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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.133, #3)

Crosslinked penicillin acylase aggregates for synthesis of β-lactam antibiotics in organic medium by Andrés Illanes; Lorena Wilson; Eduardo Caballero; Roberto Fernández-Lafuente; José Manuel Guisán (pp. 189-202).

Crosslinked enzyme aggregates (CLEAs) of a partially purified penicillin acylase from a recombinant Escherichia coli strain have been produced as a novel type of biocatalysts well endowed to perform in organic media. Different protein precipitants were studied and glutaraldehyde was used as the crosslinking agent. Precipitation curves were obtained for all precipitants to determine the concentrations at which all the protein precipitated out of the solution. The effect of the glutaraldehyde-to-protein ratio was studied with respect to process recovery and the specific activity and stability of the biocatalyst. Recovery of penicillin acylase activity was moderately high, about 50%; major losses of enzyme activity were produced at the precipitation step. Specific activities of all CLEAs were very high, which is one of the advantages of using nonsupported biocatalysts. Ammonium sulfate and tert-butyl alcohol were the best precipitants at a glutaraldehyde-protein mass ratio of 2 and were selected to perform the kinetically controlled synthesis of ampicillin in 60% (v/v) ethylene glycol medium. At comparable conversion yields, volumetric and specific antibiotic productivity were much higher for CLEAs than for carrier-bound penicillin acylases.

Keywords: Crosslinked enzyme aggregates; enzyme immobilization; penicillin acylase; ampicillin; β-lactam antibiotics; organic cosolvents

Production of ceramide with Saccharomyces cerevisiae by Kyu Hyuk Kwun; Jung-heon Lee; Kyung-ho Rho; Hyun-shik Yun (pp. 203-210).

The possibility of producing the biologically active material of the skin, ceramide, was studied using yeasts. The yeast strain that produced the most ceramide, Saccharomyces cerevisiae (KCCM 50515), was selected, and the optimal conditions for ceramide production were determined using shakeflask culture and batch fermentation. By measuring the production rate of ceramide at various pH values and temperatures, the optimal conditions for ceramide production were found to be pH 6.0 and 30°C. When heat shock was applied to the cells for 1 h by increasing the culture temperature from 30 to 40°C after cell growth, the amount of ceramide produced was increased 5.9-fold. A cell growth and ceramide production model was developed with Monod kinetics and the Leudecking-Piret model. It showed that ceramide production was increased when the cells were in the stationary phase.

Keywords: Ceramide; heat shock; optimal condition; Saccharomyces cerevisiae

Structure-function relations of carbohydrates by neoglycolipid arrays by Gang-Liang Huang; Hua-Liang Huang; Hou-Cheng Zhang; Peng-George Wang (pp. 211-215).

The work presented herein is a new noncovalent glycoarray assembly method for microplates created by simply mixing together a carbohydrate and a teradecylamine. α-d-Mannopyranoside, α-d-glucopyranoside, and α-d-galactopyranoside were utilized in model studies and product formations were detected by lectin binding. The method can be extended to study the steric hindrance effect of carbohydrate-protein interactions, namely the structure-function relations of carbohydrates.

Keywords: Glycoarray; neoglycolipid; structure-function relations

Recycling of distillery effluents in alcoholic fermentation by E. Morin Couallier; T. Payot; A. Pastore Bertin; M. L. Lameloise (pp. 217-237).

In beet distilleries, condensates arising from stillage concentration could be recycled as dilution water for the fermentation step, thus preserving groundwater resources and ensuring a quality-controlled water supply. However, the recycling of condensates has been found to cause a significant reduction in fermentation activity. This study aimed to verify that condensates are toxic to alcoholic fermentation. Ten compounds found in condensates (formic, acetic, propionic, butyric, valeric, and hexanoic acids; 2,3-butanediol, furfuryl alcohol, furfural, and 2-phenyl-ethyl-alcohol) were tested. With the exception of 2,3-butanediol, they all proved to be inhibitors. At the same molar concentration,the longer the carbonaceous chain, the stronger the inhibition by fatty acids. An experimental design was used to study the inhibitory characteristics of the 10 compounds at the concentrations found in condensates. Synergistic effects were also confirmed. In real effluents, acetic acid was so highly concentrated that it became the strongest inhibitor. It is therefore necessary to eliminate it before recycling, as well as less concentrated compounds that may accumulate, as illustrated by the simulation.

Keywords: Alcoholic fermentation; inhibition; condensates; effluent; recycling; fatty acids; 2,3-butanediol; furfuryl alcohol; furfural; 2-phenylethyl-alcohol

Phytase production by thermophilic mold sporotrichum thermophile in solid-state fermentation and its application in dephytinization of sesame oil cake by Bijender Singh; T. Satyanarayana (pp. 239-250).

The phytase production by Sporotrichum thermophile TLR50 was recorded on all the commonly used animal feed ingredients tested to varying degrees in solid-state fermentation. Enzyme production increased to 180 U/g of dry moldy residue (DMR) in sesame oil cake at 120 h and 45°C at the initial substrate-to-moisture ratio of 1∶2.5 and a w of 0.95. Supplemenetation of sesame oil cake with glucose and ammonium sulfate further enhanced phytase titer (282 U/g of DMR). An overall 76% enhancement in phytase production was achieved owing to optimization. The mold secreted acid phosphatase, amylase, xylanase, and lipase along with phytase. By the action of phytase, inorganic phosphate was liberated efficiently, leading to dephytinization of sesame oil cake.

Keywords: Phytase; thermophilic mold; Sporotrichum thermophile TLR50; sesame oil cake; solid-state fermentation; dephytinization

Lactic acid production from cellulosic material by synergetic hydrolysis and fermentation by Xueliang Shen; Liming Xia (pp. 251-262).

The hydrolysis process on corncob residue was catalyzed synergetically by the cellulase from Trichoderma reesei and the immobilized cellobiase. The feedback inhibition to cellulase reaction caused by the accumulation of cellobiose was eliminated efficiently. The hydrolysis yield of corncob residue was 82.5%, and the percentage of glucose in the reducing sugar reached 88.2%. The glucose in the cellulosic hydrolysate could be converted into lactic acid effectively by the immobilized cells of Lactobacillus delbrueckii. When the enzymatic hydrolysis of cellulose and the fermentation of lactic acid were coupled together, no glucose was accumulated in the reaction system, and the feedback inhibition caused by glucose was also eliminated. Under the batch process of synergetic hydrolysis and lactic acid fermentation with 100 g/L of cellulosic substrate, the conversion efficiency of lactic acid from cellulose and the productivity of lactic acid reached 92.4% and 0.938 g/(L·h), respectively. By using a fed-batch technique, the total concentration of cellulosic substrate and lactic acid in the synergetic process increased to 200 and 107.5 g/L, respectively, whereas the dosage of cellulase reduced from 20 to 15 IU/g of substrate in the batch process. The results of the bioconversion of renewable cellulosic resources were significant.

Keywords: Cellulase; immobilized cellobiase; cellulosic material; corncob; synergetic hydrolysis; lactic acid fermentation

Effect of aeration rate on production of xylitol from corncob hemicellulose hydrolysate by Xinghong Ding; Liming Xia (pp. 263-270).

The effects of different aeration conditions on xylitol production from corncob hemicellulose hydrolysate by Candida sp. ZU04 were investigated. Batch fermentations were carried out in a 3.7-L fermentor at 30°C, pH5.5, and agitation of 300 rpm. It was found that the two-phase aeration process was more effective than the one-phase aeration process in xylitol production. In the first 24h of the aerobic phase, a high aeration rate was applied, glucose was soon consumed, and biomass increased quickly. In the second fermentation phase, aeration rate was reduced and an improved xylitol yield was obtained. The maximum xylitol yield (0.76 g/g) was obtained with an aeration rate of 1.5 vvm (KLa of 37 h⁻¹) for the first 24 h and 0.3 vvm (KLa of 6 h⁻¹) from 24 to 96 h.

Keywords: Corncob; fermentation; hemicellulose hydrolysate; xylitol; aeration rate

Efficient production of poly-γ-glutamic acid by bacillus subtilis ZJU-7 by Feng Shi; Zhinan Xu; Peilin Cen (pp. 271-281).

A strain with high poly-γ-glutamic acid (γ-PGA) production was isolated from fermented bean curd, a traditional Chinese food. The strain was named Bacillus subtilis ZJU-7 according to 16s rDNA sequencing and its taxonomic characters. The culture conditions for γ-PGA production were evaluated. The most suitable carbon and nitrogen sources were sucrose and tryptone, respectively. Exogenous l-glutamic acid was necessary for γ-PGA production, and the production of γ-PGA increased on the addition of l-glutamic acid to the medium. In the medium containing 60 g/L of sucrose, 60 g/L of tryptone, 80 g/L of l-glutamic acid, and 10 g/L of NaCl, the yield of γ-PGA reached 54.4 g/L after cultivation at 37°C for 24h, which was the highest γ-PGA production compared with values reported in the literature. The average molecular mass of γ-PGA produced was about 1.24×10⁶ Daltons. B. subtilis ZJU-7 is genetically stable and can synthesize levan instead of γ-PGA without the addition of l-glutamic acid to the medium.

Keywords: Poly-γ-glutamic acid; 16s rDNA; l-glutamic acid; sucrose; Bacillus subtilis

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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.133, #3)

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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.133, #3)