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Showing papers on "2,3-Butanediol published in 2016"


Journal ArticleDOI
TL;DR: In this article, B. subtilis 168 was engineered to produce chiral pure meso-2,3-BD with high purity under limited oxygen conditions, and the titer of the main byproduct AC was no more than 1.1 ǫ g/L.
Abstract: 2,3-Butanediol (2,3-BD) with low toxicity to microbes, could be a promising alternative for biofuel production. However, most of the 2,3-BD producers are opportunistic pathogens that are not suitable for industrial-scale fermentation. In our previous study, wild-type Bacillus subtilis 168, as a class I microorganism, was first found to generate only d-(−)-2,3-BD (purity >99 %) under low oxygen conditions. In this work, B. subtilis was engineered to produce chiral pure meso-2,3-BD. First, d-(−)-2,3-BD production was abolished by deleting d-(−)-2,3-BD dehydrogenase coding gene bdhA, and acoA gene was knocked out to prevent the degradation of acetoin (AC), the immediate precursor of 2,3-BD. Next, both pta and ldh gene were deleted to decrease the accumulation of the byproducts, acetate and l-lactate. We further introduced the meso-2,3-BD dehydrogenase coding gene budC from Klebsiella pneumoniae CICC10011, as well as overexpressed alsSD in the tetra-mutant (ΔacoAΔbdhAΔptaΔldh) to achieve the efficient production of chiral meso-2,3-BD. Finally, the pool of NADH availability was further increased to facilitate the conversion of meso-2,3-BD from AC by overexpressing udhA gene (coding a soluble transhydrogenase) and low dissolved oxygen control during the cultivation. Under microaerobic oxygen conditions, the best strain BSF9 produced 103.7 g/L meso-2,3-BD with a yield of 0.487 g/g glucose in the 5-L batch fermenter, and the titer of the main byproduct AC was no more than 1.1 g/L. This work offered a novel strategy for the production of chiral pure meso-2,3-BD in B. subtilis. To our knowledge, this is the first report indicating that metabolic engineered B. subtilis could produce chiral meso-2,3-BD with high purity under limited oxygen conditions. These results further demonstrated that B. subtilis as a class I microorganism is a competitive industrial-level meso-2,3-BD producer.

78 citations


Journal ArticleDOI
TL;DR: This work demonstrates the potential of producing meso-2,3-BD with high titer and purity through metabolic engineering of B. licheniformis.
Abstract: 2,3-Butanediol (2,3-BD) can be used as a liquid fuel additive to replace petroleum oil, and as an important platform chemical in the pharmaceutical and plastic industries. Microbial production of 2,3-BD by Bacillus licheniformis presents potential advantages due to its GRAS status, but previous attempts to use this microorganism as a chassis strain resulted in the production of a mix of D-2,3-BD and meso-2,3-BD isomers. The aim of this work was to develop an engineered strain of B. licheniformis suited to produce the high titers of the pure meso-2,3-BD isomer. Glycerol dehydrogenase (Gdh) was identified as the catalyst for D-2,3-BD biosynthesis from its precursor acetoin in B. licheniformis. The gdh gene was, therefore, deleted from the wild-type strain WX-02 to inhibit the flux of acetoin to D-2,3-BD biosynthesis. The acoR gene involved in acetoin degradation through AoDH ES was also deleted to provide adequate flux from acetoin towards meso-2,3-BD. By re-directing the carbon flux distribution, the double-deletion mutant WX-02ΔgdhΔacoR produced 28.2 g/L of meso-2,3-BD isomer with >99 % purity. The titer was 50 % higher than that of the wide type. A bench-scale fermentation by the double-deletion mutant was developed to further improve meso-2,3-BD production. In a fed-batch fermentation, meso-2,3-BD titer reached 98.0 g/L with a purity of >99.0 % and a productivity of 0.94 g/L–h. This work demonstrates the potential of producing meso-2,3-BD with high titer and purity through metabolic engineering of B. licheniformis.

77 citations


Journal ArticleDOI
TL;DR: A wide screening of microorganisms present both into the waste water treatment system in a biodiesel industry and in culture collections was carried out in order to evaluate their potential as new 2,3-BD producer biocatalysts, and the feasibility of the process employing pure Glycerol and different samples of raw glycerol has been demonstrated.

28 citations


Journal ArticleDOI
TL;DR: Results suggested BDH1 and BDH3 played important roles in 2,3-BD formation, BDH2 and GDH have small effects on 2,1-BD production by Serratia sp.
Abstract: Serratia sp. T241, a newly isolated xylose-utilizing strain, produced three 2,3-butanediol (2,3-BD) stereoisomers. In this study, three 2,3-butanediol dehydrogenases (BDH1-3) and one glycerol dehydrogenase (GDH) involved in 2,3-BD isomers formation by Serratia sp. T241 were identified. In vitro conversion showed BDH1 and BDH2 could catalyzed (3S)-acetoin and (3R)-acetoin into (2S,3S)-2,3-BD and meso-2,3-BD, while BDH3 and GDH exhibited the activities from (3S)-acetoin and (3R)-acetoin to meso-2,3-BD and (2R,3R)-2,3-BD. Four encoding genes were assembled into E. coli with budA (acetolactate decarboxylase) and budB (acetolactate synthase), responsible for converting pyruvate into acetoin. E. coli expressing budAB-bdh1/2 produced meso-2,3-BD and (2S,3S)-2,3-BD. Correspondingly, (2R,3R)-2,3-BD and meso-2,3-BD were obtained by E. coli expressing budAB-bdh3/gdh. These results suggested four enzymes might contribute to 2,3-BD isomers formation. Mutants of four genes were developed in Serratia sp. T241. Δbdh1 led to reduced concentration of meso-2,3-BD and (2S,3S)-2,3-BD by 97.7% and 87.9%. (2R,3R)-2,3-BD with a loss of 73.3% was produced by Δbdh3. Enzyme activity assays showed the decrease of 98.4% and 22.4% by Δbdh1 and Δbdh3 compared with the wild strain. It suggested BDH1 and BDH3 played important roles in 2,3-BD formation, BDH2 and GDH have small effects on 2,3-BD production by Serratia sp. T241.

26 citations


Journal ArticleDOI
TL;DR: In this article, the effect of inhibitory materials derived from the pretreatment of lignocellulosic biomass on the production of 2,3-BDO via the fermentation of various biomasses was investigated.

23 citations


Journal ArticleDOI
TL;DR: It was found that simultaneous addition of yeast extract, Na2EDTA, and acetic acid could significantly improve 2,3-BD production and indicate the possibility of directly using the hydrolysate to effectively produce 2, 3-BD.

21 citations


01 Jan 2016
TL;DR: This work offered the first report indicating that metabolic engineered B. subtilis could produce chiral meso-2,3-BD with high purity under limited oxygen conditions, and demonstrated that B.subilis as a class I microorganism is a competitive industrial-level meso,2, 3-BD producer.
Abstract: Background2,3-Butanediol (2,3-BD) with low toxicity to microbes, could be a promising alternative for biofuel production. However, most of the 2,3-BD producers are opportunistic pathogens that are not suitable for industrial-scale fermentation. In our previous study, wild-type Bacillus subtilis 168, as a class I microorganism, was first found to generate only d-(−)-2,3-BD (purity >99 %) under low oxygen conditions.ResultsIn this work, B. subtilis was engineered to produce chiral pure meso-2,3-BD. First, d-(−)-2,3-BD production was abolished by deleting d-(−)-2,3-BD dehydrogenase coding gene bdhA, and acoA gene was knocked out to prevent the degradation of acetoin (AC), the immediate precursor of 2,3-BD. Next, both pta and ldh gene were deleted to decrease the accumulation of the byproducts, acetate and l-lactate. We further introduced the meso-2,3-BD dehydrogenase coding gene budC from Klebsiellapneumoniae CICC10011, as well as overexpressed alsSD in the tetra-mutant (ΔacoAΔbdhAΔptaΔldh) to achieve the efficient production of chiral meso-2,3-BD. Finally, the pool of NADH availability was further increased to facilitate the conversion of meso-2,3-BD from AC by overexpressing udhA gene (coding a soluble transhydrogenase) and low dissolved oxygen control during the cultivation. Under microaerobic oxygen conditions, the best strain BSF9 produced 103.7 g/L meso-2,3-BD with a yield of 0.487 g/g glucose in the 5-L batch fermenter, and the titer of the main byproduct AC was no more than 1.1 g/L.ConclusionThis work offered a novel strategy for the production of chiral pure meso-2,3-BD in B. subtilis. To our knowledge, this is the first report indicating that metabolic engineered B. subtilis could produce chiral meso-2,3-BD with high purity under limited oxygen conditions. These results further demonstrated that B. subtilis as a class I microorganism is a competitive industrial-level meso-2,3-BD producer.

18 citations


Journal ArticleDOI
TL;DR: In this paper, a wild-type Klebsiella sp. strain XRM21, which is capable of producing 2,3-butanediol from a wide variety of carbon sources including glucose, sucrose, xylose, and glycerol, was reported.
Abstract: Chemical 2,3-butanediol is an important platform compound possessing diverse industrial applications. So far, it is mainly produced by using petrochemical feedstock which is associated with high cost and adverse environmental impacts. Hence, finding alternative routes (e.g., via fermentation using renewable carbon sources) to produce 2,3-butanediol are urgently needed. In this study, we report a wild-type Klebsiella sp. strain XRM21, which is capable of producing 2,3-butanediol from a wide variety of carbon sources including glucose, sucrose, xylose, and glycerol. Among them, fermentation of sucrose leads to the highest production of 2,3-butanediol. To maximize the production of 2,3-butanediol, fermentation conditions were first optimized for strain XMR21 by using response surface methodology (RSM) in batch reactors. Subsequently, a fed-batch fermentation strategy was designed based on the optimized parameters, where 91.2 g/L of 2,3-butanediol could be produced from substrate sucrose dosing in 100 g/L for three times. Moreover, random mutagenesis of stain XMR21 resulted in a highly productive mutant strain, capable of producing 119.4 and 22.5 g/L of 2,3-butanediol and ethanol under optimized fed-batch fermentation process within 65 h with a total productivity of 2.18 g/L/h, which is comparable to the reported highest 2,3-butanediol concentration produced by previous strains. This study provides a potential strategy to produce industrially important 2,3-butanediol from low-cost sucrose.

17 citations


Journal ArticleDOI
TL;DR: Metabolic engineering strategies such as cofactor engineering and overexpression of the key enzyme butanediol dehydrogenase were attempted and both led to a statistically significant increase in the 2,3-butanediol yields for sugarcane molasses based fermentation.
Abstract: 2,3-butanediol is known to be a platform chemical with several potential industrial applications. Sustainable industrial scale production can be attained by using a sugarcane molasses based fermentation process using Bacillus subtilis. However, the accumulation of acetoin needs to be reduced to improve process efficiency. In this work, B. subtilis was genetically modified in order to increase the yield of 2,3-butanediol. Metabolic engineering strategies such as cofactor engineering and overexpression of the key enzyme butanediol dehydrogenase were attempted. Both the strategies individually led to a statistically significant increase in the 2,3-butanediol yields for sugarcane molasses based fermentation. Cofactor engineering led to a 26 % increase in 2,3-butanediol yield and overexpression of bdhA led to a 11 % increase. However, the combination of the two strategies did not lead to a synergistic increase in 2,3-butanediol yield.

16 citations


Journal ArticleDOI
19 Oct 2016-PLOS ONE
TL;DR: The effects of temperature, pH-control schemes, and agitation speeds on 2,3-BD production were explored to optimize the fermentation conditions, and a newly isolated bacterium designated as Raoultella ornithinolytica B6 was investigated with a high production of 2, 3-BD in fed-batch fermentation.
Abstract: 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.

14 citations


Journal ArticleDOI
TL;DR: K. pneumoniae knockout mutants developed with an aid of in silico investigation could produce higher amounts of 2,3-BD with increased titer, yield, and productivity.
Abstract: To improve the production of 2,3-butanediol (2,3-BD) in Klebsiella pneumoniae, the genes related to the formation of lactic acid, ethanol, and acetic acid were eliminated. Although the cell growth and 2,3-BD production rates of the K. pneumoniae ΔldhA ΔadhE Δpta-ackA strain were lower than those of the wild-type strain, the mutant produced a higher titer of 2,3-BD and a higher yield in batch fermentation: 91 g 2,3-BD/l with a yield of 0.45 g per g glucose and a productivity of 1.62 g/l.h in fed-batch fermentation. The metabolic characteristics of the mutants were consistent with the results of in silico simulation. K. pneumoniae knockout mutants developed with an aid of in silico investigation could produce higher amounts of 2,3-BD with increased titer, yield, and productivity.

Journal ArticleDOI
TL;DR: Efficient whole-cell biocatalysts from plants and microorganisms were determined in the bioconversion of acetoin to 2,3-BD and the profile of produced stereoisomers demonstrated that microorganisms produce more specific stereoisomer.
Abstract: 2,3-Butanediol (2,3-BD) is a valuable bulk chemical owing to its extensive application in chemical and pharmaceutical industry with diverse applications in drug, cosmetics and food products. In the present study, the biotransformation of acetoin to 2,3-BD by five plant species (Brassica oleracea, Brassica rapa, Daucuscarota, Pastinaca sativa, and Raphnussativus) and five microorganisms (Aspergillusfoetidus, Penicillumcitrinum, Saccharomyces carlbergensis, Pichiafermentans, and Rhodotrulaglutinis) was investigated as a method for the production of 2,3-BD, which can serve as an alternative to the common pentoses and hexoses fermentation by microorganisms. The produced 2,3-BD stereoisomers were characterized and their total conversion yields were determined. The results showed that the examined plants can be used as a green factory for the production of all 2,3-BD stereoisomers, except B. rapa. In microorganisms, P. fermentans and S. carlbergensis produced (-)-2R,3R and mesobutanediol, while P. citrinum produced (+)-2S,3S and mesobutanediol. R. glutinis and A. foetidus produced all three isomers. In conclusion, efficient whole-cell biocatalysts from plants and microorganisms were determined in the bioconversion of acetoin to 2,3-BD. The profile of produced stereoisomers demonstrated that microorganisms produce more specific stereoisomers.

Journal Article
TL;DR: Molasses appear therefore the most interesting feedstock for the production of 2,3-BDO with Bacillus licheniformis ATCC-9789 using different sugars and biowastes as substrates.
Abstract: 2,3-Butanediol (2,3-BDO) is a promising platform compound which could be used to produce valuable derivatives such as methyl ethyl ketone and 1,3-butadiene. The biotechnological production of 2,3-BDO has been mainly studied with Klebsiella sp. under microaerophilic conditions. However the pathogenicity of these strains makes this process not industrially desirable. The aim of this work was to study the possibility of producing 2,3-BDO with the non pathogenic microorganism Bacillus licheniformis ATCC-9789 using different sugars and biowastes as substrates. Shaken flask experiments were carried out using different monosaccharides commonly occurring in plant hydrolyzates (hexoses and pentoses), agroindustrial biowastes (sugar beet molasses and cheese whey) and the diand monosaccharides occurring in them. Flask fermentation of glucose produced 8.2±0,1 g/L 2,3-BDO after 18 hours, corresponding to a 40.4±1.0 g2,3-BDO/100g glucose yield. Under the same conditions, mannose was converted to 7.8±0.2 g/L 2,3-BDO with a yield of 39.4±1.8 g2,3-BDO/100g mannose. Pentoses (xylose and arabinose) were little or not consumed with no production of 2,3-BDO. Biowastes used at 20 g/L provided the corresponding sugars at approximately 10 g/L initial concentrations. While lactose in cheese whey was not used and converted into 2,3-BDO, 2.6±0.3 g/L 2,3-BDO were produced after 14.5 hours from sucrose occurring in molasses, corresponding to a yield of 26.6±3.4 g2,3-BDO/100g sucrose. Molasses appear therefore the most interesting feedstock for the production of 2,3-BDO with Bacillus licheniformis ATCC 9789.

Patent
29 Apr 2016
TL;DR: In this paper, an antibacterial composition containing meso-2,3-butanediol represented by chemical formula 1, which is an isomer of 2, 3-Butanediol, was presented.
Abstract: The present invention relates to an antibacterial composition containing meso-2,3-butanediol represented by chemical formula 1, which is an isomer of 2,3-butanediol, thereby being capable of effectively controlling bacteria and exhibiting superior antibacterial effects even in small amounts.

Patent
29 Jun 2016
TL;DR: In this article, a preparation method of 2,3-butanediol through microbial fermentation is presented, which can effectively remove residual glucose in the fermentation liquid, and is in favor of implementation of next-step twoaqueous-phase extraction, rectification and other refining ways.
Abstract: The invention discloses a preparation method of 2,3-butanediol through microbial fermentation; the method includes the steps: (1) strain culture: culturing lactobacillus with an MRS culture medium, to obtain a lactobacillus seed liquid; and culturing klebsiella pneumoniae with an LB culture medium, to obtain a klebsiella pneumoniae seed liquid; (2) fermentation culture: inoculating a fermentation culture medium with the klebsiella pneumoniae seed liquid, carrying out micro-aerobic fermentation, batch-feeding a glucose solution in the fermentation process to control the concentration of glucose in the fermentation system to be 20 g/L-40 g/L; and (3) fermentation control: in a fermentation last stage, adjusting the pH of the fermentation liquid to 5-6, inoculating with the lactobacillus seed liquid, and carrying out aerobic fermentation under a low-pH condition. The method can effectively remove residual glucose in the fermentation liquid, and is in favor of implementation of next-step two-aqueous-phase extraction, rectification and other refining ways, and is suitable for mass production and application.

Patent
23 Nov 2016
TL;DR: In this paper, an engineering bacterium lacking organic acid production paths and use thereof in co-production of 1, 3-propanediol, 2,3-butanediol and ethanol was revealed.
Abstract: The invention discloses an engineering bacterium lacking organic acid production paths and use thereof in co-production of 1,3-propanediol, 2,3-butanediol and ethanol. Lactic dehydrogenase gene of lactic acid synthesis path of Klebsiella, acetate kinase gene and pyruvic oxidase gene in acetic acid synthesis path, and fumarate reductase gene and isocitrate lyase gene of succinic acid synthesis path are inactivated, so that production of organic acids in metabolism of glycerol of the Klebsiella can be reduced, and yield and conversion rate of alcohols such as1,3-propanediol, 2,3-butanediol and ethanol can be improved, and the co-production of the 1,3-propanediol, 2,3-butanediol and ethanol can be achieved. As a significant reduction in the production of the organic acids, the extraction process of the alcohols can be simplified. The production efficiency of production of high value products by microbial conversion of glycerol can be improved, the production cost is reduced, and the engineering bacterium has important application value.

Patent
03 Aug 2016
TL;DR: In this article, a method for producing acetoin and 2, 3- butanediol through high temperature fermentation has been proposed, with the advantages that raw materials are low in cost, the fermentation process is resistant to infectious microbe contamination, cooling water is saved, and operation is conducted coarsely.
Abstract: The invention discloses bacillus and application thereof to production of acetoin and 2, 3- butanediol through high temperature fermentation. A strain is Bacillus sp. H15-1CGMCC No. 12389, the gram faerbung is positive, spores are produced, the cellular morphology is in a rod shape, and the stain can grow at the temperature of 40-60 DEG C and proliferate at the highest speed at the temperature of 50 DEG C. By means of the strain, degerminated corn flour hydrolysate is adopted as the main raw material, a mechanical stirring and ventilation fermentation tank is adopted for conducting fermentation for 68 hours at the temperature of 50 DEG C, and acetoin with the concentration being 50.8 g/L and 2, 3- butanediol with the concentration being 32.1 g/L can be generated. A method for producing acetoin and 2, 3- butanediol through high temperature fermentation has the advantages that raw materials are low in cost, the fermentation process is resistant to infectious microbe contamination, cooling water is saved, and operation is conducted coarsely.


Patent
03 Aug 2016
TL;DR: In this article, a recombinant microorganism for producing D(-) 2,3-butanediol is described. But the present method is based on the same approach as the one described in this paper.
Abstract: The present invention relates to a recombinant microorganism for producing D(-) 2,3-butanediol, wherein a gene encoding an enzyme for converting acetoin into D(-) 2,3-butanediol is introduced into a microorganism having a pathway for converting acetoin into 2,3-butanediol. In addition, the present invention relates to a method for producing D(-) 2,3-butanediol by using the recombinant microorganism.