One‐step production of 2,3‐butanediol from starch by secretory over‐expression of amylase in Klebsiella pneumoniae

BACKGROUND: 2,3‐Butanediol (2,3‐BD) is a valuable chemical that can be biosynthesized from many kinds of substrates. For commercial biological production of 2,3‐BD, it is desirable to use cheap substrate without pretreatment, such as starch. However, there have been few reports on the production of 2,3‐BD directly from starch. RESULTS: In this work, gene malS coding for α‐amylase (EC 3.2.1.1) precursor was inserted into plasmid pUC18K, and secretory over‐expression of α‐amylase was achieved by engineered Klebsiella pneumoniae. The extracellular recombinant amylase accelerated the hydrolyzation of starch, and one‐step production of 2,3‐BD from starch was carried out by engineered K. pneumoniae. A 2,3‐BD concentration of 3.8 g L^?1 and yield of 0.19 g 2,3‐BD g^?1 starch were obtained after 24 h fermentation. CONCLUSION: The one‐step production of 2,3‐BD from starch was achieved by secretory over‐expression of amylase in K. pneumoniae. This would simplify the process and reduce the production cost considerably by enabling use of starch with minimal pretreatment. Copyright © 2008 Society of Chemical Industry     

Structure and catalytic properties of Co/ porcelain in the synthesis of 2,3‐dihydrofuran

The cyclodehydration of 1,4‐butanediol over cobalt catalysts in the liquid phase is used for the production of 2,3‐dihydrofuran. The catalyst preparation parameters considered were the metal loading, precipitation pH and reduction temperature of cobalt salt. It was found that the use of Co(NO₃)₂ together with Na₂CO₃ in a 1:1 ratio yielded better catalysts. Under the conditions used in this study the optimum cobalt loading for the selective production of 2,3‐dihydrofuran is in the range 15–50 wt%. The optimum reduction temperature of Co/porcelain catalyst depends on cobalt loading. The optimum reduction temperatures for 15 and 50 wt% cobalt loading are 773 and 723 K (reduction time 20 min), respectively. © 2001 Society of Chemical Industry     

The reaction mechanism of diethyl‐2,3‐dicyano‐2,3‐di(p,m‐dimethoxylphenyl) succinate with styrene

The radical polymerization of styrene (ST) can be initiated by diethyl‐2,3‐dicyano‐2,3‐di(dimethoxyphenyl) succinate (ECPS). The reaction mechanism has been studied by means of UV, H¹‐NMR, product analysis, gel permeation chromatography, electronic spin resonance (ESR), and the conversion of monomer via time. These experimental results indicate that ECPS probably takes the place of complex with ST, and the complex interaction between ECPS and ST can take advantage of the dissociation of the C C bond. The complex interaction and thermal effect are the important factors causing the dissociation of C C bond. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 74: 1140–1145, 1999     

Optimization of agro‐residual medium for α‐amylase production from a hyper‐producing Bacillus subtilis KCC103 in submerged fermentation

BACKGROUND: Although submerged fermentation (SmF) is the conventional method in industry, use of low‐cost agro‐residues for α‐amylase production in SmF has not been well established. Here we optimized agro‐residue‐based medium and culture conditions for α‐amylase production in SmF using a hyper‐producing Bacillus subtilis KCC103. RESULTS: B. subtilis KCC103 produced α‐amylase in SmF by utilizing agro‐residues. Wheat bran (WB) and sunflower oil cake (SFOC) were selected as the best substrates using shake flasks. Medium containing WB (carbohydrate rich) and SFOC (rich in protein and free amino acids) at 1:1 (w/w) ratio produced high levels (90 IU mL⁻¹) of α‐amylase at 30–36 h in a shake flask. The α‐amylase yield was 14‐fold enhanced (1258 IU mL⁻¹) by optimizing process parameters and medium composition following response surface methodology in a bioreactor. The optimal conditions were: WB 1.27%, SFOC 1.42%, pH 7, 37 °C and 10–12 h. Both in shake flask and bioreactor α‐amylase synthesis was not repressed by the release of simple sugars into the medium. CONCLUSION: KCC103 with catabolite derepression and hyperproducing ability is useful for economic α‐amylase production using low‐cost agro‐residual substrates in conventional SmF. Since the production time (10–12 h) is much shorter than other strains this would improve productivity and further reduce the cost of α‐amylase production. Copyright © 2008 Society of Chemical Industry     

Pervaporative enrichment of 2,3‐butanediol from its mixture with 1‐butanol using a polydimethylsiloxane and ZSM‐5 mixed matrix membrane: Effects of ethanol as a by‐product

A ZSM‐5 filled polydimethylsiloxane membrane with 44.4 wt.% zeolite loading was used in the pervaporative removal of 1‐butanol from its mixtures with 1‐butanol. A small quantity of ethanol was added to the feed as a by‐product to test the response of the membrane. It was found that the permeation behaviour of other feed components was changed and membrane selectivity decreased. This change was attributed to the frequently‐observed inter‐component coupled transport in multi‐component feed systems. The impact of ethanol on recovery of 2,3‐butanediol was evaluated using a simulated continuous operation, which enriched 2,3‐butanediol to 99.5 wt.% from a feed containing 5 wt.% 2,3‐butanediol and less than 1.0 wt.% ethanol. It was observed that membrane selectivity improves as ethanol concentration decreases in the stream due to its preferential removal. The final recovery of 2,3‐butanediol was not significantly reduced as the concentration of ethanol was below 1.0 wt.%. © 2011 Canadian Society for Chemical Engineering     

Optimization of medium and process parameters for a constitutive α‐amylase production from a catabolite derepressed Bacillus subtilis KCC103

BACKGROUND: In Bacillus subtilis KCC103 α‐amylase is hyper‐produced and not catabolite repressed by glucose. Various sugars, raw starches and nitrogen sources were tested for their repression effect on α‐amylase synthesis. Enhancement of α‐amylase production by supplementing micronutrients and surfactants was studied. Using optimized medium, process parameters were optimized for improved α‐amylase production. RESULTS: α‐Amylase was produced from KCC103 utilizing simple sugars indicating the absence of catabolite repression. Raw potato and yeast extract were best carbon and nitrogen sources for α‐amylase production. α‐Amylase synthesis was enhanced by micronutrients cysteine, thiamine, Mg²⁺ and SDS. Maximum α‐amylase (394 IU mL^?1) was produced in the optimized medium consisting of (in g L^?1) raw potato (30.0), yeast extract (20.0), cysteine (0.3), thiamine (0.2), SDS (0.2) and MgSO₄ (0.5 mmol L^?1) at 36–48 h under optimal conditions (pH 7.0, 37 °C, 200 rpm). The α‐amylase production was further enhanced to 537.7 IU mL^?1 with shorter time (15–18 h) in a bioreactor with optimized agitation rate of 700 rpm at 30% dissolved oxygen. CONCLUSION: Since there was no carbon catabolite repression of α‐amylase synthesis, sugar mixture from various agro‐residues hydrolysates could be utilized for α‐amylase production. The study showed the feasibility of utilization of raw potato for α‐amylase production from the KCC103, which would lead to a significant reduction in process cost. Copyright © 2008 Society of Chemical Industry     

Fermentation characteristics of a low‐oxygen inducible hmp promoter system in Bacillus subtilis LAB1886

In an attempt to find a low‐cost promoter system which could be used for producing recombinant proteins, a low oxygen inducible hmp promoter system in Bacillus subtilis (LAB1886) was characterized by studying the effects of cell density (OD₆₀₀), glucose concentration, pH, dissolved oxygen (DO) level, nitrate and nitrite concentration on cell growth and β‐galactosidase expression. The optimal cultivation strategy was found to be to grow the cells in the presence of nitrate to OD₆₀₀ = 0.2 at a high DO level (80%), and then to induce the hmp promoter by lowering the DO level to 1–2%. As a result, the specific β‐galactosidase activity increased continuously to the maximal value of 1750 Miller Units. Nitrite, as the apparent inducer of the hmp promoter, was produced from nitrate reduction, and had profound effects on cell growth and β‐galactosidase expression. Copyright © 2006 Society of Chemical Industry     

Polymerization of methyl methacrylate with a thermal iniferter: Diethyl 2,3‐dicyano‐2,3‐di(p‐tolyl)succinate

A hexa‐substituted ethane type compound, diethyl‐2,3‐dicyano‐2,3‐di(p‐tolyl)succinate (DCDTS), was successfully synthesized and used for initiation of methyl methacrylate (MMA) polymerization. The reaction demonstrated the characteristics of a “living” polymerization; i.e., both the yield and the molecular weight of the resulting polymers increased linearly with increasing reaction time, the molecular‐weight distribution of PMMA obtained was ～1.60 and almost unaffected by the conversion, and the resultant polymer can be chain extended by adding fresh MMA. End group analysis of the resultant PMMA confirmed that DCDTS behaves as a thermal iniferter for MMA polymerization. A block copolymer was prepared from the resultant PMMA, which contains a hexa‐substituted C? C bond functional end group. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 80: 2566–2572, 2001     

Enrichment of glutaminase production by Bacillus subtilis RSP‐GLU in submerged cultivation based on neural network—genetic algorithm approach

BACKGROUND: Owing to the importance of glutaminase in biotech product production, its production with isolated Bacillus subtilis RSP‐GLU (MTCC 9727) was investigated. Fermentation factors play an important role in product enhancement. Hence, glutaminase production was optimized using an artificial neural network (ANN) coupled genetic algorithm (GA). RESULTS: A ‘6–12–1’ topology ANN was constructed to identify the nonlinear relationship between fermentation factors and enzyme yield. ANN‐predicted values were optimized for glutaminase production using a GA. The overall mean absolute predictive error (MAPE) and the mean square errors (MSE) were observed to be 0.00125 and 1.77 and 0.002 and 3.06 for training and testing, respectively. The goodness of neural network prediction (coefficient of R²) was found to be 0.996. The maximum interactive impact on glutaminase production was noted with rpm versus medium volume. The use of ANN–GA hybrid methodology resulted in a significant improvement (47%) in glutaminase yield. CONCLUSION: Five different optimum fermentation conditions out of 500 revealed maximum enzyme production. Glutaminase enzyme production in this Bacillus subtilis RSP‐GLU is strongly influenced by aeration of the fermentation. A hybrid ANN‐GA effectively identifies the different fermentation conditions for optimum production of enzyme in a given large set of conditions. Copyright © 2009 Society of Chemical Industry