Simultaneous production of citric acid and invertase by Yarrowia lipolytica SUC+ transformants

Simultaneous production of citric acid (CA) and invertase by Yarrowia lipolytica A-101-B56-5 (SUC⁺ clone) growing from sucrose, mixture of glucose and fructose, glucose or glycerol was investigated. Among the tested substrates the highest concentration of CA was reached from glycerol (57.15 g/L) with high yield (Y_CA/S = 0.6 g/g). When sucrose was used, comparable amount of CA was secreted (45 g/L) with slightly higher yield (Y_CA/S = 0.643 g/g). In all cultures amount of isocitrate (ICA) was below 2% of total citrates. Considering invertase production, the best carbon source appeared to be sucrose (72 380 U/L). The highest yield of CA and invertase biosynthesis calculated for 1 g of biomass was obtained for cells growing from glycerol (9.9 g/g and 4325 U/g, respectively). Concentrates of extra- and intracellular invertase of the highest activity were obtained from sucrose as substrate (0.5 and 1.8 × 10⁶ U/L, respectively).     

Continuous culture of the microalgae Schizochytrium limacinum on biodiesel-derived crude glycerol for producing docosahexaenoic acid

Crude glycerol is a major byproduct of the biodiesel industry; previous research has proved the feasibility of producing docosahexaenoic acid (DHA, 22:6 n − 3) through fermentation of the algae Schizochytrium limacinum on crude glycerol. The objective of this work is to investigate the cell growth kinetics, substrate utilization efficiency, and DHA production of the algae through a continuous culture. Steady-state biomass yield, biomass productivity, growth yield on glycerol, specific glycerol consumption rate, and fatty acid composition were investigated within the range of dilution rate (D) from 0.2 to 0.6 day⁻¹, and the range of feed crude glycerol concentration (S₀) from 15 to 120 g/L. The maximum specific growth rate was determined as 0.692 day⁻¹. The cells had a true growth yield of 0.283 g/g but with a relatively high maintenance coefficient (0.2216 day⁻¹). The highest biomass productivity of 3.88 g/L-day was obtained at D = 0.3 day⁻¹ and S₀ = 60 g/L, while the highest DHA productivity (0.52 g/L-day) was obtained at D = 0.3 day⁻¹ and S₀ = 90 g/L due to the higher DHA content at S₀ = 90 g/L. The biomass and DHA productivity of the continuous culture was comparable to those of batch culture, while lower than the fed-batch culture, mainly because of the lower DHA content obtained by the continuous culture. Overall, the results show that continuous culture is a powerful tool to investigate the cell growth kinetics and physiological behaviors of the algae growing on biodiesel-derived crude glycerol.     

Oil production by oleaginous yeasts using the hydrolysate from pretreatment of wheat straw with dilute sulfuric acid

This paper explores the use of the hydrolysate from the dilute sulfuric acid pretreatment of wheat straw for microbial oil production. The resulting hydrolysate was composed of pentoses (24.3 g/L) and hexoses (4.9 g/L), along with some other degradation products, such as acetic acid, furfural, and hydroxymethylfurfural (HMF). Five oleaginous yeast strains, Cryptococcus curvatus, Rhodotorula glutinis, Rhodosporidium toruloides, Lipomyces starkeyi, and Yarrowia lipolytica, were evaluated by using this hydrolysate as substrates. The results showed that all of these strains could use the detoxified hydrolysate to produce lipids while except R. toruloides non-detoxified hydrolysate could also be used for the growth of all of the selective yeast strains. C. curvatus showed the highest lipid concentrations in medium on both the detoxified (4.2 g/L) and non-detoxified (5.8 g/L) hydrolysates. And the inhibitory effect studies on C. curvatus indicated HMF had insignificant impacts at a concentration of up to 3 g/L while furfural inhibited cell growth and lipid content by 72.0% and 62.0% at 1 g/L, respectively. Our work demonstrates that lipid production is a promising alternative to utilize hemicellulosic sugars obtained during pretreatment of lignocellulosic materials.     

Plasmodium falciparum: In vitro interaction of quassin and neo-quassin with artesunate, a hemisuccinate derivative of artemisinin

Quassia amara L. (Family Simaroubaceae) is known to have several medicinal properties including the activity against malaria. An HPLC method was employed for purification of the biologically active quassinoids; quassin (Q) and neo-quassin (NQ), further characterized by MALDI-TOF analyses. Purified Q, NQ and the crude bark extract (S1) along with artesunate (AS) were studied for their in vitro anti-plasmodial activity. The in vivo toxicity studies at intraperitoneal doses with higher concentrations of the crude bark extract (S1) in Balb/C mice ruled out the apprehension of toxicity. Interaction studies between the test compounds among themselves (Q + NQ) and individually with artesunate (AS + Q, AS + NQ), were carried out in vitro at four ratios (1:5, 1:2, 2:1 and 5:1) on chloroquine sensitive (MRC-pf-20) and resistant (MRC-pf-303) strains of Plasmodium falciparum. The crude bark extracts of Q. amara exhibited higher P. falciparum inhibitory activity (IC₅₀ = 0.0025 μg/ml) as compared to that of the isolated compounds, quassin (IC₅₀ = 0.06 μg/ml, 0.15 μM), neo-quassin (IC₅₀ = 0.04 μg/ml, 0.1 μM) and also to the positive control, artesunate (IC₅₀ = 0.02 μg/ml, 0.05 μM). The in vitro drug interaction study revealed the compounds, quassin and neo-quassin to be additive to each other. At lower ratios, artesunate was found to be a potential combination partner with both the compounds. It was interesting to note that none of the combinations exhibited antagonistic interactions. This phenomenon offers the opportunity for further exploration of novel therapeutic concentrations and combinations.     

Chemical composition and larvicidal activity of essential oils from Piper species

The larvicidal activity of essential oils of four species of Piper from the Amazon Forest was tested using third-instar larvae of Aedes aegypti. The oils were extracted by steam distillation and analyzed by GC and GC–MS. The main components isolated from each Piper species were as follows: viridiflorol (27.50%), aromadendrene (15.55%) and β-selinene (10.50%) from Piper gaudichaudianum; β-selinene (15.77%) and caryophyllene oxide (16.63%) from Piper humaytanum; dillapiol (54.70%) and myristicin (25.61%) from Piper permucronatum; and asaricin (27.37%) and myristicin (20.26%) from Piper hostmanianum. Amongst all essential oils tested, the most active against larvae of A. aegypti was the oil extracted from P. permucronatum, with a LC₅₀ = 36 μg/ml (LC₉₀ = 47 μg/ml), followed by the essential oil of P. hostmanianum, with a LC₅₀ = 54 μg/ml (LC₉₀ = 72 μg/ml). The oils with higher content of arylpropanoids were more active against larvae of A. aegypti.     

Colonization of Aspergillus japonicus on synthetic materials and application to the production of fructooligosaccharides

The ability of Aspergillus japonicus ATCC 20236 to colonize different synthetic materials (polyurethane foam, stainless steel sponge, vegetal fiber, pumice stones, zeolites, and foam glass) and to produce fructooligosaccharides (FOS) from sucrose (165 g/L) is described. Cells were immobilized in situ by absorption, through direct contact with the carrier particles at the beginning of fermentation. Vegetal fiber was the best immobilization carrier as A. japonicus grew well on it (1.25 g/g carrier), producing 116.3 g/L FOS (56.3 g/L 1-kestose, 46.9 g/L 1-nystose, and 13.1 g/L 1-β-fructofuranosyl nystose) with 69% yield (78% based only in the consumed sucrose amount), giving also elevated activity of the β-fructofuranosidase enzyme (42.9 U/mL). In addition, no loss of material integrity, over a 2 day-period, was found. The fungus also immobilized well on stainless steel sponge (1.13 g/g carrier), but in lesser extents on polyurethane foam, zeolites, and pumice stones (0.48, 0.19, and 0.13 g/g carrier, respectively), while on foam glass no cell adhesion was observed. When compared with the FOS and β-fructofuranosidase production by free A. japonicus, the results achieved using cells immobilized on vegetal fiber were closely similar. It was thus concluded that A. japonicus immobilized on vegetal fiber is a potential alternative for high production of FOS at industrial scale.     

Microbial production of N-acetylneuraminic acid by genetically engineered Escherichia coli

Previously, we described the production of N-acetylneuraminic acid (NeuAc) from N-acetylglucosamine (GlcNAc) in a system combining recombinant Escherichia coli expressing GlcNAc 2-epimerase (slr1975), E. coli expressing NeuAc synthetase (neuB), and Corynebacterium ammoniagenes. However, this system was unsuitable for large-scale production because of its complexity and low productivity. To overcome these problems, we constructed a recombinant E. coli simultaneously overexpressing slr1975 and neuB. This recombinant E. coli produced 81 mM (25 g/L) NeuAc in 22 h without the addition of C. ammoniagenes cells. For manufacturing on an industrial scale, it is preferable to use unconcentrated culture broth as the source of enzymes, and therefore, a high-density cell culture is required. An acetate-resistant mutant strain of E. coli (HN0074) was selected as the host strain because of its ability to grow to a high cell density. The NeuAc aldolase gene of E. coli HN0074 was disrupted by homologous recombination yielding E. coli N18-14, which cannot degrade NeuAc. After a 22 h reaction with 540 mM (120 g/L) GlcNAc in a 5 L jar fermenter, the culture broth of E. coli N18-14 overexpressing slr1975 and neuB contained 172 mM (53 g/L) NeuAc.     

The energy density of jellyfish: Estimates from bomb-calorimetry and proximate-composition

Two techniques are described to calculate energy densities for the bell, gonad and oral arm tissues of three scyphozoan jellyfish (Cyanea capillata, Rhizostoma octopus and Chrysaora hysoscella). First, bomb-calorimetry was used, a technique that is readily available and inexpensive. However, the reliability of this technique for gelatinous material is contentious. Second, further analysis involving the more labour intensive proximate-composition analysis (protein, fat and carbohydrate) was carried out on two species (C. capillata and R. octopus). These proximate data were subsequently converted to energy densities. The two techniques (bomb-calorimetry and proximate-composition) gave very similar estimates of energy density. Differences in energy density were found both amongst different species and between different tissues of the same species. Mean (± S.D.) energy density estimates for whole animals from bomb-calorimetry were 0.18 ± 0.05, 0.11 ± 0.04, and 0.10 ± 0.03 kJ g wet mass^− 1 for C. capillata, R. octopus, and C. hysoscella respectively. The implications of these low energy densities for species feeding on jellyfish are discussed.     

Characterization of a new insect cell line (NTU-YB) derived from the common grass yellow butterfly, Eurema hecabe (Linnaeus) (Pieridae: Lepidoptera) and its susceptibility to microsporidia

A new lepidopteran cell line, NTU-YB, was derived from pupal tissue of Eurema hecabe (Linnaeus) (Pieridae: Lepidoptera). The doubling time of YB cells in TNM-FH medium supplemented with 8% FBS at 28 °C was 26.87 h. The chromosome numbers of YB cells varied widely from 21 to 196 with a mean of 86. Compared to other insect cell lines, the YB cells produced distinct esterase, malate dehydrogenase, and lactate dehydrogenase isozyme patterns. Identity of the internal transcribed spacer region-I (ITS-I) of YB cells to E. hecabe larvae was 96% and to Eurema blanda larvae (tissue isolated from head) was 81%. The YB cells were permissive to Nosema sp. isolated from E. blanda and the infected YB cells showed obvious cytopathic effects after 3 weeks post inoculation. The highest level of spore production was at 4 weeks post inoculation when cells were infected with the Nosema isolate, and spore production was 1.34 ± 0.9 × 10⁶ spore/ml. Ultrastructrual studies showed that YB cells can host in vitro propagation of the E. blanda Nosema isolate, and developing stages were observed in the host cell nuclei as observed in the natural host, E. blanda. The NTU-YB cell line is also susceptible to Nosema bombycis.     

Semicircular canal system in early primates

Mammals with more rapid and agile locomotion have larger semicircular canals relative to body mass than species that move more slowly. Measurements of semicircular canals in extant mammals with known locomotor behaviours can provide a basis for testing hypotheses about locomotion in fossil primates that is independent of postcranial remains, and a means of reconstructing locomotor behaviour in species known only from cranial material. Semicircular canal radii were measured using ultra high resolution X-ray CT data for 9 stem primates (“plesiadapiforms”; n = 11), 7 adapoids (n = 12), 4 omomyoids (n = 5), and the possible omomyoid Rooneyia viejaensis (n = 1). These were compared with a modern sample (210 species including 91 primates) with known locomotor behaviours. The predicted locomotor agilities for extinct primates generally follow expectations based on known postcrania for those taxa. “Plesiadapiforms” and adapids have relatively small semicircular canals, suggesting they practiced less agile locomotion than other fossil primates in the sample, which is consistent with reconstructions of them as less specialized for leaping. The derived notharctid adapoids (excluding Cantius) and all omomyoids sampled have relatively larger semicircular canals, suggesting that they were more agile, with Microchoerus in particular being reconstructed as having had very jerky locomotion with relatively high magnitude accelerations of the head. Rooneyia viejaensis is reconstructed as having been similarly agile to omomyids and derived notharctid adapoids, which suggests that when postcranial material is found for this species it will exhibit features for some leaping behaviour, or for a locomotor mode requiring a similar degree of agility.     

Enhancement of ethanol production from glycerol in a Klebsiella pneumoniae mutant strain by the inactivation of lactate dehydrogenase

Previously, using γ-irradiation treatment, we isolated a mutant strain of Klebsiella pneumoniae (named GEM167) that showed high-level ethanol production from glycerol. In the present study, in an effort to enhance ethanol production, we used a deletion of the lactate dehydrogenase gene to engineer a mutant strain incapable of lactate synthesis. In the ΔldhA mutant of GEM167, the production of ethanol was significantly increased from 21.5 g/l to 28.9 g/l and from 0.93 g/(l h) to 1.2 g/(l h). Introduction of the Zymomonas mobilis pdc and adhII genes encoding pyruvate decarboxylase and aldehyde dehydrogenase, respectively, further improved the ethanol production level from glycerol to 31.0 g/l; this is the highest level reported to date.     

Characterisation of copolymer, poly (hydroxybutyrate-co-hydroxyvalerate) (PHB-co-PHV) produced by Halomonas campisalis (MCM B-1027), its biodegradability and potential application

Characterisation of polyhydroxyalkanoate (PHA) film produced by haloalkalitolerant Halomonascampisalis (MCM B-1027) in 14 L SS fermenter revealed it to have composition of monomer units, HB:HV as 96:4 as analysed by ¹H NMR indicating the PHA as a co-polymer of PHB-co-PHV, molecular weight by gel permeation chromatography as 2.08 × 10⁶, melting temperature 166.51 °C, tensile strength 18.8 MPa; two relaxations namely beta transition corresponding to the glass rubber transition and alpha transition corresponding to crystalline relaxation by Dynamic Mechanical Thermal analysis and only one relaxation corresponding to MWS interfacial polarisation with activation energy of 129 kJ/mol by broadband dielectric spectroscopy. Optical microscopic studies showed typical Maltese-cross pattern of spherulites. The PHA film was found to be biodegradable by standard ASTM method as well as by soil burial method. The leak proof polymer bags prepared from the film could be used as a packaging material.     

Use of glycerol for producing 1,3-dihydroxyacetone by Gluconobacter oxydans in an airlift bioreactor

1,3-Dihydroxyacetone can be produced by biotransformation of glycerol with glycerol dehydrogenase from Gluconobacter oxydans cells. Firstly, improvement the activity of glycerol dehydrogenase was carried out by medium optimization. The optimal medium for cell cultivation was composed of 5.6 g/l yeast extract, 4.7 g/l glycerol, 42.1 g/l mannitol, 0.5 g/l K₂HPO₄, 0.5 g/l KH₂PO₄, 0.1 g/l MgSO₄·7H₂O, and 2.0 g/l CaCO₃ with the initial pH of 4.9. Secondly, an internal loop airlift bioreactor was applied for DHA production from glycerol by resting cells of G. oxydans ZJB09113. Furthermore, the effects of pH, aeration rate and cell content on DHA production and glycerol feeding strategy were investigated. 156.3 ± 7.8 g/l of maximal DHA concentration with 89.8 ± 2.4% of conversion rate of glycerol to DHA was achieved after 72 h of biotransformation using 10 g/l resting cells at 30 °C, pH 5.0 and 1.5 vvm of aeration rate.     

Lactococcus lactis as a cell factory: a twofold increase in phosphofructokinase activity results in a proportional increase in specific rates of glucose uptake and lactate formation

Despite the fact that the area of glycolysis in Lactococcus lactis has been intensively studied, only a limited number of studies have been focused on the regulation of uptake of glucose itself. Using the tool of the glucostat fed-batch mode of culture, it was demonstrated in our earlier work that the concentration of glucose regulates its uptake rate and that the control of the glycolytic flux resides to a large extent in processes outside the pathway itself, like glucose transport and the ATP consuming reactions, while allosteric properties of key enzymes like phosphofructokinase (PFK) have a significant influence on the control. Extending our work, we report here the results of fermentations with engineered L. lactis strains with altered PFK activity in which the pfkA gene from Aspergillus niger, and its truncated version pfk13 that encodes a shorter PFK1 fragment were cloned. The results in this study suggest that, under the optimum for the microorganism applied microaerobic conditions, the glycolytic capacity of L. lactis was significantly increased in engineered strains with increased PFK activity. The transformant strain in which the truncated pfk13 gene of A. niger was expressed performed more efficiently as it was able to grow successfully in glucostat cultures with 277 mM glucose - while the optimum glucose concentration for the parental strain was 55 mM. The present work demonstrates the direct effect of PFK activity on the glycolytic flux in L. lactis since a twofold increase in specific PFK activity (from 7.1 to 14.5 U/OD₆₀₀) resulted in a proportional increase of the maximum specific rates of glucose uptake (from 0.8 to 1.7 μM s⁻¹ g CDW⁻¹) and lactate formation (from 15 to 22.8 g lactate (g CDW)⁻¹ h⁻¹).     

Effects of arbuscular mycorrhizal inoculation and fertilization on mycorrhizal Statute of Jacaranda mimosifolia D.Don cultivated in nurseries

The effects of fertilization and the nature of the inoculum as well as the variation of the dose intake of the latter on the level of Jacaranda mimosifolia D.Don mycorhization were tested. Young plants were treated with two inoculums presenting different origins, compositions and modes of application: one is a commercial product containing Glomus irregulare, and the other is a composite indigenous inoculum resulting from trapping five species of genus Glomus and also from multiplication on mycotrophic plants: leek (Allium porrum L.) and vetch (Vicia sativa L.). For each inoculum, two doses were tested and for each dose of inoculum, four levels of fertilization based on a complete commercial fertilizer (Osmocote) were tested: 0 g/plant, 2 g/plant, 4 g/plant, and 6 g/plant. Three repetitions were performed for each combination treatment of inoculum/fertilizer. One-year-old young Jacaranda plants, being about 40 cm high, were cultured under greenhouse in 10/12 cm caliber pots. After six months, all the inoculated plants were mycorrhized. According to endomycorrhizal structures found on their roots, plants receiving doses of composite indigenous inoculum reached a more advanced stage of mycorrhization than those treated with the commercial inoculum. The existence of an interaction effect between the inoculum dose and the level of fertilization on Jacaranda mycorhization rate was excluded. These two parameters of variation were studied as simple effects. The increase in commercial inoculum dose had a significant positive influence on the level of Jacaranda plants mycorrhization (P = 0.05). The rate of mycorrhization jumped from 12.69% to 21.92%. Nonetheless, for plants receiving increasing doses of composite indigenous inoculum, the level of mycorrhization has varied randomly. In both instances of inoculum treatments, increasing the dose of fertilizer significantly inhibited endomycorrhizal colonization of Jacaranda roots (P = 0.01). Thus, the rate of root colonization decreased from 47.43% to 2.41% for plants receiving the composite indigenous inoculums. It decreased from 32.35% to 3.95% for those treated with the commercial inoculum. Mycorrhization had a positive effect on root dry biomass of Jacaranda, as in the case of unfertilize ave the highest rates of colonization.     

Production of succinic acid from sucrose and sugarcane molasses by metabolically engineered Escherichia coli

Sucrose-utilizing genes (cscKB and cscA) from Escherichia coli KO11 were cloned and expressed in a metabolically engineered E. coli KJ122 to enhance succinate production from sucrose. KJ122 harboring a recombinant plasmid, pKJSUC, was screened for the efficient sucrose utilization by growth-based selection and adaptation. KJ122-pKJSUC-24T efficiently utilized sucrose in a low-cost medium to produce high succinate concentration with less accumulation of by-products. Succinate concentrations of 51 g/L (productivity equal to 1.05 g/L/h) were produced from sucrose in anaerobic bottles, and concentrations of 47 g/L were produced in 10 L bioreactor within 48 h. Antibiotics had no effect on the succinate production by KJ122-pKJSUC-24T. In addition, succinate concentrations of 62 g/L were produced from sugarcane molasses in anaerobic bottles, and concentrations of 56 g/L in 10 L bioreactor within 72 h. These results demonstrated that KJ122-pKJSUC-24T would be a potential strain for bio-based succinate production from sucrose and sugarcane molasses.     

Glutamic acid independent production of poly-γ-glutamic acid by Bacillus amyloliquefaciens LL3 and cloning of pgsBCA genes

A new glutamic acid independent poly-γ-glutamic acid (γ-PGA) producing strain, which was identified as Bacillusamyloliquefaciens LL3 by analysis of 16S rDNA and gyrase subunit A gene (gyrA), was isolated from fermented food. The product had a molecular weight of 470, 801 and l-glutamate monomer content of 98.47%. The pre-optimal medium, based on single-factor tests and orthogonal design, contained 50 g/L sucrose, 2 g/L (NH₄)₂SO₄, 0.6 g/L MgSO₄, and provided well-balanced changes in processing parameters and a γ-PGA yield of 4.36 g/L in 200 L system. The γ-PGA synthetase genes pgsBCA were cloned from LL3, and successfully expressed by pTrcLpgs vector in Escherichia coli JM109, resulting the synthesis of γ-PGA without glutamate. This study demonstrates the designedly improved yield of γ-PGA in 200 L system and the first report of pgsBCA from glutamic acid independent strain, which will benefit the metabolized mechanism investigation and the wide-ranging application of γ-PGA.     

Cardiodepressive effect elicited by the essential oil of Alpinia speciosa is related to L-type Ca current blockade

This study was undertaken to elucidate the effect of the essential oil from Alpinia speciosa (EOAs) on cardiac contractility and the underlying mechanisms. The essential oil was obtained from Alpinia speciosa leaves and flowers and the oil was analyzed by GC-MS method. Chemical analysis revealed the presence of at least 18 components. Terpinen-4-ol and 1,8-cineole corresponded to 38% and 18% of the crude oil, respectively. The experiments were conducted on spontaneously-beating right atria and on electrically stimulated left atria isolated from adult rats. The effect of EOAs on the isometric contractions and cardiac frequency in vitro was examined. EOAs decreased rat left atrial force of contraction with an EC₅₀ of 292.2 ± 75.7 μg/ml. Nifedipine, a well known L-type Ca²⁺ blocker, inhibited in a concentration-dependent manner left atrial force of contraction with an EC₅₀ of 12.1 ± 3.5 μg/ml. Sinus rhythm was diminished by EOAs with an EC₅₀ of 595.4 ± 56.2 μg/ml. Whole-cell L-type Ca²⁺ currents were recorded by using the patch-clamp technique. EOAs at 25 μg/ml decreased I_Ca,L by 32.6 ± 9.2% and at 250 μg/ml it decreased by 89.3 ± 7.4%. Thus, inhibition of L-type Ca²⁺ channels is involved in the cardiodepressive effect elicited by the essential oil of Alpinia speciosa in rat heart.     

Microbial oil production from rice straw hydrolysate by Trichosporon fermentans

Microbial oil production from sulphuric acid treated rice straw hydrolysate (SARSH) by Trichosporon fermentans was performed for the first time. Fermentation of SARSH without detoxification gave a poor lipid yield of 1.7 g/l, which was much lower than the result with glucose or xylose as the single carbon source (13.6 g/l or 9.9 g/l). The detoxification pretreatment, including overliming, concentration, and adsorption by Amberlite XAD-4 improved the fermentability of SARSH significantly by removing the inhibitors in SARSH. A total biomass of 28.6 g/l with a lipid content of 40.1% (corresponding to a lipid yield of 11.5 g/l) could be achieved after cultivation of T. fermentans on the detoxified SARSH for 8 days. Moreover, besides SARSH, T. fermentans could also utilize mannose, galactose, or cellobiose, in hydrolysates of other natural lignocellulosic materials as the single carbon source to grow and accumulate lipid with a high yield (at least 10.4 g/l). Hence, it is a promising strain for microbial oil production and thus biodiesel preparation from agro-industrial residues, especially lignocellulosic materials.     

Engineering Bacillus subtilis for acetoin production from glucose and xylose mixtures

As a vital flavor compound, acetoin is extensively used in dairy products and drinks industry. In this study, Bacillus subtilis was engineered to metabolize glucose and xylose as substrates for acetoin production. Initially, gene araE from B. subtilis, encoding the xylose transport protein AraE, was placed under the control of the constitutive promoter P₄₃ for over-expression. Batch cultures showed that 10 g/L xylose was depleted completely in 32 h. Subsequently, genes xylA and xylB from Escherichia coli, encoding xylose isomerase and xylulokinase respectively, were introduced into B. subtilis, and the recombinant turned out to assimilate glucose and xylose without preference. In shake-flask fermentations, 5.5 g/L acetoin with a yield of 0.70 mol (mol sugar)⁻¹ was obtained by the optimum strain BSUL13 under microaerobic conditions, which offered a metabolic engineering strategy on engineering microbe as cell factory for the production of high-valued chemicals from renewable resource.