Immobilization of Yeast Alcohol Dehydrogenase on Weakly Basic Anion Exchange Resin Beads

Yeast alcohol dehydrogenase was immobilized on weakly basic macroporous anion exchange resin beads Lewatit MP-64. After the adsorption the enzyme was crosslinked by glutaraldehyde: The activity of the immobilized enzyme was investigated in the pH 8.9 recirculation reactor system at 303 K. It was found that the immobilized enzyme was destabilized upon addition of semicarbazide hydrochloride to the buffer solution. A greater amount of protein was attached to the support when ethanol was present in the enzyme solution, but the activity of the bound enzyme was lower than in the absence of ethanol

Immobilization of Yeast Alcohol Dehydrogenase on Weakly Basic Anion Exchange Resin Beads

Yeast alcohol dehydrogenase was immobilized on weakly basic macroporous anion exchange resin beads Lewatit MP-64. After the adsorption the enzyme was crosslinked by glutaraldehyde: The activity of the immobilized enzyme was investigated in the pH 8.9 recirculation reactor system at 303 K. It was found that the immobilized enzyme was destabilized upon addition of semicarbazide hydrochloride to the buffer solution. A greater amount of protein was attached to the support when ethanol was present in the enzyme solution, but the activity of the bound enzyme was lower than in the absence of ethanol

Modelling and Optimization of the (R)-(+)-3,4-dihydroxyphenyllactic Acid Production Catalyzed with D-lactate Dehydrogenase from Lactobacillus leishmannii Using Genetic Algorithm

A mathematical model for the enzymatic kinetics of the synthesis of (R)–(+)–3,4-dihydroxyphenyllactic acid (DHPL) was developed. The synthesis was catalyzed by D-lactate dehydrogenase from Lactobacillus leishmannii. Since this enzyme requires NADH as a coenzyme, formate dehydrogenase system was used for NADH regeneration. Kinetic constants of both enzymes were estimated independently from initial reaction rate experiments. The developed mathematical model was verified by the batch reactor experiment (volumetric productivity in this experiment was 4.76 g dm–3 d–1). Optimization of initial reaction conditions for DHPL synthesis was performed using the genetic algorithm (GA). The genetic algorithm as a flexible optimization tool had been used to obtain the experimental conditions where maximal volumetric productivity could be achieved. The optimal initial conditions were found in the investigated parameter area: c3,4-dihydroxyphenylpyruvic acid = 4.69 mmol dm–3, cNAD+= 4.95 mmol dm–3, cformate = 36.85 mmol dm–3, D-lactate dehydrogenase = 3 mg cm–3, formate dehydrogenase = 2.94mg cm–3 and the reaction time 8.5 min. At these conditions volumetric productivity of 93.06 g dm–3 d–1 can be achieved

Stabilization of D-Amino Acid Oxidase via Covalent Immobilization and Mathematical Model of D-Methionine Oxidative Deamination Catalyzed by Immobilized Enzyme

Porcine kidney D-amino acid oxidase was stabilized by covalent immobilization on spherical particles of Eupergit C because of its low stability in soluble form. The focus of this work was to evaluate operational stability of the immobilized enzyme. To evaluate D-amino acid oxidase’s operational stability during process conditions, repetitive batch reactor experiments of D-methionine oxidation reaction were carried out with continuous aeration for oxygen supply at air-flow rates of 5 and 10 dm3 h–1. Kinetic analysis of the immobilized enzyme was done as well. The mathematical model of D-methionine oxidative deamination catalyzed by the immobilized D-amino acid oxidase was developed and it described the data well. It enabled the estimation of operational stability decay rate constant. It was possible to achieve 100 % substrate conversion in all batch experiments

White-rot fungi in phenols, dyes and other xenobiotics treatment – a brief review

Bioremediation is an attractive technology that utilizes the metabolic potential of microorganisms in order to clean up the environmental pollutants to the less hazardous or non-hazardous forms with less input of chemicals, energy and time. White-rot fungi are unique organisms that show the capacities of degrading and mineralizing lignin as well as organic, highly toxic and recalcitrant compounds. The key enzymes of their metabolism are extracellular lignolytic enzymes that enable fungi to tolerate a relatively high concentration of toxic substrates. This paper gives a brief review of many aspects concerning the application of white-rot fungi with the purpose of the industrial contaminants removal

Modelling and Optimization of the (R)-(+)-3,4-dihydroxyphenyllactic Acid Production Catalyzed with D-lactate Dehydrogenase from Lactobacillus leishmannii Using Genetic Algorithm

A mathematical model for the enzymatic kinetics of the synthesis of (R)–(+)–3,4-dihydroxyphenyllactic acid (DHPL) was developed. The synthesis was catalyzed by D-lactate dehydrogenase from Lactobacillus leishmannii. Since this enzyme requires NADH as a coenzyme, formate dehydrogenase system was used for NADH regeneration. Kinetic constants of both enzymes were estimated independently from initial reaction rate experiments. The developed mathematical model was verified by the batch reactor experiment (volumetric productivity in this experiment was 4.76 g dm–3 d–1). Optimization of initial reaction conditions for DHPL synthesis was performed using the genetic algorithm (GA). The genetic algorithm as a flexible optimization tool had been used to obtain the experimental conditions where maximal volumetric productivity could be achieved. The optimal initial conditions were found in the investigated parameter area: c3,4-dihydroxyphenylpyruvic acid = 4.69 mmol dm–3, cNAD+= 4.95 mmol dm–3, cformate = 36.85 mmol dm–3, D-lactate dehydrogenase = 3 mg cm–3, formate dehydrogenase = 2.94mg cm–3 and the reaction time 8.5 min. At these conditions volumetric productivity of 93.06 g dm–3 d–1 can be achieved

Modelling of Continuous L-Malic Acid Production by Porcine Heart Fumarase and Fumarase in Yeast Cells

Continuous production of L-malic acid will be presented in this paper. The fumarase isolated from porcine heart, fumarase in the permeabilized non-growing cells of baker’s yeast and Saccharomyces bayanus (UVAFERM BC) were used as biocatalysts. In the production of L-malic acid with fumarase isolated from porcine hearts, there was no enzyme deactivation for a period of two days.At the average residence time of 4 hours, the conversion of about 80 % was achieved.Inactivation of the enzyme was observed using permeabilized cells.Thi s inactivation is described as a reversible process.C onversion of about 50 % was achieved with the remaining enzyme activity.A mathematical model that describes the production of L-malic acid, which contains the enzyme inactivation rate, was developed. Based on simulations, the used biocatalysts were compared. The results show that in the continuous production of L-malic acid, one milligram of purified enzyme corresponds to 68 g (wet weight) cells of Saccharomyces bayanus or 120 g (wet weight) cells of baker’s yeast

Overview on Reactions with Multi-enzyme Systems

Production of special chemicals and pharmaceuticals includes multi-step procedures that are still carried out in the traditional way: by isolating the intermediary product of each step and using it as a substrate for the next step, which is money and time consuming. These are also procedures that require many chemicals, energy and labour. Over the last couple of decades scientists have been working on new, integrated processes that require fewer resources and are more close to nature, as they produce less waste. These kinds of processes are discussed in this paper. The advantages of enzyme catalyzed reactions are well documented and numerous. Enzyme reactions are carried out at mild reaction conditions. The enzymes are enantioselective and stereoselective, which is important particularly for the pharmaceutical industry. By combining the action of different enzymes we can imitate the processes in the living cells and produce the desired compounds. Enzyme catalyzed reactions can also be combined with chemical reactions in one-pot chemo-enzymatic synthesis. The progress in the development of these reactions will be presented

Modelling of L-DOPA Oxidation Catalyzed by Laccase

Enzymatic oxidation of 3,4-dihydroxyphenyl-L-alanine (L-DOPA) with laccase from Trametes versicolor was investigated. The highest enzyme activity at pH 5.4 and at 25 ºC was found. The reaction kinetics and the effect of dissolved oxygen concentration on the reaction rate were evaluated. A mathematical model, comprised of double-substrate Michealis-Menten kinetics and mass balances for L-DOPA and dissolved oxygen concentrations, was developed in order to describe and predict the process of L-DOPA oxidation. Kinetic parameters, , and were estimated and experimentally verified by a set of experiments with constant additional aeration for different initial concentrations of L-DOPA and dissolved oxygen. A significant increase in reaction rate was established at a higher oxygen concentration in the inlet gas. The developed model was used to investigate the influence of dissolved oxygen concentration on L-DOPA conversion

Modelling of L-DOPA Oxidation Catalyzed by Laccase

Enzymatic oxidation of 3,4-dihydroxyphenyl-L-alanine (L-DOPA) with laccase from Trametes versicolor was investigated. The highest enzyme activity at pH 5.4 and at 25 ºC was found. The reaction kinetics and the effect of dissolved oxygen concentration on the reaction rate were evaluated. A mathematical model, comprised of double-substrate Michealis-Menten kinetics and mass balances for L-DOPA and dissolved oxygen concentrations, was developed in order to describe and predict the process of L-DOPA oxidation. Kinetic parameters, , and were estimated and experimentally verified by a set of experiments with constant additional aeration for different initial concentrations of L-DOPA and dissolved oxygen. A significant increase in reaction rate was established at a higher oxygen concentration in the inlet gas. The developed model was used to investigate the influence of dissolved oxygen concentration on L-DOPA conversion