of the xylitol reaction pathways. These results led to the proposition that xylitol hydrogenolysis to ethylene glycol and propylene glycol apparently involves kinetically relevant dehydrogenation of xylitol to xylose on the metal surfaces, and subsequent base-catalyzed retro-aldol condensation of xylose to form glycolaldehyde and glyceraldehyde, followed by direct glycolaldehyde hydrogenation to ethylene glycol and by sequential glyceraldehyde dehydration and hydrogenation to propylene glycol. Clearly, the relative rates between the hydrogenation of the aldehyde intermediates and their competitive reactions with the bases dictate the selectivities to the two glycols. This study provides directions towards efficient synthesis of the two glycols from not only xylitol, but also other lignocellulose-derived polyols, which can be achieved, for example, by optimizing the reaction parameters, as already shown by the observed effects of the catalysts, pH values, and H2 pressures.
1/1/11 12:00:00 AM
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1.3.8 Xylitol
Selective hydrogenolysis of xylitol to ethylene glycol and propylene glycol over copper catalysts
Abstract Cu SiO 2 catalysts were prepared by homogeneous deposition–precipitation with a wide range of Cu contents (8.8–100 wt%) and Cu particle sizes (2.1–111.1 nm). These catalysts were evaluated in the selective hydrogenolysis of biomass-derived xylitol to ethylene glycol and propylene glycol under the promotion of Ca(OH) 2 base. Their catalytic activity and selectivity to the two glycols depended strongly on the Cu particle sizes, which increased with the particle sizes and reached the maximum values at around 20–35 nm. Such size effects are apparently attributed to the effects on the dehydrogenation and hydrogenation activities of the Cu catalysts, and consequently on the xylitol hydrogenolysis pathways, reflecting the structural requirement for the xylitol hydrogenolysis. The effects of the reaction parameters including H 2 pressure (0–8.0 MPa), temperature (433–493 K) and pH values (7.0–12.4, adjusted by changing the amount of Ca(OH) 2 ) were examined. These effects confirmed the reaction pathways previously proposed for the xylitol hydrogenolysis to the two glycols, involving the dehydrogenation of xylitol to xylose on Cu as the rate-determining step, followed by the retro-aldol condensation of xylose with Ca(OH) 2 to glycolaldehyde and glyceraldehyde, and their subsequent hydrogenation to ultimately form glycols in competition with their side reactions to glycolic acid and lactic acid in the presence of Ca(OH) 2 . Upon optimizing the reaction conditions (473 K, 6.0 MPa H 2 and sufficient Ca(OH) 2 ), nearly 100% xylitol conversion and 54.4% combined selectivity to ethylene glycol and propylene glycol were obtained on Cu SiO 2 with Cu size of 35.7 nm, comparable to those on the previously reported Ni- and Ru- based catalysts. Clearly, this study provides directions for the design of more efficient Cu catalysts and the optimization of the reaction parameters toward the efficient polyol hydrogenolysis into glycols.
4/1/14 12:00:00 AM
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1.3.9 Xylitol
Selective reduction of xylose to xylitol from a mixture of hemicellulosic sugars.
Abstract The biocatalytic reduction of d -xylose to xylitol requires separation of the substrate from l -arabinose, another major component of hemicellulosic hydrolysate. This step is necessitated by the innate promiscuity of xylose reductases, which can efficiently reduce l -arabinose to l -arabinitol, an unwanted byproduct. Unfortunately, due to the epimeric nature of d -xylose and l -arabinose, separation can be difficult, leading to high production costs. To overcome this issue, we engineered an E. coli strain to efficiently produce xylitol from d -xylose with minimal production of l -arabinitol byproduct. By combining this strain with a previously engineered xylose reductase mutant, we were able to eliminate l -arabinitol formation and produce xylitol to near 100% purity from an equiweight mixture of d -xylose, l -arabinose, and d -glucose.
9/1/10 12:00:00 AM
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1.3.10 Xylitol
Xylitol production from waste xylose mother liquor containing miscellaneous sugars and inhibitors: one-pot biotransformation by Candida tropicalis and recombinant Bacillus
Background: The process of industrial xylitol production is a massive source of organic pollutants, such as waste xylose mother liquor (WXML), a viscous reddish-brown liquid. Currently, WXML is difficult to reuse due to its miscellaneous low-cost sugars, high content of inhibitors and complex composition. WXML, as an organic pollutant of hemicellulosic hydrolysates, accumulates and has become an issue of industrial concern in China. Previous studies have focused only on the catalysis of xylose in the hydrolysates into xylitol using one strain, without considering the removal of other miscellaneous sugars, thus creating an obstacle to subsequent large-scale purification. In the present study, we aimed to develop a simple one-pot biotransformation to produce high-purity xylitol from WXML to improve its economic value. Results: In the present study, we developed a procedure to produce xylitol from WXML, which combines detoxification, biotransformation and removal of by-product sugars (purification) in one bioreactor using two complementary strains, Candida tropicalis X828 and Bacillus subtilis Bs12. At the first stage of micro-aerobic biotransformation, the yeast cells were allowed to grow and metabolized glucose and the inhibitors furfural and hydroxymethyl furfural (HMF), and converted xylose into xylitol. At the second stage of aerobic biotransformation, B. subtilis Bs12 was activated and depleted the by-product sugars. The one-pot process was successfully scaled up from shake flasks to 5, 150 L and 30 m 3 bioreactors. Approximately 95 g/L of pure xylitol could be obtained from the medium containing 400 g/L of WXML at a yield of 0.75 g/g xylose consumed, and the by-product sugars glucose, l-arabinose and galactose were depleted simultaneously. Conclusions: Our results demonstrate that the one-pot procedure is a viable option for the industrial application of WXML to produce value-added chemicals. The integration of complementary strains in the biotransformation of hemicellulosic hydrolysates is efficient under optimized conditions. Moreover, our study of one-pot biotransformation also provides useful information on the combination of biotechnological processes for the biotransformation of other compounds.
1/1/16 12:00:00 AM
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1.3.11 Xylitol
Xylose fermentation efficiency of industrial Saccharomyces cerevisiae yeast with separate or combined xylose reductase/xylitol dehydrogenase and xylose isomerase pathways
Background Xylose isomerase (XI) and xylose reductase/xylitol dehydrogenase (XR/XDH) pathways have been extensively used to confer xylose assimilation capacity to Saccharomyces cerevisiae and tackle one of the major bottlenecks in the attainment of economically viable lignocellulosic ethanol production. Nevertheless, there is a lack of studies comparing the efficiency of those pathways both separately and combined. In this work, the XI and/or XR/XDH pathways were introduced into two robust industrial S. cerevisiae strains, evaluated in synthetic media and corn cob hemicellulosic hydrolysate and the results were correlated with the differential enzyme activities found in the xylose-pathway engineered strains.
1/28/19 12:00:00 AM
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2. Biological routes to organic acids

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2.1 Glycolic acid

1

Glycolic acid (hydroacetic acid or hydroxyacetic acid); chemical formula C2H4O3 (also written as HOCH2CO2H), is the smallest α-hydroxy acid (AHA). This colourless, odourless, and hygroscopic crystalline solid is highly soluble in water. [Wiki](https://en.wikipedia.org/wiki/Glycolic_acid) **Applications:** Glycolic acid is used in the textile industry as a dyeing and tanning agent, in food processing as a flavouring agent and as a preservative, and in the pharmaceutical industry as a skin care agent. It is also used in adhesives and plastics. Glycolic acid is often included into emulsion polymers, solvents and additives for ink and paint in order to improve flow properties and impart gloss. It is used in surface treatment products that increase the coefficient of friction on tile flooring. It is the active ingredient in the household cleaning liquid Pine-Sol. Glycolic acid is a useful intermediate for organic synthesis, in a range of reactions including: oxidation-reduction, esterification and long chain polymerization. It is used as a monomer in the preparation of polyglycolic acid (PGA) and other biocompatible copolymers (e.g. PLGA). Commercially, important derivatives include the methyl and ethyl esters which are readily distillable, unlike the parent acid. The butyl ester is a component of some varnishes, being desirable because it is nonvolatile and has good dissolving properties. [\[Wiki\]](https://en.wikipedia.org/wiki/Glycolic_acid) **Process:** * Yeast (Engineered S. cerevisiae & K. lactis): Biostat CT-DCU bioreactor (max. working volume 5000 ml, Sartorius, Göttingen, Germany) at pH 5.0, 30°C, 1 volume air \[volume culture\]-1 min-1 (vvm) and 500 rpm agitation with rushton turbines. The pH was maintained constant by addition of 2 M NaOH. Silicone antifoaming agent (BDH, 0.2 ml l-1) was added to prevent excess foaming. Batch medium was SC medium [\[SC medium\]](http://cshprotocols.cshlp.org/content/2016/11/pdb.rec090589.full?rss=1) lacking uracil with 20 g l-1D-xylose and 15 g l-1 ethanol. D-Xylose and ethanol were fed to the bioreactor as separate feeds; 30% ethanol and 4% D-xylose feed. Yield 32%, titer 15 g/L. Art. [#ARTNUM](#article-25190-2039878265) * Engineered E. coli: 40 g/L glycolate at a yield of 0.63 g/g. Bioreactor fermentations were carried out in a 3-L Bioflo culture vessel (New Brunswick, CT, USA) with a 2-L working volume. These fermentations utilized minimal medium (MM2) consisting of 2.0 g/L NH4Cl, 5.0 g/L (NH4)2SO4, 2.0 g/L KH2PO4, 0.5 g/L NaCl, 2 mL/L 1 M MgSO4, 1 mL/L mineral solution, 0.1 mL/L 4 mM Na2MoO4, and specified sugar; additionally, silicone antifoaming B emulsion was used to prevent foaming. Aerobic conditions were maintained by sparging air at 0.5 or 1 L/min, and the pH was maintained at 7.0 with 6 N NaOH. For all cultures, 50 mg/L spectinomycin, 34 mg/L chloramphenicol, and 50 mg/L kanamycin were added as appropriate. the initial D-xylose concentration was approximately 110 g/L for GA-03 and 65 g/L for GA-10 and GA-11. Temperature was maintained at 37 °C for strain GA-03 and 30 °C for strains GA-10 and GA-11. Dissolved oxygen content was maintained at 20% by altering agitation from 200 to 800 rpm. Art. [#ARTNUM](#article-25190-2214411608) **Current production:** * Synthetic: e.g. from formaldehyde and syngas. * Bio-based: isolated from several crops (e.g. sugar cane, beets) * Biological: from ethylene glycol, from glucose, xylose in research, Pilot from glucose. Art. [#ARTNUM](#article-25190-EP2027277B1) **Purification:** Several patents exist from Metabolic explorer and Roquette, no research papers. These contain multiple steps \~3.

2.1.1 Glycolic acid
Biotechnological production of glycolic acid and ethylene glycol: current state and perspectives
Glycolic acid (GA) and ethylene glycol (EG) are versatile two-carbon organic chemicals used in multiple daily applications. GA and EG are currently produced by chemical synthesis, but their biotechnological production from renewable resources has received a substantial interest. Several different metabolic pathways by using genetically modified microorganisms, such as Escherichia coli, Corynebacterium glutamicum and yeast have been established for their production. As a result, the yield of GA and EG produced from sugars has been significantly improved. Here, we describe the recent advancement in metabolic engineering efforts focusing on metabolic pathways and engineering strategies used for GA and EG production.
2/1/19 12:00:00 AM
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2.1.2 Glycolic acid
ESCHERICHIA COLI PRODUCING GLYCOLATE FROM XYLOSE METHOD FOR PREPARING THE SAME AND METHOD FOR PRODUCING GLYCOLATE USING THE SAME
The present invention relates to Escherichia coli having the ability to produce glycolic acid from xylose, a method for preparing the same, and a method for producing glycolic acid using the same. More specifically, the present invention relates to transformed Escherichia coli which can mass-produce glycolic acid from xylose, a method for preparing the same, and a method for producing glycolic acid from xylose using the transformed Escherichia coli. According to the present invention, transformed Escherichia coli which can mass-produce glycolic acid from xylose can be provided. Also, according to the present invention, a by-product generated in the process of producing glycolic acid from xylose can be introduced to the glycolic acid biosynthesis pathway, and thus the yield of by-products can be significantly reduced while producing glycolic acid in high yields.
11/27/18 12:00:00 AM
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2.1.3 Glycolic acid
Efficient utilization of pentoses for bioproduction of the renewable two-carbon compounds ethylene glycol and glycolate.
Abstract The development of lignocellulose as a sustainable resource for the production of fuels and chemicals will rely on technology capable of converting the raw materials into useful compounds; some such transformations can be achieved by biological processes employing engineered microorganisms. Towards the goal of valorizing the hemicellulose fraction of lignocellulose, we designed and validated a set of pathways that enable efficient utilization of pentoses for the biosynthesis of notable two-carbon products. These pathways were incorporated into Escherichia coli , and engineered strains produced ethylene glycol from various pentoses, including simultaneously from D -xylose and L -arabinose; one strain achieved the greatest reported titer of ethylene glycol, 40 g/L, from D -xylose at a yield of 0.35 g/g. The strategy was then extended to another compound, glycolate. Using D -xylose as the substrate, an engineered strain produced 40 g/L glycolate at a yield of 0.63 g/g, which is the greatest reported yield to date.
3/1/16 12:00:00 AM
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2.1.4 Glycolic acid
Engineering Escherichia coli for glycolic acid production from D-xylose through the Dahms pathway and glyoxylate bypass
Glycolic acid (GA) is an ⍺-hydroxy acid used in cosmetics, packaging, and medical industries due to its excellent properties, especially in its polymeric form. In this study, Escherichia coli was engineered to produce GA from D-xylose by linking the Dahms pathway, the glyoxylate bypass, and the partial reverse glyoxylate pathway (RGP). Initially, a GA-producing strain was constructed by disrupting the xylAB and glcD genes in the E. coli genome and overexpressing the xdh(Cc) from Caulobacter crescentus. This strain was further improved through modular optimization of the Dahms pathway and the glyoxylate bypass. Results for module 1 showed that the rate-limiting step of the Dahms pathway was the xylonate dehydratase reaction, and the overexpression of yagF was sufficient to overcome this bottleneck. Furthermore, the appropriate aldolase gene for module 1 was proven to be yagE. The results also show that overexpression of the lactaldehyde dehydrogenase gene, aldA, is needed to increase the GA production while the overexpression of glyoxylate reductase gene, ycdW, was only essential when the glyoxylate bypass was active. On the other hand, the module 2 enzymes AceA and AceK were vital in activating the glyoxylate bypass, while the RGP enzymes were dispensable. The final strain (GA19) produced 4.57 g/L GA with a yield of 0.46 g/g from D-xylose. So far, this is the highest value achieved for GA production in engineered E. coli through the Dahms pathway.
3/1/18 12:00:00 AM
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2.1.5 Glycolic acid
Glycolic acid production in the engineered yeasts Saccharomyces cerevisiae and Kluyveromyces lactis
Background Glycolic acid is a C2 hydroxy acid that is a widely used chemical compound. It can be polymerised to produce biodegradable polymers with excellent gas barrier properties. Currently, glycolic acid is produced in a chemical process using fossil resources and toxic chemicals. Biotechnological production of glycolic acid using renewable resources is a desirable alternative.
1/1/13 12:00:00 AM
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2.1.6 Glycolic acid
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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2.1.7 Glycolic acid
PRODUCTION OF ACID(S) AND ALCOHOL FROM SUGARS USING YEAST
The present invention relates to method of producing glycolic acid using the Dahms pathway, as well as to a microorganism, which is able to convert D-xylose derived from biomass to 2-keto-3-deoxy pentanoic acid, 3-deoxy pentonoic acid, glycolic acid, or concomitantly to glycolic and lactic acid, and to ethylene glycol. The starting material, pentose sugar D-xylose, is a major component in lignocellulosic hydrolysates and its efficient conversion to value-added products is essential in the context of biomass utilisation and cost-effective biorefineries. Further, the present invention also relates to a glycolic acid product and to a use of said micro-organism to produce such glycolic acid.
4/7/14 12:00:00 AM
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2.1.8 Glycolic acid
Production of ethylene glycol or glycolic acid from D-xylose in Saccharomyces cerevisiae
The important platform chemicals ethylene glycol and glycolic acid were produced via the oxidative D-xylose pathway in the yeast Saccharomyces cerevisiae. The expression of genes encoding D-xylose dehydrogenase (XylB) and D-xylonate dehydratase (XylD) from Caulobacter crescentus and YagE or YjhH aldolase and aldehyde dehydrogenase AldA from Escherichia coli enabled glycolic acid production from D-xylose up to 150 mg/L. In strains expressing only xylB and xylD, 29 mg/L 2-keto-3-deoxyxylonic acid [(S)-4,5-dihydroxy-2-oxopentanoic acid] (2K3DXA) was produced and D-xylonic acid accumulated to ca. 9 g/L. A significant amount of D-xylonic acid (ca. 14%) was converted to 3-deoxypentonic acid (3DPA), and also, 3,4-dihydroxybutyric acid was formed. 2K3DXA was further converted to glycolaldehyde when genes encoding by either YagE or YjhH aldolase from E. coli were expressed. Reduction of glycolaldehyde to ethylene glycol by an endogenous aldo-keto reductase activity resulted further in accumulation of ethylene glycol of 14 mg/L. The possibility of simultaneous production of lactic and glycolic acids was evaluated by expression of gene encoding lactate dehydrogenase ldhL from Lactobacillus helveticus together with aldA. Interestingly, this increased the accumulation of glycolic acid to 1 g/L. The D-xylonate dehydratase activity in yeast was notably low, possibly due to inefficient Fe–S cluster synthesis in the yeast cytosol, and leading to D-xylonic acid accumulation. The dehydratase activity was significantly improved by targeting its expression to mitochondria or by altering the Fe–S cluster metabolism of the cells with FRA2 deletion.
11/1/17 12:00:00 AM
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2.1.9 Glycolic acid
GLYCOLIC ACID PRODUCTION BY FERMENTATION FROM RENEWABLE RESOURCES

A method for the fermentative production of glycolic acid by culturing a recombinant microorganism modified for converting a source of carbon to glycolic acid, in an appropriate culture medium comprising a fermentable source of carbon capable of being metabolized by the microorganism, said method comprising the steps of: a) Fermentation of the microorganism to produce glycolic acid by converting the source of carbon into glycolic acid, b) Concentration of the glycolic acid in the microorganism or in the medium and c) Recovery of glycolic acid from the fermentation broth and/or the biomass optionally remaining in portions or in the total amount (0-100%) in the end product, wherein the fermentable source of carbon is a sugar selected among monosaccharides, disaccharides, trisaccharides, oligosaccharides and polysaccharides, and wherein the microorganism is modified to attenuate the conversion of glyoxylate to products other than glycolate, with the attenuation of the expression of at least one gene, selected among the following genes involved in glyoxylate metabolism: • _ace_ B encoding malate synthase • _glc_ B encoding the second malate synthase • _gcl_ encoding glyoxylate carboligase • _eda_ encoding 2-keto-3-deoxygluconate 6-phosphate aldolase. A method as claimed in claim 1 wherein glycolate is isolated through a step of polymerization to at least glycolate dimers. A method as claimed in claim 2 wherein glycolate is recovered by depolymerization from glycolate dimers, oligomers and/or polymers. A method as claimed in claim 1 wherein the microorganism contains at least one gene encoding a polypeptide catalyzing the conversion of glyoxylate to glycolate. A method as claimed in claim 4 wherein the gene encodes a NADPH dependent glyoxylate reductase. A method as claimed in claim 5 in which the gene encoding a polypeptide with NADPH dependent glyoxylate reductase activity is endogenous. A method as claimed in any one of claims 4 to 6 wherein the expression of said gene is increased. A method as claimed in any one of claims 5 to 7 wherein the gene encoding a polypeptide with NADPH dependent glyoxylate reductase activity is selected among _ycd_ W and _yia_ E. A method as claimed in any one of claims 1 to 8 wherein the microorganism is modified in such a way that it is unable to substantially metabolize glycolate, the expression of at least one gene, selected among the following genes involved in glycolate metabolism, being attenuated in the microorganism: • _glc_ DEF encoding glycolate oxidase • _aldA_ encoding glycoaldehyde dehydrogenase. A method as claimed in any one of claims 1 to 9 wherein the microorganism is transformed to increase the glyoxylate pathway flux, by: - attenuating the activity of the enzyme isocitrate dehydrogenase, or - attenuating the expression of at least one of the following genes: • _pta_ encoding phospho- transacetylase • _ack_ encoding acetate kinase • _pox_ B encoding pyruvate oxidase, or - increasing the activity of aceA. A method as claimed in claim 10 wherein the expression of _ace_ A is increased by the attenuation of the expression of the genes _icl_ R _or fad_ R _._ A method as claimed in claim 10 wherein the expression of _ace_ A is increased by introducing an artificial promoter upstream of the gene _ace_ A _._ A method as claimed in any one of claims 1 to 12 wherein the availability of NADPH is increased, by attenuating the expression of at least one gene, selected among the following: • _pgi_ encoding the glucose-6-phosphate isomerase • _udh_ A encoding the soluble transhydrogenase • _edd_ encoding phosphogluconate dehydratase. A method as claimed in any one of claims 1 to 13, wherein the carbon source is at least one of the following: glucose, sucrose, mono- or oligosaccharides, or starch. A recombinant microorganism as defined in any one of claims 1 to 14. A microorganism as claimed in claim 15 that is selected among the group consisting of _E. coli, C. glutamicum_ or _S. cerevisiae._
6/7/07 12:00:00 AM
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2.2 3-hydroxypropionic acid

1

3-Hydroxypropionic acid (3-HP) is a non-chiral isomer of lactic acid with a β-hydroxyl group. It is a precursor for the synthesis of value-added chemicals including 1,3-propanediol, acrylic acid, and a high value-added biocompatible polymer for medical applications called poly(3-HP). 3HP has been produced from combined glucose and xylose in research. **Process:** * Just xylose with engineered yeast: 7.37 g/L titel, yield: 0.29 cmol/cmol: The batch and fed-batch fermentations were performed in 2.7-L DASGIP Bioreactors (Dasgip, Jülich, Germany). The working volume for batch fermentations was 1 L, the temperature set-point was controlled at 30 °C, the airflow was set at 1 vvm (gas volume flow per unit of liquid volume per minute), the pH was maintained at 5 by feedback controlled addition of 10% NH4OH, the dissolved oxygen was kept above 30% of saturation by feedback control of the stirring speed from 600 rpm until a maximum of 1200 rpm. With a mineral medium (50 g L−1 of xylose, 5 g L−1 of (NH4)2SO4, 3 g L−1 of KH2PO4, 0.5 g L−1 of MgSO4·7H2O, 0.05 mL of antifoam, 1 mL of a vitamin solution and 1 mL of a trace metal solution). Art. [#ARTNUM](#article-25151-2219621454) * Glucose and xylose: Engineered E. coli: 37.6 g/L 3-HP, 0.63 g/L/h and yield of 0.17 g/g. Fed-batch fermentation was conducted in a 2.5 L jar fermenter (Kobiotech, Incheon, Korea) with an initial working volume of 1 L R/5 medium containing 13 g/L glucose and 7 g/L xylose. The main culture was performed at 37 °C with an aeration rate of 2 vvm and agitation speed of 1200 rpm. When the OD600 reached 30, IPTG and coenzyme B12 were added and the temperature was decreased to 25 °C while maintaining the same aeration rate and agitation speed. The feed solution comprised 500 g/L glucose and 250 g/L xylose with 20 g/L MgSO4·7H2O. After induction, the feed solution was introduced continuously. The pH of the medium was automatically adjusted between 6.78 and 6.82 by the addition of 28% ammonia water. Art. [#ARTNUM](#article-25151-2928897536) * Glucose and xylose: Engineered Corynebacterium glutamicum: 62.6 g/L of 3-HP was produced from glucose at a yield of 0.51 g/g. The fed-batch fermentation was carried out in 5 L Braun Sartorius bioreactor with the working volume of 2 L. The medium consists of (per liter) 50 g glucose (or 25 g glucose and 25 g xylose), 24 g NH4Cl, 20 g corn steep liquor, 1 g urea, 2.5 g KH2PO4, 0.75 g MgSO4·7H2O, 50 mg FeSO4·7H2O, 13 mg MnSO4·5H2O, 50 mg CaCl2·2H2O, 6.3 mg CuSO4·5H2O, 1.3 mg ZnSO4·7H2O, 5 mg NiCl2·6H2O, 1.3 mg CoCl2·6H2O, 1.3 mg (NH4)6Mo7O24·4H2O, 14 mg nicotinic acid, 7 mg thiamine-HCl, and 0.5 mg D-biotin, 5 mg chloramphenicol and 25 mg kanamycin. The fermentations were maintained at 30 °C, pH 7.2 (controlled by feeding NH4OH), and aeration of 1vvm. The feeding of concentrated glucose (600 g/L) or glucose and xylose mixture (400 g/L of glucose and 200 g/L of xylose) started after the initial sugar was consumed, and the feeding rate was controlled to keep the glucose concentration lower than 10 g/L. The agitation speed was changed during the fermentation to keep a sufficient oxygen supply (dissolved oxygen higher than 10% of saturation). The cells were induced at 16 h with 0.1 mM IPTG. A total of 60 μM of filter sterilized vitamin B12 was added at 16, 32, and 48 h of cultivation. Art. [#ARTNUM](#article-25151-2558107736) **Current production** * Synthetic: Not cost-effective: not used for bulk production. * Biological: research, pilot from glucose by Cargill (OPXBio), announced commercialization in 2016.

2.2.1 3-hydroxypropionic acid
A platform for Acetyl-CoA synthesis using xylose as feedstock in Saccharomyces cerevisiae
Global rise in temperature and diminishing oil reserves has stimulated a market of alternative replacements to traditional petroleum based products. An alternative is the use of a bio refinery capable of converting biomass to the normally petroleum based products. The baker yeast Saccharomyces cerevisiae is an attractive cell factory as its existing large-scale infrastructures for bioethanol production. However, it cannot utilize xylose, an otherwise unusable part of the plant biomass, which represents of utmost importance in the bio-refinery development. To also have a strain capable to produce a wide range of products, it could be used as a platform to base a bio refinery upon. Therefore, the aim of this study is to generate platform strains capable of forming acetyl-CoA, an intermediate metabolite in many of the cells metabolic reactions and also for many other industrially relevant bio-chemicals. With this goal in mind, the metabolism of S. cerevisiae was engineered. The genes encoding an isomerase-based xylose assimilation pathway (RTG, XI, XKS), and a phosphoketolase pathway (XPK, PTA), were cloned into the yeast strain CEN.PK113-5D to enable the yeast to take up and convert xylose into acetyl-CoA. The functionality of this synthetic pathway were evaluated for the production of 3-hydroxypropionic acid via introduction of ACC1** and MCR genes into the engineered strains. By characterisation of all the engineered strains on glucose growth we found increase of acetate production in strains with the phosphoketolase pathway expressed, indicating the in vivo activity of this pathway. However, expression of the xylose assimilation pathway through genome integration did not render the strains able to grow on xylose, suggesting the low efficiency of the assembled xylose assimilation pathway. To overcome this adaptive laboratory evolution is recommended.
1/1/15 12:00:00 AM
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2.2.2 3-hydroxypropionic acid
Bio-based C-3 Platform Chemical: Biotechnological Production and -Conversion of 3-Hydroxypropionaldehyde
Popular Abstract in English Microbes are present everywhere in the environment and have quite an intimate relationship with humans. Much of our perception about microbes is as disease causing agents but on the other hand it is also the microbes that present cure for the disease. Besides using microbes for traditional applications, for example for providing antibiotics, production of fermented foods, etc., humans are increasingly taking advantage of the ”good” microbes as probiotics for improving health, wastewater treatment, mining of metals, cleaning the environment, and for producing bioenergy, chemicals and materials. The micro-sized organisms contain a complex network of metabolic pathways involving a large number of chemical reactions catalysed by enzymes for utilizing different substances in the environment and converting them to a variety of products. Lactic acid bacteria comprise an important group of microbes that humans have used for thousands of years to conserve and enhance the nutritional value of sensitive foods. Lactobacillus species are a major part of this group. Some Lactobacillus species are used for the production of yoghurt, cheese, sauerkraut, pickles, beer, wine, cider several fermented foods, as well as animal feeds, such as silage. Lactobacillus reuteri is a major component of the bacteria present in guts of mammals and birds. It has been shown that several different strains of L. reuteri have a positive effect on health, including various types of gastrointestinal disorders and oral health. In the late 1980s, it was discovered that L. reuteri produced a novel broad-spectrum antibiotic substance by fermentation of glycerol, which was named as "reuterin” after Gerhard Reuter. Reuterin can inhibit the growth of some harmful Gram-negative and Gram-positive bacteria, along with yeasts, fungi and protozoa. Reuterin is a mixture of three components, made of 3-hydroxypropionaldehyde (3HPA) and its derivatives. This thesis is about 3HPA as a molecule of interest for the chemical industry based on renewable resources. Today, as we become increasingly aware of our dependence on fossil resources to fulfill our needs, and the environmental problems associated with the use of these non-renewable resources, there is a growing interest in the use of renewable resources as raw materials and environment-friendly methods for the production of chemicals, materials and energy. 3HPA is currently not a commercial product. If it could be economically produced from glycerol using the bacteria it can potentially be used as a building block or ”platform” for several other chemicals with 3 carbon atoms (C3), e.g. 1,3-propanediol (1,3PDO), 3-hydroxypropionic acid (3HP), acrolein, etc. Glycerol, commonly known as glycerine, is produced as a side product of hydrolysis of fats, production of ethanol and biodiesel. Over the past decade or more, biodiesel is being produced from several plant oils such as rapeseed-, soybean- and palm oil, and also from used oils. In this thesis, conversion of glycerol to 3HPA using L. reuteri is investigated. When the 3HPA level reaches a certain limit, it starts to affect the cell viability and activity, hence inhibiting its own production. Different strategies to complex 3HPA were studied to improve its production. L. reuteri has also the ability to convert 3HPA to 1,3PDO and 3HP via different pathways. In the thesis, the pathway for 3HP production has been introduced in standard bacteria, Escherichia coli by recombinant DNA technology and shown to be active. One of the enzymes of the pathway has further been studied. The work in this thesis was done in collaboration with Perstorp AB, and was supported by Vinnova, the Swedish Governmental Agency for Innovation Systems. (Less)
1/1/13 12:00:00 AM
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2.2.3 3-hydroxypropionic acid
Enhanced production of 3-hydroxypropionic acid from glucose and xylose by alleviation of metabolic congestion due to glycerol flux in engineered Escherichia coli
Abstract Among platform chemicals obtained from renewable biomass, 3-hydroxypropionic acid (3-HP) has attracted considerable attention. A GC/TOF-MS study revealed that the intracellular metabolites of the TCA cycle and fatty acid synthesis increased in JHS01302, a galP -overexpressing strain of Escherichia coli , during glucose and xylose co-fermentation. Decreased intracellular glycerol levels and increased intracellular biosynthesis of 3-HP were also detected in the strain. Based on these results, the yeast GPD1 gene was replaced with the endogenous gpsA gene to modulate the rate of glycerol metabolism. In flask cultures, JHS01304 containing the gpsA gene displayed 43% lower glycerol accumulation and 52% higher 3-HP production than the control. JHS01304 produced 37.6 g/L 3-HP with a productivity rate of 0.63 g/L/h and yield of 0.17 g/g in the fed-batch fermentation. The metabolome analysis provided valuable information for alleviating the metabolic burden of glycerol flux to improve the production of 3-HP during glucose and xylose co-fermentation.
4/1/19 12:00:00 AM
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2.2.4 3-hydroxypropionic acid
METHOD FOR PRODUCTION OF 3-HYDROXYPROPIONIC ACID USING RECOMBINANT E. COLI WITH HIGH YIELD
The present invention relates to a method for producing 3-hydroxypropionic acid using recombinant escherichia coli with high yield. According to one embodiment of the present invention, provided is a method for producing 3-hydroxypropionic acid. According to one embodiment of the present invention, the method for producing 3-hydroxypropionic acid with high yield uses recombinant escherichia coli as a host. According to one embodiment of the present invention, the method for producing 3-hydroxypropionic acid with high yield uses glycerol, xylose, and glucose.
7/30/14 12:00:00 AM
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2.2.5 3-hydroxypropionic acid
Metabolic engineering of Corynebacterium glutamicum for the production of 3-hydroxypropionic acid from glucose and xylose
Abstract 3-Hydroxypropionic acid (3-HP) is a promising platform chemical which can be used for the production of various value-added chemicals. In this study, Corynebacterium glutamicum was metabolically engineered to efficiently produce 3-HP from glucose and xylose via the glycerol pathway. A functional 3-HP synthesis pathway was engineered through a combination of genes involved in glycerol synthesis (fusion of  gpd and  gpp from  Saccharomyces cerevisiae ) and 3-HP production ( pduCDEGH from  Klebsiella pneumoniae and aldehyde dehydrogenases from various resources). High 3-HP yield was achieved by screening of active aldehyde dehydrogenases and by minimizing byproduct synthesis ( gapA A1G Δ ldhA Δ pta-ackA Δ poxB Δ glpK ). Substitution of phosphoenolpyruvate-dependent glucose uptake system (PTS) by inositol permeases ( iolT1 ) and glucokinase ( glk ) further increased 3-HP production to 38.6 g/L, with the yield of 0.48 g/g glucose. To broaden its substrate spectrum, the engineered strain was modified to incorporate the pentose transport gene  araE and xylose catabolic gene  xylAB , allowing for the simultaneous utilization of glucose and xylose. Combination of these genetic manipulations resulted in an engineered C. glutamicum strain capable of producing 62.6 g/L 3-HP at a yield of 0.51 g/g glucose in fed-batch fermentation. To the best of our knowledge, this is the highest titer and yield of 3-HP from sugar. This is also the first report for the production of 3-HP from xylose, opening the way toward 3-HP production from abundant lignocellulosic feedstocks.
1/1/17 12:00:00 AM
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2.2.6 3-hydroxypropionic acid
Production of 3-hydroxypropionic acid from glucose and xylose by metabolically engineered Saccharomyces cerevisiae
Biomass, the most abundant carbon source on the planet, may in the future become the primary feedstock for production of fuels and chemicals, replacing fossil feedstocks. This will, however, require development of cell factories that can convert both C6 and C5 sugars present in lignocellulosic biomass into the products of interest. We engineered Saccharomyces cerevisiae for production of 3-hydroxypropionic acid (3HP), a potential building block for acrylates, from glucose and xylose. We introduced the 3HP biosynthetic pathways via malonyl-CoA or β-alanine intermediates into a xylose-consuming yeast. Using controlled fed-batch cultivation, we obtained 7.37±0.17g 3HPL-1 in 120hours with an overall yield of 29±1%Cmol 3HPCmol-1 xylose. This study is the first demonstration of the potential of using S. cerevisiae for production of 3HP from the biomass sugar xylose.
12/1/15 12:00:00 AM
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2.2.7 3-hydroxypropionic acid
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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2.2.8 3-hydroxypropionic acid
Compositions and methods for 3-hydroxypropionic acid production

1. A transformed yeast cell selected from _I. orientalis, C. lambica_ , and _S. bulderi_ , comprising an active 3-HP fermentation pathway, wherein the pathway comprises an exogenous gene that is not present in the native form of the cell which encodes for an aspartate 1-decarboxylase (ADC) wherein the cell is capable of growing at a pH of less than 4 in media containing 75 g/L or greater 3-HP. 2. The transformed yeast cell of claim 1, wherein the exogenous ADC gene is operatively linked to one or more exogenous regulatory elements. 3. The transformed yeast cell of claim 1, wherein said cell further comprises one or more deletions or disruptions of a native gene selected from PDC, ADH, GAL6, CYB2A, CYB2B, GPD, GPP, ALD, and PCK genes. 4. The transformed yeast cell of claim 3, wherein one or more of the deletions or disruptions results from insertion of the exogenous ADC gene. 5. The transformed yeast cell of claim 1, wherein the cell is a 3-HP- resistant yeast cell. 6. The transformed yeast cell of claim 1, wherein the cell has undergone mutation and/or selection, such that the mutated and/or selected cell possess a higher degree of resistance to 3-HP than a wild-type cell of the same species. 7. The transformed yeast cell of claim 6, wherein the cell has undergone mutation and/or selection before being genetically modified with the exogenous ADC gene. 8. The transformed yeast cell of claim 6, wherein the cell has undergone selection in the presence of lactic acid or 3-HP. 9. The transformed yeast cell of claim 8, wherein the selection is chemostat selection. 10. The transformed yeast cell of claim 1, wherein the yeast cell is an _I. orientalis_ yeast cell. 11. The transformed yeast cell of claim 1, wherein the yeast cell is an _I. orientalis_ CNB1 yeast cell. 12. The transformed yeast cell of claim 11, wherein the modified yeast cell is unable to ferment pentose sugars. 13. The transformed yeast cell of claim 1, wherein the yeast cell is a _C. lambica_ yeast cell. 14. The transformed yeast cell of claim 1, wherein the yeast cell is an _S. bulderi_ yeast cell. 15. The transformed yeast cell of claim 1, further comprising an exogenous PYC gene. 16. The transformed yeast cell of claim 1, further comprising an exogenous AAT gene. 17. The transformed yeast cell of claim 1, further comprising an exogenous BAAT gene or an exogenous gabT gene. 18. The transformed yeast cell of claim 17, wherein said BAAT gene or gabT gene is a BAAT gene that is also a gabT gene. 19. The transformed yeast cell of claim 1, further comprising an exogenous 3-HPDH gene. 20. The transformed yeast cell of claim 10, further comprising an exogenous 3-HPDH gene. 21. The transformed yeast cell of claim 19, wherein the 3-HPDH gene is also a HIBADH gene. 22. The transformed yeast cell of claim 19, wherein the 3-HPDH gene is also a 4-hydroxybutyrate dehydrogenase gene. 23. The transformed yeast cell of claim 1, further comprising an exogenous PPC gene. 24. A method of producing 3-HP comprising:(i) culturing the transformed yeast cell of claim 1 in the presence of medium comprising at least one carbon source; and(ii) isolating 3-HP from the culture. 25. The method of claim 24, wherein said carbon source is selected from glucose, xylose, arabinose, sucrose, fructose, cellulose, glucose oligomers, and glycerol. 26. The method of claim 24, wherein the medium is at a pH of less than 5. 27. The method of claim 24, wherein the transformed yeast cell is an _I. orientalis_ CNB1 yeast cell. 28. The method of claim 27, wherein the transformed yeast cell is unable to ferment pentose sugars. 29. The method of claim 24, wherein the transformed yeast cell is an _I. orientalis_ yeast cell. 30. The method of claim 24, wherein the transformed yeast cell is a _C. lambica_ yeast cell. 31. The method of claim 24, wherein the transformed yeast cell is an _S. bulderi_ yeast cell. 32. The method of claim 29, wherein the transformed yeast cell further comprises an exogenous 3-HPDH gene.
11/21/11 12:00:00 AM
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2.3 Lactic acid

1

Lactic acid (2-hydroxypropanoic acid) is a three‑carbon carboxylic acid used in food, cosmetic, pharmaceutical, and polymer industries. Current commercial production of lactic acid largely depends on the microbial culture because of mild operating conditions and the high chiral purity of the product. **Applications:** Lactic acid and its salts have many long known applications in everyday life ranging from food additives to processing fluids. For instance, in the food industry it is used as an acidulant, a preservative and an emulgator. Next to these existing applications, lactic acid has a major potential for the synthesis of the biopolymer PLA. Given the right conditions and catalytic functionalities, lactic acid may be converted into a wide range of useful intermediates such as acrylic acid, propylene glycol, 2,3-pentanedione, acetaldehyde, pyruvic acid, and lactide (the monomer in PLA synthesis). Besides the latter cyclic ester, a wide range of linear esters (alkyl lactates) are easily produced as well and these possess unique solvation properties. **Process:** * Yeast S. cerevisiae: 0.69 g/g, 50 g/L. The bioreactor fermentations were conducted in YP medium containing 80 g/L of xylose using a BioFlo/CelliGen 115 bioreactor (New Brunswick Scientific Co., USA). An initial yeast cell concentration of OD = 10 (A600 nm) was used. Working volume was set at 1 L inside of a 2-L glass vessel. Aeration was maintained at 1.5 L/min using microfiltered (0.22 μm) ambient air and an impeller rotation of 200 rpm. Temperature was maintained at 30 °C. NaOH (10 N) was added as needed to maintain a pH value of 6. Art. [#ARTNUM](#article-25163-935779216) **Current prodution:** * Biological: >90% of total production, mainly from glucose and sucrose, fermentation has issues with pH , resulting in base addition and gypsum formation which is a big waste product and gives issues for purification * Chemocatalytic from biobased sources: subject to research because of the difficulties in fermentation for larger scale production.

2.3.1 Lactic acid
Catalytic conversion of hemicellulosic biomass to lactic acid in pH neutral aqueous phase media
Abstract The conversion of lignocellulosic biomass into value-added chemicals using non-toxic heterogeneous catalysts and water as solvent is an attractive green process. Biomass-derived lactic acid is an important renewable chemical building block for synthesizing commodity chemicals, e.g. biodegradable plastics. This paper reports that hemicellulosic biomass, xylan and xylose, can be converted to lactic acid over a ZrO 2 catalyst starting from pH neutral aqueous solutions. The effects of reaction conditions, including temperature, oxygen partial pressure, biomass loading, and catalyst loading, etc., on the conversions of hemicellulosic biomass and the corresponding yields of lactic acid have been investigated. Molar yields of lactic acid, up to 42% and 30% were produced from xylose and xylan, respectively, under the investigated reaction conditions and with the ZrO 2 catalyst. The key intermediates such as glyceraldehyde, glycolaldehyde and pyruvaldehyde were used as the reactants to probe the reaction mechanism. The role of the ZrO 2 catalyst in the retro-aldol condensation of xylose, as well as the catalyst stability, has been discussed.
1/1/15 12:00:00 AM
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2.3.2 Lactic acid
Efficient production of L-lactic acid from xylose by Pichia stipitis.
Microbial conversion of renewable raw materials to useful products is an important objective in industrial biotechnology. Pichia stipitis , a yeast that naturally ferments xylose, was genetically engineered for l-(+)-lactate production. We constructed a P. stipitis strain that expressed the l-lactate dehydrogenase (LDH) from Lactobacillus helveticus under the control of the P. stipitis fermentative ADH1 promoter. Xylose, glucose, or a mixture of the two sugars was used as the carbon source for lactate production. The constructed P. stipitis strain produced a higher level of lactate and a higher yield on xylose than on glucose. Lactate accumulated as the main product in xylose-containing medium, with 58 g/liter lactate produced from 100 g/liter xylose. Relatively efficient lactate production also occurred on glucose medium, with 41 g/liter lactate produced from 94 g/liter glucose. In the presence of both sugars, xylose and glucose were consumed simultaneously and converted predominantly to lactate. Lactate was produced at the expense of ethanol, whose production decreased to ∼15 to 30% of the wild-type level on xylose-containing medium and to 70 to 80% of the wild-type level on glucose-containing medium. Thus, LDH competed efficiently with the ethanol pathway for pyruvate, even though the pathway from pyruvate to ethanol was intact. Our results show, for the first time, that lactate production from xylose by a yeast species is feasible and efficient. This is encouraging for further development of yeast-based bioprocesses to produce lactate from lignocellulosic raw material.
1/1/07 12:00:00 AM
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2.3.3 Lactic acid
Enhanced L-(+)-Lactic Acid Production by an Adapted Strain of Rhizopus oryzae using Corncob Hydrolysate
Corncob is an economic feedstock and more than 20 million tons of corncobs are produced annually in China. Abundant xylose can be potentially converted from the large amount of hemicellulosic materials in corncobs, which makes the crop residue an attractive alternative substrate for a value-added production of a variety of bioproducts. Lactic acid can be used as a precursor for poly-lactic acid production. Although current industrial lactic acid is produced by lactic acid bacteria using enriched medium, production by Rhizopus oryzae is preferred due to its exclusive formation of the L-isomer and a simple nutrition requirement by the fungus. Production of L-(+)-lactic acid by R. oryzae using xylose has been reported; however, its yield and conversion rate are poor compared with that of using glucose. In this study, we report an adapted R. oryzae strain HZS6 that significantly improved efficiency of substrate utilization and enhanced production of L-(+)-lactic acid from corncob hydrolysate. It increased L-(+)-lactic acid final concentration, yield, and volumetric productivity more than twofold compared with its parental strain. The optimized growth and fermentation conditions for Strain HZS6 were defined.
1/1/08 12:00:00 AM
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2.3.4 Lactic acid
In vitro reconstitution and characterisation of the oxidative d -xylose pathway for production of organic acids and alcohols
The oxidative d-xylose pathway, i.e. Dahms pathway, can be utilised to produce from cheap biomass raw material useful chemical intermediates. In vitro metabolic pathways offer a fast way to study the rate-limiting steps and find the most suitable enzymes for each reaction. We have constructed here in vitro multi-enzyme cascades leading from d-xylose or d-xylonolactone to ethylene glycol, glycolic acid and lactic acid, and use simple spectrophotometric assays for the read-out of the efficiency of these pathways. Based on our earlier results, we focussed particularly on the less studied xylonolactone ring opening (hydrolysis) reaction. The bacterial Caulobacter crescentus lactonase (Cc XylC), was shown to be a metal-dependent enzyme clearly improving the formation of d-xylonic acid at pH range from 6 to 8. The following dehydration reaction by the ILVD/EDD family d-xylonate dehydratase is a rate-limiting step in the pathway, and an effort was made to screen for novel enolase family d-xylonate dehydratases, however, no suitable replacing enzymes were found for this reaction. Concerning the oxidation of glycolaldehyde to glycolic acid, several enzyme candidates were also tested. Both Escherichia coli aldehyde dehydrogenase (Ec AldA) and Azospirillum brasilense α-ketoglutarate semialdehyde dehydrogenase (Ab AraE) proved to be suitable enzymes for this reaction.
4/11/19 12:00:00 AM
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2.3.5 Lactic acid
Isolation and Characterization of Acid-Tolerant, Thermophilic Bacteria for Effective Fermentation of Biomass-Derived Sugars to Lactic Acid
Biomass-derived sugars, such as glucose, xylose, and other minor sugars, can be readily fermented to fuel ethanol and commodity chemicals by the appropriate microbes. Due to the differences in the optimum conditions for the activity of the fungal cellulases that are required for depolymerization of cellulose to fermentable sugars and the growth and fermentation characteristics of the current industrial microbes, simultaneous saccharification and fermentation (SSF) of cellulose is envisioned at conditions that are not optimal for the fungal cellulase activity, leading to a higher-than-required cost of cellulase in SSF. We have isolated bacterial strains that grew and fermented both glucose and xylose, major components of cellulose and hemicellulose, respectively, to l(+)-lactic acid at 50°C and pH 5.0, conditions that are also optimal for fungal cellulase activity. Xylose was metabolized by these new isolates through the pentose-phosphate pathway. As expected for the metabolism of xylose by the pentose-phosphate pathway, [13C]lactate accounted for more than 90% of the total 13C-labeled products from [13C]xylose. Based on fatty acid profile and 16S rRNA sequence, these isolates cluster with Bacillus coagulans, although the B. coagulans type strain, ATCC 7050, failed to utilize xylose as a carbon source. These new B. coagulans isolates have the potential to reduce the cost of SSF by minimizing the amount of fungal cellulases, a significant cost component in the use of biomass as a renewable resource, for the production of fuels and chemicals.
5/1/06 12:00:00 AM
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2.3.6 Lactic acid
Lactic acid production from biomass-derived sugars via co-fermentation of Lactobacillus brevis and Lactobacillus plantarum.
Lignocellulosic biomass is an attractive alternative resource for producing chemicals and fuels. Xylose is the dominating sugar after hydrolysis of hemicellulose in the biomass, but most microorganisms either cannot ferment xylose or have a hierarchical sugar utilization pattern in which glucose is consumed first. To overcome this barrier, Lactobacillus brevis ATCC 367 was selected to produce lactic acid. This strain possesses a relaxed carbon catabolite repression mechanism that can use glucose and xylose simultaneously; however, lactic acid yield was only 0.52 g g −1 from a mixture of glucose and xylose, and 5.1 g L −1 of acetic acid and 8.3 g L −1 of ethanol were also formed during production of lactic acid. The yield was significantly increased and ethanol production was significantly reduced if L. brevis was co-cultivated with Lactobacillus plantarum ATCC 21028. L. plantarum outcompeted L. brevis in glucose consumption, meaning that L. brevis was focused on converting xylose to lactic acid and the by-product, ethanol, was reduced due to less NADH generated in the fermentation system. Sequential co-fermentation of L. brevis and L. plantarum increased lactic acid yield to 0.80 g g −1 from poplar hydrolyzate and increased yield to 0.78 g lactic acid per g of biomass from alkali-treated corn stover with minimum by-product formation. Efficient utilization of both cellulose and hemicellulose components of the biomass will improve overall lactic acid production and enable an economical process to produce biodegradable plastics.
6/1/15 12:00:00 AM
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2.3.7 Lactic acid
Lactic acid production from xylose by engineered Saccharomyces cerevisiae without PDC or ADH deletion
Production of lactic acid from renewable sugars has received growing attention as lactic acid can be used for making renewable and bio-based plastics. However, most prior studies have focused on production of lactic acid from glucose despite that cellulosic hydrolysates contain xylose as well as glucose. Microbial strains capable of fermenting both glucose and xylose into lactic acid are needed for sustainable and economic lactic acid production. In this study, we introduced a lactic acid-producing pathway into an engineered Saccharomyces cerevisiae capable of fermenting xylose. Specifically, ldhA from the fungi Rhizopus oryzae was overexpressed under the control of the PGK1 promoter through integration of the expression cassette in the chromosome. The resulting strain exhibited a high lactate dehydrogenase activity and produced lactic acid from glucose or xylose. Interestingly, we observed that the engineered strain exhibited substrate-dependent product formation. When the engineered yeast was cultured on glucose, the major fermentation product was ethanol while lactic acid was a minor product. In contrast, the engineered yeast produced lactic acid almost exclusively when cultured on xylose under oxygen-limited conditions. The yields of ethanol and lactic acid from glucose were 0.31 g ethanol/g glucose and 0.22 g lactic acid/g glucose, respectively. On xylose, the yields of ethanol and lactic acid were <0.01 g ethanol/g xylose and 0.69 g lactic acid/g xylose, respectively. These results demonstrate that lactic acid can be produced from xylose with a high yield by S. cerevisiae without deleting pyruvate decarboxylase, and the formation patterns of fermentations can be altered by substrates.
10/1/15 12:00:00 AM
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2.3.8 Lactic acid
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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2.3.9 Lactic acid
Production of fuels and chemicals from xylose by engineered Saccharomyces cerevisiae : a review and perspective
Efficient xylose utilization is one of the most important pre-requisites for developing an economic microbial conversion process of terrestrial lignocellulosic biomass into biofuels and biochemicals. A robust ethanol producing yeast Saccharomyces cerevisiae has been engineered with heterologous xylose assimilation pathways. A two-step oxidoreductase pathway consisting of NAD(P)H-linked xylose reductase and NAD+-linked xylitol dehydrogenase, and one-step isomerase pathway using xylose isomerase have been employed to enable xylose assimilation in engineered S. cerevisiae. However, the resulting engineered yeast exhibited inefficient and slow xylose fermentation. In order to improve the yield and productivity of xylose fermentation, expression levels of xylose assimilation pathway enzymes and their kinetic properties have been optimized, and additional optimizations of endogenous or heterologous metabolisms have been achieved. These efforts have led to the development of engineered yeast strains ready for the commercialization of cellulosic bioethanol. Interestingly, xylose metabolism by engineered yeast was preferably respiratory rather than fermentative as in glucose metabolism, suggesting that xylose can serve as a desirable carbon source capable of bypassing metabolic barriers exerted by glucose repression. Accordingly, engineered yeasts showed superior production of valuable metabolites derived from cytosolic acetyl-CoA and pyruvate, such as 1-hexadecanol and lactic acid, when the xylose assimilation pathway and target synthetic pathways were optimized in an adequate manner. While xylose has been regarded as a sugar to be utilized because it is present in cellulosic hydrolysates, potential benefits of using xylose instead of glucose for yeast-based biotechnological processes need to be realized.
12/1/17 12:00:00 AM
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2.3.10 Lactic acid
d-lactic acid production from renewable lignocellulosic biomass via genetically modified Lactobacillus plantarum.
d-lactic acid is of great interest because of increasing demand for biobased poly-lactic acid (PLA). Blending poly-l-lactic acid with poly-d-lactic acid greatly improves PLA's mechanical and physical properties. Corn stover and sorghum stalks treated with 1% sodium hydroxide were investigated as possible substrates for d-lactic acid production by both sequential saccharification and fermentation and simultaneous saccharification and cofermentation (SSCF). A commercial cellulase (Cellic CTec2) was used for hydrolysis of lignocellulosic biomass and an l-lactate-deficient mutant strain Lactobacillus plantarum NCIMB 8826 ldhL1 and its derivative harboring a xylose assimilation plasmid (ΔldhL1-pCU-PxylAB) were used for fermentation. The SSCF process demonstrated the advantage of avoiding feedback inhibition of released sugars from lignocellulosic biomass, thus significantly improving d-lactic acid yield and productivity. d-lactic acid (27.3 g L−1) and productivity (0.75 g L−1 h−1) was obtained from corn stover and d-lactic acid (22.0 g L−1) and productivity (0.65 g L−1 h−1) was obtained from sorghum stalks using ΔldhL1-pCU-PxylAB via the SSCF process. The recombinant strain produced a higher concentration of d-lactic acid than the mutant strain by using the xylose present in lignocellulosic biomass. Our findings demonstrate the potential of using renewable lignocellulosic biomass as an alternative to conventional feedstocks with metabolically engineered lactic acid bacteria to produce d-lactic acid. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:271–278, 2016
3/1/16 12:00:00 AM
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2.4 Malic acid

1

Malic acid is an organic compound with the molecular formula C4H6O5. It is a dicarboxylic acid that is made by all living organisms, contributes to the sour taste of fruits, and is used as a food additive. [Wiki](https://en.wikipedia.org/wiki/Malic_acid) **Process:** * A. oryzae: 40 g/L, 0.19 g/L/h, 0.49 mol/mol. For the bioreactor cultivations, 1.5 L of main culture medium was used. Medium: 120 g/L xylose, 1.2 g/L (NH4)2SO4, 0.1 g/L KH2PO4, 0.17 g/L K2HPO4 · 3H2O, 0.1 g/L MgSO4 · 7H2O, 0.1 g/L CaCl2 · 2H2O, 5 mg/L NaCl, and 60 mg/L FeSO4 · 7H2O. For pH regulation, 90 g/L CaCO3 was added. The process was operated in the small-scale bioreactor (vessel volume 2.0 L) Minifors (Infors, Switzerland) at either 30 or 35 °C, an aeration rate of 0.5 vvm, and a stirrer speed of 300 rpm. Additionally, 120 g CaCO3 for pH regulation and 200 μL of antifoam reagent (Contraspum A 4050 HAC, Tschimmer und Schwarz) were added before autoclaving. Art. [#ARTNUM](#article-25195-2057194747) * Aspergillus parasiticus: The optimal medium components were as follows:the xylose,(NH4)2SO4,yeast extract powder,MgSO4,MnSO4·H2O,FeSO4·7H2O and CaCO3 were 100.0,2.0,3.0,0.20,0.15,0.08 and 80.00 g/L,respectively.The yield of malic acid from the optimal condition was 53.58 g/L and it was 40.5% higher than that of original condition.The reasonable fermentation conditions were inoculum ratio 8%(V/V),the liquid volume in the shake flask 60 mL/250 mL,fermentation temperature 32 °C,rotation speed 170 r/min,leading to the 55.47 g/L yield of L-malic acid. Art. [#ARTNUM](#article-25195-2356769566) **Current production:** * Synthetic: Racemic malic acid is produced industrially by the double hydration of maleic anhydride (petrochemically). * Biological: research **Research Findings** * Molds of the genus Aspergillus are able to produce malic acid in large quantities from glucose and other carbon sources. In order to enhance the production potential of Aspergillus oryzae DSM 1863, production and consumption rates in an established bioreactor batchprocess based on glucose were determined. At 35 °C, up to 42 g/L malic acid was produced in a 168h batch process with fumaric acid as a byproduct. In prolonged shaking flask experiments (353 h), the suitability of the alternative carbon sources xylose and glycerol at a carbontonitrogen (C/N) ratio of 200:1 and the influence of different C/N ratios in glucose cultivations were tested. When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Art. [#ARTNUM](#article-25195-2057194747)

2.4.1 Malic acid
Optimization of fermentation process for L-malic acid with xylose by Aspergillus parasiticus CICC40365
[Objective] To increase the yield of L-malic acid and the utilization rate of xylose,the Aspergillus parasiticus CICC40365 was used as the strain to produce L-malic acid with xylose,and the fermentation technology in shake flask was investigated in the work.[Methods] The medium components and fermentation conditions were optimized through single factor experiments and response surface methodology.[Results] The optimal medium components were as follows:the xylose,(NH4)2SO4,yeast extract powder,MgSO4,MnSO4·H2O,FeSO4·7H2O and CaCO3 were 100.0,2.0,3.0,0.20,0.15,0.08 and 80.00 g/L,respectively.The yield of malic acid from the optimal condition was 53.58 g/L and it was 40.5% higher than that of original condition.The reasonable fermentation conditions were inoculum ratio 8%(V/V),the liquid volume in the shake flask 60 mL/250 mL,fermentation temperature 32 °C,rotation speed 170 r/min,leading to the 55.47 g/L yield of L-malic acid.Meanwhile,the effect of Mg2+,Mn2+ on the relative enzymes in the xylose metabolism indicated that the xylulokinase played an important role in the process of xylose metabolism.[Conclusion] The xylose can be better utilized to produce L-malic acid through fermentation with Aspergillus parasiticus CICC40365,and the yield of L-malic acid and utilization rate of xylose were improved effectively through this experiment.
1/1/14 12:00:00 AM
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2.4.2 Malic acid
Physiological characterization of the high malic acid-producing Aspergillus oryzae strain 2103a-68
Malic acid is a C4 dicarboxylic acid that is currently mainly used in the food and beverages industry as an acidulant. Because of the versatility of the group of C4 dicarboxylic acids, the chemical industry has a growing interest in this chemical compound. As malic acid will be considered as a bulk chemical, microbial production requires organisms that sustain high rates, yields, and titers. Aspergillus oryzae is mainly known as an industrial enzyme producer, but it was also shown that it has a very competitive natural production capacity for malic acid. Recently, an engineered A. oryzae strain, 2103a-68, was presented which overexpressed pyruvate carboxylase, malate dehydrogenase, and a malic acid transporter. In this work, we report a detailed characterization of this strain including detailed rates and yields under malic acid production conditions. Furthermore, transcript levels of the genes of interest and corresponding enzyme activities were measured. On glucose as carbon source, 2103a-68 was able to secrete malic acid at a maximum specific production rate during stationary phase of 1.87 mmol (g dry weight (DW))−1 h−1 and with a yield of 1.49 mol mol−1. Intracellular fluxes were obtained using 13C flux analysis during exponential growth, supporting the success of the metabolic engineering strategy of increasing flux through the reductive cytosolic tricarboxylic acid (rTCA) branch. Additional cultivations using xylose and a glucose/xylose mixture demonstrated that A. oryzae is able to efficiently metabolize pentoses and hexoses to produce malic acid at high titers, rates, and yields.
4/1/14 12:00:00 AM
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2.4.3 Malic acid
Process characterization and influence of alternative carbon sources and carbon-to-nitrogen ratio on organic acid production by Aspergillus oryzae DSM1863
l-Malic acid and fumaric acid are C4 dicarboxylic organic acids and considered as promising chemical building blocks. They can be applied as food preservatives and acidulants in rust removal and as polymerization starter units. Molds of the genus Aspergillus are able to produce malic acid in large quantities from glucose and other carbon sources. In order to enhance the production potential of Aspergillus oryzae DSM 1863, production and consumption rates in an established bioreactor batch-process based on glucose were determined. At 35 °C, up to 42 g/L malic acid was produced in a 168-h batch process with fumaric acid as a by-product. In prolonged shaking flask experiments (353 h), the suitability of the alternative carbon sources xylose and glycerol at a carbon-to-nitrogen (C/N) ratio of 200:1 and the influence of different C/N ratios in glucose cultivations were tested. When using glucose, 58.2 g/L malic acid and 4.2 g/L fumaric acid were produced. When applying xylose or glycerol, both organic acids are produced but the formation of malic acid decreased to 45.4 and 39.4 g/L, respectively. Whereas the fumaric acid concentration was not significantly altered when cultivating with xylose (4.5 g/L), it is clearly enhanced by using glycerol (9.3 g/L). When using glucose as a carbon source, an increase or decrease of the C/N ratio did not influence malic acid production but had an enormous influence on fumaric acid production. The highest fumaric acid concentrations were determined at the highest C/N ratio (300:1, 8.44 g/L) and lowest at the lowest C/N ratio (100:1, 0.7 g/L).
6/1/14 12:00:00 AM
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2.5 Succinic acid

2

Succinic acid is a dicarboxylic acid with the chemical formula (CH2)2(CO2H)2. The name derives from Latin succinum, meaning amber. [Wiki](https://en.wikipedia.org/wiki/Succinic_acid#Applications) Succinic acid is a precursor to some polyesters and a component of some alkyd resins. 1,4-Butanediol (BDO) can be synthesized using succinic as a precursor. The automotive and electronics industries heavily rely on BDO to produce connectors, insulators, wheel covers, gearshift knobs and reinforcing beams. Succinic acid also serves as the bases of certain biodegradable polymers, which are of interest in tissue engineering applications. Succinic acid has mainly been produced with hydrolysates or combined sugars. **Process:** * Engineered E. coli: 0.5 g/g, 25 g/L. Batch fermentations were conducted in a 12 L bioreactor (BR12, Belach Bioteknik AB, Sweden) with a total starting volume of 8 L (including 0.4 L inoculum and 2 L sugar solution). The medium used for strain AFP184 contained the following components (g L‐1):  K2HPO4, 1.4; KH2 PO4, 0.6; (NH4)2SO4, 3.3; MgSO4·7H2O, 0.4; corn steep liquor (50% solids, Sigma‐Aldrich), 15; and 3 mL antifoam agent (Antifoam 204, Sigma‐Aldrich). The fermentation temperature was controlled at 37 °C, and the pH was maintained between 6.6 and 6.7 by automatic addition of NH4OH (15% NH3 solution). The dissolved oxygen concentration (%DO), measured by a pO2 electrode, was regulated by varying the agitation speed between 500 and 1000 rpm. The total fermentation time was 32 h and consisted of an aerobic growth phase and an anaerobic production phase. Art. [#ARTNUM](#article-25162-2031359270) **Current production:** * Synthetic: petrochemically: Historically, succinic acid was obtained from amber by distillation and has thus been known as spirit of amber. Common industrial routes include hydrogenation of maleic acid, oxidation of 1,4-butanediol, and carbonylation of ethylene glycol. * Biological: several commercial and pilot facilities produce SA from sugars (mainly glucose). **Research Findings** * Metabolically engineered E. coli M6PM was constructed and fermentation with pure sugars revealed that it could utilize xylose and glucose efficiently. E. coli M6PM produced a final succinate concentration of 30.03 ± 0.02 g/L and a yield of 1.09 mol/mol during 72 h dualphase fermentation using elephant grass stalk hydrolysate, which resulted in 64% maximum theoretical yield of succinic acid. Art. [#ARTNUM](#article-25162-2904282411) * In this study, xylose mother liquor was utilized to produce succinic acid by recombinant Escherichia coli strain SD121, and the response surface methodology was used to optimize the fermentation media. The optimal conditions of succinic acid fermentation were as follows: 82.62 g L−1 total initial sugars, 42.27 g L−1 MgCO3 and 17.84 g L−1 yeast extract. The maximum production of succinic acid was 52.09 ± 0.21 g L−1 after 84 h with a yield of 0.63 ± 0.03 g g−1 total sugar, approaching the predicted value (53.18 g L−1). It was 1.78-fold of the production of that obtained with the basic medium. Art. [#ARTNUM](#article-25162-1985419561)

2.5.1 Succinic acid
Development of succinic acid production from corncob hydrolysate by Actinobacillus succinogenes
Succinic acid is one of the most important platform chemicals since it has great potential in industrial applications. In this study, corncob hydrolysate was used for succinic acid production. After diluted acid treatment, xylose was released from hemicellulose as the predominant monosaccharide in the hydrolysate, whereas glucose was released very little and most was retained as cellulose in the raw material. Without any detoxification, corncob hydrolysate was used directly as the carbon source in the fermentation. Actinobacillus succinogenes could utilize the sugars in the hydrolysate to produce succinic acid efficiently. Through medium optimization, yeast extract was selected as the nitrogen source and MgCO3 was used to control pH. A total of 23.64 g/l of succinic acid was produced with a yield of 0.58 g/g based on consumed sugar, indicating that the waste corncob residue can be used to produce value-added chemicals practically.
10/1/10 12:00:00 AM
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2.5.2 Succinic acid
Effect of Different Carbon Sources on the Production of Succinic Acid Using Metabolically Engineered Escherichia coli
Succinic acid (SA) is an important platform molecule in the synthesis of a number of commodity and specialty chemicals. In the present work, dual-phase batch fermentations with the E. coli strain AFP184 were performed using a medium suited for large-scale industrial production of SA. The ability of the strain to ferment different sugars was investigated. The sugars studied were sucrose, glucose, fructose, xylose, and equal mixtures of glucose and fructose and glucose and xylose at a total initial sugar concentration of 100 g L -1 . AFP184 was able to utilize all sugars and sugar combinations except sucrose for biomass generation and succinate production. For sucrose as a substrate no succinic acid was produced and none of the sucrose was metabolized. The succinic acid yield from glucose (0.83 g succinic acid per gram glucose consumed anaerobically) was higher than the yield from fructose (0.66 g g -1 ). When using xylose as a carbon source, a yield of 0.50 g g -1 was obtained. In the mixed-sugar fermentations no catabolite repression was detected. Mixtures of glucose and xylose resulted in higher yields (0.60 g g -1 ) than use of xylose alone. Fermenting glucose mixed with fructose gave a lower yield (0.58 g g -1 ) than fructose used as the sole carbon source. The reason is an increased pyruvate production. The pyruvate concentration decreased later in the fermentation. Final succinic acid concentrations were in the range of 25-40 g L -1 . Acetic and pyruvic acid were the only other products detected and accumulated to concentrations of 2.7-6.7 and 0-2.7 g L -1 . Production of succinic acid decreased when organic acid concentrations reached approximately 30 g L -1 . This study demonstrates that E. coli strain AFP184 is able to produce succinic acid in a low cost medium from a variety of sugars with only small amounts of byproducts formed.
4/9/07 12:00:00 AM
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2.5.3 Succinic acid
Efficient production of succinic acid from corn stalk hydrolysates by a recombinant Escherichia coli with ptsG mutation
Succinic acid is considered to be one of the key platform chemicals used in a variety of industrial applications. The exploitation of biomass to produce succinic acid requires a microbial type that can ferment the mixture of reducing sugars derived from lignocellulose. The recombinant Escherichia coli strains with homologous or cyanobacterial ppc overexpression and IdhA, pflB, ptsG mutations were constructed, and the mixed sugar fermentations were carried out with the prominent strain SD121. Then, a modeled corn stalk hydrolysates containing 30 g l(-1) glucose, 10 g l(-1) xylose and 2.5 g l(-1) arabinose was applied for succinic acid fermentation with SD121. A yield of 0.77 g succinic acid g(-1) total sugar was achieved. Fermentation of corn stalk hydrolysates with SD121 produced a final succinic acid concentration of 36.55 g l(-1) with a higher yield of 0.83 g g(-1) total sugar in anaerobic bottles. In two-stage fermentation process in bioreactor, initial aerobic growth facilitated the subsequent anaerobic succinic acid production with a final concentration of 57.81 g l(-1), and a yield of 0.87 g g(-1), total sugar. This was the first report of succinic acid production from corn stalk hydrolysates by metabolically engineered Escherichia coli. The higher succinic acid yield from corn stalk hydrolysates compared to modeled sugar mixtures, showed a great potential usage of renewable biomass as a feedstock for an economical succinic acid production using E. coli. (C) 2010 Elsevier Ltd. All rights reserved.
1/1/11 12:00:00 AM
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2.5.4 Succinic acid
Enzyme Activity Analysis and Directive Breeding of Actinobacillus succinogenes Fermenting Crop Straw Hydrolysate Containing Pentose and Hexose for Production of Succinic Acid
Objective:It is very important to obtain high yield mutant strain on the base of key enzyme activity analysis of A. succinogenes for the industrial bioconversion of succinic acid from crop straw.Methods:The pentose and hexose hydrolyzed from crop straw hydrolyzed by dilute sulfuric acid were determinated using HPLC.During the fermentation of the hydrolysate, the activities of key enzymes were detenninated and regulated.In order to decrease ethanol yield,those strains mutated by soft X-ray of synchronous radiation were screened out on the plates with high concentration of allyl alcohol.Then the alcohol dehydrogenase activity of the mutant strain was compared with that of the parent strain.Results:Determination of the hydrolysate showed that about 47 g glucose and 26 g xylose are from 200 g hydrolyzed crop straw.Pepck,Pc and Mdh are identified as the key enzymes of succinic acid metabolism and the high activity of Adh causes the cumulation of ethanol. Determination of end products from the parent strain indicated that the concentration of ethanol is the highest among those byproducts and that the yield of succinic acid and byproduct ethanol are 54.2 g/L and 8.9 g/L,respectively.Compared with the parent strain,the ethanol concentration produced by anti-allyl alcohol mutant strain S.JST01 decreases by 84%,from 8.9 to 1.4 g/L.Meanwhile the yield of succinic acid increases by 16e,from 54.2 to 63.1 g/L.Enzyme determination showed that the activity unit of alcohol dehydrogenase (Adh)decreases from 614 to 108.Conclusions:The interdiction of metabolic pathway of Adh decreases the metabolism prosuced by ethanol accordingly,thus the succinic acid flux is strengthened by the redundant carbon flux from the byproduct.Furthermore,the mutant strain S.JST01 with the end product yield of 63.1 g/L succinic acid is worth applying to industrial fermentation.
1/1/09 12:00:00 AM
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2.5.5 Succinic acid
Modular pathway engineering of Corynebacterium glutamicum to improve xylose utilization and succinate production
Abstract Xylose-negative Corynebacterium glutamicum has been engineered to utilize xylose as the sole carbon source via either the xylose isomerase (XI) pathway or the Weimberg pathway. Heterologous expression of xylose isomerase and overexpression of a gene encoding for xylulose kinase enabled efficient xylose utilization. In this study, we show that two functionally-redundant transcriptional regulators (GntR1 and GntR2) present on xylose repress the pentose phosphate pathway genes. For efficient xylose utilization, pentose phosphate pathway genes and a phosphoketolase gene were overexpressed with the XI pathway in C. glutamicum . Overexpression of the genes encoding for transaldolase (Tal), 6-phosphogluconate dehydrogenase (Gnd), or phosphoketolase (XpkA) enhanced the growth and xylose consumption rates compared to the wild-type with the XI pathway alone. However, co-expression of these genes did not have a synergetic effect on xylose utilization. For the succinate production from xylose, overexpression of the tal gene with the XI pathway in a succinate-producing strain improved xylose utilization and increased the specific succinate production rate by 2.5-fold compared to wild-type with the XI pathway alone. Thus, overexpression of the tal , gnd , or xpkA gene could be helpful for engineering C. glutamicum toward production of value-added chemicals with efficient xylose utilization.
9/1/17 12:00:00 AM
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2.5.6 Succinic acid
Processes and apparatus for producing furfural, levulinic acid, and other sugar-derived products from biomass
In some variations, the invention provides a process for producing furfural, 5-hydroxymethylfurfural, and/or levulinic acid from cellulosic biomass, comprising: fractionating the feedstock in the presence of a solvent for lignin, sulfur dioxide, and water, to produce a liquor containing hemicellulose, cellulose-rich solids, and lignin; hydrolyzing the hemicellulose contained in the liquor, to produce hemicellulosic monomers; dehydrating the hemicellulose to convert at least a portion of C5 hemicelluloses to furfural and to convert at least a portion of C6 hemicelluloses to 5-hydroxymethylfurfural; converting at least some of the 5-hydroxymethylfurfural to levulinic acid and formic acid; and recovering at least one of the furfural, the 5-hydroxymethylfurfural, or the levulinic acid. Other embodiments provide a process for dehydrating hemicellulose to convert oligomeric C5 hemicelluloses to furfural and to convert oligomeric C6 hemicelluloses to 5-hydroxymethylfurfural. The furfural may be converted to succinic acid, or to levulinic acid, for example.
11/18/13 12:00:00 AM
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2.5.7 Succinic acid
Production of Succinic Acid for Lignocellulosic Hydrolysates
The purpose of this Cooperative Research and Development Agreement (CRADA) is to add and test new metabolic activities to existing microbial catalysts for the production of succinic acid from renewables. In particular, they seek to add to the existing organism the ability to utilize xylose efficiently and simultaneously with glucose in mixtures of sugars or to add succinic acid production to another strain and to test the value of this new capability for production of succinic acid from industrial lignocellulosic hydrolyasates. The Contractors and Participant are hereinafter jointly referred to as the 'Parties'. Research to date in succinic acid fermentation, separation and genetic engineering has resulted in a potentially economical process based on the use of an Escherichia coli strain AFP111 with suitable characteristics for the production of succinic acid from glucose. Economic analysis has shown that higher value commodity chemicals can be economically produced from succinic acid based on repliminary laboratory findings and predicted catalytic parameters. The initial target markets include succinic acid itself, succinate salts, esters and other derivatives for use as deicers, solvents and acidulants. The other commodity products from the succinic acid platform include 1,4-butanediol, {gamma}-butyrolactone, 2-pyrrolidinone and N-methyl pyrrolidinone. Current economic analyses indicate that thismore » platform is competitive with existing petrochemical routes, especially for the succinic acid and derivatives. The report presents the planned CRADA objectives followed by the results. The results section has a combined biocatalysis and fermentation section and a commercialization section. This is a nonproprietary report; additional proprietary information may be made available subject to acceptance of the appropriate proprietary information agreements.« less
6/1/02 12:00:00 AM
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2.5.8 Succinic acid
Succinate Production with Metabolically Engineered Escherichia coli Using Elephant Grass Stalk (Pennisetum purpureum) Hydrolysate as Carbon Source
Succinic acid is a spectacular chemical that can be used as the precursor of various industrial products including pharmaceuticals and biochemicals. The improvement of the succinic acid market depends on strains engineering that is capable of producing succinic acid at high yield and excellent growth rate which could utilize the wide range of carbon sources such as renewable biomass. Here we use counter selection using catAsacB for pathway design and strains developments. In this investigation, metabolically engineered Escherichia coli M6PM strain was constructed for the synthesis of succinic acid using elephant grass stalk (Pennisetum purpureum) as a carbon source. Elephant grass stalk hydrolysate was prepared which comprised of 11.60 ± 0.04 g/L glucose, 27.22 ± 0.04 g/L xylose and 0.65 ± 0.04 g/L arabinose. Metabolically engineered E. coli M6PM was constructed and fermentation with pure sugars revealed that it could utilize xylose and glucose efficiently. E. coli M6PM produced a final succinate concentration of 30.03 ± 0.02 g/L and a yield of 1.09 mol/mol during 72 h dual-phase fermentation using elephant grass stalk hydrolysate, which resulted in 64% maximum theoretical yield of succinic acid. The high succinate yield from elephant grass stalk demonstrated possible application of renewable biomass as feedstock for the synthesis of succinic acid using recombinant E. coli.
12/7/18 12:00:00 AM
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2.5.9 Succinic acid
Succinic acid production from lignocellulosic hydrolysate by Basfia succiniciproducens
The production of chemicals alongside fuels will be essential to enhance the feasibility of lignocellulosic biorefineries. Succinic acid (SA), a naturally occurring C4-diacid, is a primary intermediate of the tricarboxylic acid cycle and a promising building block chemical that has received significant industrial attention. Basfia succiniciproducens is a relatively unexplored SA-producing bacterium with advantageous features such as broad substrate utilization, genetic tractability, and facultative anaerobic metabolism. Here B. succiniciproducens is evaluated in high xylose-content hydrolysates from corn stover and different synthetic media in batch fermentation. SA titers in hydrolysate at an initial sugar concentration of 60 g/L reached up to 30 g/L, with metabolic yields of 0.69 g/g, and an overall productivity of 0.43 g/L/h. These results demonstrate that B. succiniciproducens may be an attractive platform organism for bio-SA production from biomass hydrolysates.
8/1/16 12:00:00 AM
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2.5.10 Succinic acid
Succinic acid production from xylose mother liquor by recombinant Escherichia coli strain.
Succinic acid (1,4-butanedioic acid) is identified as one of important building-block chemicals. Xylose mother liquor is an abundant industrial residue in xylitol biorefining industry. In this study, xylose mother liquor was utilized to produce succinic acid by recombinant Escherichia coli strain SD121, and the response surface methodology was used to optimize the fermentation media. The optimal conditions of succinic acid fermentation were as follows: 82.62 g L−1 total initial sugars, 42.27 g L−1 MgCO3 and 17.84 g L−1 yeast extract. The maximum production of succinic acid was 52.09 ± 0.21 g L−1 after 84 h with a yield of 0.63 ± 0.03 g g−1 total sugar, approaching the predicted value (53.18 g L−1). It was 1.78-fold of the production of that obtained with the basic medium. This was the first report on succinic acid production from xylose mother liquor by recombinant E. coli strains with media optimization using response surface methodology. This work suggested that the xylose mother liquor could be an alte...
11/2/14 12:00:00 AM
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3. Other biological routes

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3.1 Polyhydroxyalkanoates

1

Polyhydroxyalkanoates or PHAs are polyesters produced in nature by numerous microorganisms, including through bacterial fermentation of sugar or lipids. When produced by bacteria they serve as both a source of energy and as a carbon store. PHA polymers are thermoplastic, can be processed on conventional processing equipment, and are, depending on their composition, ductile and more or less elastic. They differ in their properties according to their chemical composition (homo-or copolyester, contained hydroxy fatty acids). They are UV stable, in contrast to other bioplastics from polymers such as polylactic acid, partial ca. temperatures up to 180 °C, and show a low permeation of water. The crystallinity can lie in the range of a few to 70%. Processability, impact strength and flexibility improves with a higher percentage of valerate in the material. PHAs are soluble in halogenated solvents such chloroform, dichloromethane or dichloroethane. PHB is similar in its material properties to polypropylene (PP), has a good resistance to moisture and aroma barrier properties. Polyhydroxybutyric acid synthesized from pure PHB is relatively brittle and stiff. PHB copolymers, which may include other fatty acids such as beta-hydroxyvaleric acid, may be elastic.[Wiki](https://en.wikipedia.org/wiki/Polyhydroxyalkanoates) PHA production is usually done in different stages to separate growth and production, these can be done in different reactors. PHAs can be produced from several sources, mostly combinations of sugars are employed, hydrolysates or glucose/sucrose are used. **Process:** * R. eutropha: 33.70 g/L of P(3HB) in 108 h with a P(3HB) content of 79.02 wt%. Fed-batch fermentation of recombinant R. eutropha (pKM212-XylAB) strain was carried out at 30 °C in 2.5 L jar fermentors (CNS Co. Ltd., Korea) containing 1 L of MR-B medium supplemented with 7.48 g/L of xylose and 16.01 g/L of glucose. A seed culture (100 mL) was prepared in LB medium overnight at 30 °C. The feeding solution containing 375 g/L of glucose and 125 g/L of xylose was added to the medium to maintain the glucose concentration at from 6 to 8 g/L. The culture pH was controlled at 6.8 by the automatic addition of 6 N NaOH and the agitation speed was controlled at 200 rpm. Art. [#ARTNUM](#article-25172-2414017267) * Bacterial strains IPT 048 and IPT 101: 62% polymer content and 0.39 g g−1 PHB yield. They were submitted to two-step cultivations (cell growth, PHA accumulation) in a 10-l bench scale bioreactor at 30°C under conditions of controlled pH (7.0) and dissolved oxygen concentration above 20% of air saturation. The medium used contained 0.132 g l−1 KH2PO4, 1 g l−1 (NH4)2SO4, 0.109 g l−1 MgSO4·7H2O, 0.06 g l−1 CaCl2·2H2O, 0.02 g l−1 ferric ammonium citrate and 2 ml l−1 trace element solution. The trace element solution contained 0.3 g l−1 H3BO3, 0.2 g/l CoCl2·6H2O, 0.1 g l−1 ZnSO4·7H2O, 30.0 mg l−1 MnCl2·4H2O, 30.0 mg l−1 NaMoO4·2H2O, 20.0 mg l−1 NiCl2·6H2O and 10.0 mg l−1 CuSO4·5H2O. The pH was automatically controlled at 7.0 by adding 4 N H2SO4 or 4 N NaOH. Art. [#ARTNUM](#article-25172-2121549329) **Current production:** * biologically: Several companies are producing PHA from different biobased streams, pilot, demonstration and commercial plants exist. **Research findings:** * Ralstonia eutropha was successfully engineered to utilize xylose as a sole carbon source as well as to co-utilize it in the presence of glucose for the synthesis of P(3HB). In addition, R. eutropha engineered to utilized xylose could synthesize P(3HB) from the sunflower stalk hydrolysate solution containing glucose and xylose as major sugars, which suggests that xylose utilizing R. eutropha developed in this study should be useful for development of lignocellulose based microbial processes. Art. [#ARTNUM](#article-25172-2414017267)

3.1.1 Polyhydroxyalkanoates
Anaerobic poly-3- d -hydroxybutyrate production from xylose in recombinant Saccharomyces cerevisiae using a NADH-dependent acetoacetyl-CoA reductase
Poly-3-d-hydroxybutyrate (PHB) that is a promising precursor for bioplastic with similar physical properties as polypropylene, is naturally produced by several bacterial species. The bacterial pathway is comprised of the three enzymes β-ketothiolase, acetoacetyl-CoA reductase (AAR) and PHB synthase, which all together convert acetyl-CoA into PHB. Heterologous expression of the pathway genes from Cupriavidus necator has enabled PHB production in the yeast Saccharomyces cerevisiae from glucose as well as from xylose, after introduction of the fungal xylose utilization pathway from Scheffersomyces stipitis including xylose reductase (XR) and xylitol dehydrogenase (XDH). However PHB titers are still low. In this study the acetoacetyl-CoA reductase gene from C. necator (CnAAR), a NADPH-dependent enzyme, was replaced by the NADH-dependent AAR gene from Allochromatium vinosum (AvAAR) in recombinant xylose-utilizing S. cerevisiae and PHB production was compared. A. vinosum AAR was found to be active in S. cerevisiae and able to use both NADH and NADPH as cofactors. This resulted in improved PHB titers in S. cerevisiae when xylose was used as sole carbon source (5-fold in aerobic conditions and 8.4-fold under oxygen limited conditions) and PHB yields (4-fold in aerobic conditions and up to 5.6-fold under oxygen limited conditions). Moreover, the best strain was able to accumulate up to 14% of PHB per cell dry weight under fully anaerobic conditions. This study reports a novel approach for boosting PHB accumulation in S. cerevisiae by replacement of the commonly used AAR from C. necator with the NADH-dependent alternative from A. vinosum. Additionally, to the best of our knowledge, it is the first demonstration of anaerobic PHB synthesis from xylose.
12/1/16 12:00:00 AM
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3.1.2 Polyhydroxyalkanoates
Engineering xylose metabolism for production of polyhydroxybutyrate in the non-model bacterium Burkholderia sacchari
Despite its ability to grow and produce high-value molecules using renewable carbon sources, two main factors must be improved to use Burkholderia sacchari as a chassis for bioproduction at an industrial scale: first, the lack of molecular tools to engineer this organism and second, the inherently slow growth rate and poly-3-hydroxybutyrate [P(3HB)] production using xylose. In this work, we have addressed both factors. First, we adapted a set of BglBrick plasmids and showed tunable expression in B. sacchari. Finally, we assessed growth rate and P(3HB) production through overexpression of xylose transporters, catabolic or regulatory genes. Overexpression of xylR significantly improved growth rate (55.5% improvement), polymer yield (77.27% improvement), and resulted in 71% of cell dry weight as P(3HB). These values are unprecedented for P(3HB) accumulation using xylose as a sole carbon source and highlight the importance of precise expression control for improving utilization of hemicellulosic sugars in B. sacchari.
12/1/18 12:00:00 AM
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3.1.3 Polyhydroxyalkanoates
Hemicellulose as a potential substrate for production of
Pseudomonas cepacia was evaluated for its ability to utilize xylose, a major hemicellulosic sugar of hardwoods, for the production of the biodegradable, thermoplastic poly(P-hydroxybutyrate) (PHB). This culture produced 2.6 g . L-I of biomass containing 60% (w/w) PHB when grown in shake flasks on an ammonium-limited, mineral salts medium containing 10 g . L-l of xylose. Batch fermentation data showed that growth and PHB production kinetics on xylose were similar to previously published results for the same microorganism on fructose. On xylose, the maximum specific growth rate, the maximum specific PHB production rate (based on total biomass minus PHB biomass), the overall yield of biomass produced from substrate consumed, the yield of PHB produced from substrate consumed (YPHBIS), and the percentage of PHB were 0.22 h-l, 0.072 g . g-I. h-l, 0.29 g . g-l, 0.11 g . g-I and 45% (w/w), respectively. A high maintenance energy (0.119 g of xylose . g of biomass-I. h-I) is probably responsible for the low overall yield. However, the product yield, YPHBIS, was still the highest reported for any microorganism grown on pentosic sugars. Using the YPHBIS of 0.1 1 g . g-l, it was estimated that the substrate cost (in terms of hydrolyzed hemicellulose) for PHB production would be similar to that of cane molasses and half that of bulk glucose.
1/1/95 12:00:00 AM
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3.1.4 Polyhydroxyalkanoates
Metabolic engineering of an E. coli ndh knockout strain for PHB production from mixed glucose–xylose feedstock
BACKGROUND Poly(3-hydroxybutyrate) (PHB), which is completely biodegradable, is considered a potential candidate to replace a number of petroleum-derived polymers due to similar mechanical properties. In a previous study, inactivation of ndh gene in E. coli, which encodes the NDH-II dehydrogenase, resulted in significantly increased PHB production from either glucose or xylose as substrate. RESULTS In this study, the xylose isomerase (EC:5.3.1.5), xylulokinase (EC:2.7.1.17) and the arabinose/xylose transport protein from Bacillus subtilis 168 (encoded by xylA, xylB and araE, respectively) were co-expressed in the ndh knockout strain, E. coli LJ03(pBHR68), which harbors the PHB biosynthesis genes from Ralstonia eutropha. The resulting strain E. coli LJ03(pBHR68+pM-ABE) was able to simultaneously utilize glucose and xylose to accumulate PHB. In flask cultivation, 3.67 g L−1 PHB was produced from a glucose–xylose mixture (10 g L−1 glucose and 5 g L−1 xylose), which was 2.09-fold higher than the production of the control strain E. coli JM109(pBHR68+pM-ABE). Ultimately, PHB production in fed-batch fermentation reached a maximum titer of 21.0 g L−1, representing a 1.93-fold increase relative to the control strain. CONCLUSION Results indicated that the engineered E. coli LJ03 strain is significantly more efficient than the parent strain E. coli JM109 in producing PHB from mixed glucose–xylose feedstock. To the best of our knowledge, this is the first study describing the implementation of an exogenous xylose utilization pathway in E. coli for the production of PHB from mixed glucose–xylose feedstock – a model of lignocellulosic hydrolysate. © 2017 Society of Chemical Industry
10/1/17 12:00:00 AM
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3.1.5 Polyhydroxyalkanoates
Model Study To Assess Softwood Hemicellulose Hydrolysates as the Carbon Source for PHB Production in Paraburkholderia sacchari IPT 101
Softwood hemicellulose hydrolysates are a cheap source of sugars that can be used as a feedstock to produce polyhydroxybutyrates (PHB), which are biobased and compostable bacterial polyesters. To assess the potential of the hemicellulosic sugars as a carbon source for PHB production, synthetic media containing softwood hemicellulose sugars (glucose, mannose, galactose, xylose, arabinose) and the potentially inhibitory lignocellulose degradation products (acetic acid, 5-hydroxymethylfurfural (HMF), furfural, and vanillin) were fermented with the model strain Paraburkholderia sacchari IPT 101. Relative to pure glucose, individual fermentation for 24 h with 20 g/L mannose or galactose exhibited maximum specific growth rates of 97% and 60%, respectively. On the other hand, with sugar mixtures of glucose, mannose, galactose, xylose, and arabinose, the strain converted all sugars simultaneously to reach a maximum PHB concentration of 5.72 g/L and 80.5% PHB after 51 h. The addition of the inhibitor mixture at th...
1/8/18 12:00:00 AM
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3.1.6 Polyhydroxyalkanoates
Pentose-rich hydrolysate from acid pretreated rice straw as a carbon source for the production of poly-3-hydroxybutyrate
Abstract The aim of this work was to evaluate three different bacterial strains for their ability to accumulate poly-3-hydroxybutyrate (PHB) using pentose sugar rich hydrolysate generated from acid pretreated rice straw as the sole carbon source. Out of these, Bacillus firmus NII 0830 showed maximum PHB production. Acid pretreated black liquor contained sugars and sugar degradation products such as formic acid, acetic acid, furfural and hydroxymethyl furfural. The bacterium grew in the hydrolysate medium without any detoxification and it could accumulate 1.9 g/l biomass with 1.697 g/l PHB and the PHB content in the cell was 89%. This was the highest value ever reported from Bacillus species. The optimum conditions for the fermentation media were an inoculum concentration of 6.5%, 90 h of incubation and 0.75% of xylose concentration. The characterization of extracted polymer was carried out by FTIR, 1 H and 13 C NMR which showed characteristics similar to that of the standard PHB from Sigma.
9/1/13 12:00:00 AM
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3.1.7 Polyhydroxyalkanoates
Poly-3-hydroxybutyrate (P3HB) production by bacteria from xylose, glucose and sugarcane bagasse hydrolysate
Fifty-five bacterial strains isolated from soil were screened for efficient poly-3-hydroxybutyrate (P3HB) biosynthesis from xylose. Three strains were also evaluated for the utilization of bagasse hydrolysate after different detoxification steps. The results showed that activated charcoal treatment is pivotal to the production of a hydrolysate easy to assimilate. Burkholderia cepacia IPT 048 and B. sacchari IPT 101 were selected for bioreactor studies, in which higher polymer contents and yields from the carbon source were observed with bagasse hydrolysate, compared with the use of analytical grade carbon sources. Polymer contents and yields, respectively, reached 62% and 0.39 g g−1 with strain IPT 101 and 53% and 0.29 g g−1 with strain IPT 048. A higher polymer content and yield from the carbon source was observed under P limitation, compared with N limitation, for strain IPT 101. IPT 048 showed similar performances in the presence of either growth-limiting nutrient. In high-cell-density cultures using xylose plus glucose under P limitation, both strains reached about 60 g l−1 dry biomass, containing 60% P3HB. Polymer productivity and yield from this carbon source reached 0.47 g l−1 h−1 and 0.22 g g−1, respectively.
7/1/04 12:00:00 AM
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3.1.8 Polyhydroxyalkanoates
Production of biofuels and chemicals from xylose using native and engineered yeast strains
Abstract Numerous metabolic engineering strategies have allowed yeasts to efficiently assimilate xylose, the second most abundant sugar component of lignocellulosic biomass. During the investigation of xylose utilization by yeasts, a global rewiring of metabolic networks upon xylose cultivation has been captured, as opposed to a pattern of glucose repression. A clear understanding of the xylose-induced metabolic reprogramming in yeast would shed light on the optimization of yeast-based bioprocesses to produce biofuels and chemicals using xylose. In this review, we delved into characteristics of yeast xylose metabolism, and potential benefits of using xylose as a carbon source to produce various biochemicals with examples. Transcriptomic and metabolomic patterns of xylose-grown yeast cells were distinct from those on glucose—a conventional sugar of industrial biotechnology—and the gap might lead to opportunities to produce biochemicals efficiently. Indeed, limited glycolytic metabolic fluxes during xylose utilization could result in enhanced production of metabolites whose biosynthetic pathways compete for precursors with ethanol fermentation. Also, alleviation of glucose repression on cytosolic acetyl coenzyme A (acetyl-CoA) synthesis, and respiratory energy metabolism during xylose utilization enhanced production of acetyl-CoA derivatives. Consideration of singular properties of xylose metabolism, such as redox cofactor imbalance between xylose reductase and xylitol dehydrogenase, is necessary to maximize these positive xylose effects. This review argues the importance and benefits of xylose utilization as not only a way of expanding a substrate range, but also an effective environmental perturbation for the efficient production of advanced biofuels and chemicals in yeasts.
12/1/18 12:00:00 AM
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3.1.9 Polyhydroxyalkanoates
Recombinant Ralstonia eutropha engineered to utilize xylose and its use for the production of poly(3-hydroxybutyrate) from sunflower stalk hydrolysate solution
Background Lignocellulosic raw materials have extensively been examined for the production of bio-based fuels, chemicals, and polymers using microbial platforms. Since xylose is one of the major components of the hydrolyzed lignocelluloses, it is being considered a promising substrate in lignocelluloses based fermentation process. Ralstonia eutropha, one of the most powerful and natural producers of polyhydroxyalkanoates (PHAs), has extensively been examined for the production of bio-based chemicals, fuels, and polymers. However, to the best of our knowledge, lignocellulosic feedstock has not been employed for R. eutropha probably due to its narrow spectrum of substrate utilization. Thus, R. eutropha engineered to utilize xylose should be useful in the development of microbial process for bio-based products from lignocellulosic feedstock.
12/1/16 12:00:00 AM
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3.1.10 Polyhydroxyalkanoates
The composition analysis and preliminary cultivation optimization of a PHA-producing microbial consortium with xylose as a sole carbon source.
This work aimed at using xylose as sole substrate, and combining feast-famine process with Nile blue staining as well as denaturing gradient gel electrophoresis (DGGE) analysis to screen polyhydroxyalkanoate (PHA)-producing bacteria from waste activated sludge (WAS). Composition changes of the microbial consortium during domestication were analyzed by DGGE, and the results indicated that there were mainly four classes of bacteria in the final stable system, which were γ-Proteobacteria, Cellvibrio sp., an uncultured bacterium and Pseudomonas sp., respectively. After preliminary optimization, the optimal conditions for the microbial consortium to produce PHA were also obtained as follows: temperature 33 °C, pH 8, xylose concentration 2.4 g/L, C/N ratio 160 and C/P ratio 125. The final PHA accumulation was up to 31% of dry cell weight (DCW), compared to 23.8% of the original consortia. Though our process is at the very beginning and the PHA yield is relatively low, producing PHA from xylose by using microbial consortia is a promising way to save the PHA production cost.
6/1/16 12:00:00 AM
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3.1.11 Polyhydroxyalkanoates
Upgrading wheat straw to HOMO and co-polyhydroxyalkanoates
Polyhydroxyalkanoates (PHAs) are biodegradable and thus environmentally friendly thermoplastics that are synthesized by various microbial strains as intracellular storage materials. These polyesters present a broad range of properties varying from very crystalline to more elastomeric polymers and find applications from agriculture to medicine. Despite their versatility, they are still not competitive due to the high production costs, of which the C-source accounts for circa 30%. To decrease raw materials costs, lignocellulosic agro-industrial residues rich in cellulose and hemicelluloses can be used as the C-source after being processed to yield simple sugars. Wheat straw lignocellulosic hydrolysates (LCH) were prepared (biorefinery.de GmbH) by pre-treating this residual biomass using the AFEX process followed by enzymatic hydrolysis. A hydrolysate rich in glucose and xylose and with low titres of inhibitory compounds is produced that can be used as carbon source for PHA production. Burkholderia sacchari DSM 17165 was selected for its ability to use both hexoses and pentoses. Polymer production was optimized in fed-batch cultivations in stirred-tank reactors (STR). Polymer concentration, volumetric productivity and polymer cell content of respectively 84 g/L, 1.6 g L −1 h −1 and 68 % (w/w) were attained [1]. Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P3HB-co-4HB) copolymers exhibit attractive thermal and mechanical properties due to the 4HB monomer. Synthesis of this monomer was achieved upon the addition of gamma-butyrolactone (GBL) as co-substrate to fed-batch cultures. Using a DOstat feeding strategy for LCH and a continuous addition of GBL, the maximum attained P(3HB-co-4HB) productivity and 4HB molar % were 0.5 g/(L.h) and 5.0 molar %, respectively [2]. Extraction of P(3HB) from the cells usually involves the use of halogenated solvents to attain high recovery yields and purities. However, the use of these solvents causes health and environmental hazards. To lessen this drawback green solvents were tested and high recovery yields and purities were achieved. Lignocellulosic agricultural residues can thus be ugraded with high yields and productivities to value-added products using the biorefinery concept.
2/1/15 12:00:00 AM
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