Eui Yong Kim

Synthesis and characterization of a novel extracellular polysaccharide by Rhodotorula glutinis.

The aim of this work was to characterize an exopolysaccharide by Rhodotorula glutinis KCTC 7989 and to investigate the effect of the culture conditions on the production of this polymer. The extracellular polysaccharide (EPS) produced from this strain was a novel acidic heteropolysaccharide composed of neutral sugars (85%) and uronic acid (15%). The neutral sugar composition was identified by gas chromatography as mannose, fucose, glucose, and galactose in a 6.7:0.2:0.1:0.1 ratio. The molecular weight of purified EPS was estimated to be 1.0−3.8×105 Dalton, and the distribution of the molecular weight was very homogeneous (polydispersity index =1.32). The EPS solution showed a characteristic of pseudoplastic non-Newtonian fluid at a concentration >2.0% in distilled water. The maximum EPS production was obtained when the strain was grown on glucose (30 g/L). Ammonium sulfate was the best suitable nitrogen source for EPS production. The highest yield of EPS was obtained at a carbon to nitrogen ratio of 15. The EPS synthesis was activated at the acidic range of pH 3.0–5.0 and increased when the pH of the culture broth decreased naturally to <2.0 during the fermentation. When the yeast was grown on glucose (30 g/L) and ammonium sulfate (2 g/L) at 22°C at an initial pH of 4.0, EPS production was maximized (4.0 g/L), and the glucose-based production yield coefficient and carbon-based production yield coefficient were 0.30 g of EPS/g of glucose and 0.34 g (carbon of EPS)/g (carbon of glucose), respectively.

Optimization of protease immobilization by covalent binding using glutaraldehyde

Immobilization of a protease, Flavourzyme, by covalent binding on various carriers was investigated. Lewatit R258-K, activated with glutaraldehyde, was selected among the tested carriers, because of the highest immobilized enzyme activity. The optimization of activation and immobilization conditions was performed to obtain high recovery yield. The activity recovery decreased with increasing carrier loading over an optimal value, indicating the inactivation of enzymes by their reaction with uncoupled aldehyde groups of carriers. The buffer concentrations for carrier activation and enzyme immobilization were optimally selected as 500 and 50 mM, respectively. With increasing enzyme loading, the immobilized enzyme activity increased, but activity recovery decreased. Immobilization with a highly concentrated enzyme solution was advantageous for both the immobilized enzyme activity and activity recovery. Consequently, the optimum enzyme and carrier loadings for the immobilization of Flavourzyme were determined as 1.8 mg enzyme/mL and 0.6 g resin/mL, respectively.

Steady-State Oxidation Model by Horseradish Peroxidase for the Estimation of the Non-Inactivation Zone in the Enzymatic Removal of Pentachlorophenol

A theoretical model for the rate of oxidation of pentachlorophenol (PCP) catalyzed by horseradish peroxidase (HRP), was investigated to account for the influence of hydrogen peroxide (H2O2) concentration on the catalytic activity. To evaluate the maximum allowable H2O2 concentration, a relatively simple steady-state model was developed based on the Ping-Pong Bi-Bi mechanism considering the effect of excess H2O2. Several sets of experimental data obtained from batch reactions using an equimolar concentration of H2O2 and PCP were used to estimate the kinetic parameters by a nonlinear regression method. The model profiles acquired using the estimated parameters were in good agreement with experimental data at different initial enzyme and substrate concentrations. The best-fitted parameters were used to predict the initial rate of the enzyme reaction. The model prediction was coincident with the experimental results of other studies, indicating that the proposed model could be used for the optimization of reaction conditions. The maximum allowable H2O2 concentration to prevent H2O2 inhibition was calculated from the proposed model equation: [H2O2](0,max) = (square root)KmH2O2Ki[PCP]0/KmPCP+[PCP]0. Using this equation, a curve depicting the non-inactivation zone for the two substrates (hydrogen peroxide and PCP) was plotted and it could be used for experimental design and optimal process operation. To minimize enzyme inactivation by H2O2, it was determined that the concentration of H2O2 should be lower than 2.78 mM, regardless of the stoichiometric ratio.

Effect of phenol on Β-carotene content in total carotenoids production in cultivation ofRhodotorula glutinis

The composition of carotenoids produced byR. glutinis was observed to be dependent upon the addition of phenol into medium. A stimulatory effect of phenol on Β-carotene ofRhodotorula glutins K-501 grown on glucose was investigated. Carotenoids produced byRhodotorula glutinis K-501 were identified to torularhodin, torulene and Β-carotene, whose composition was 79.5%, 6.4% and 14.1%, respectively. The Β-carotene content increased up to 35% when phenol was added to culture media at 500 ppm. The ratio of torularhodin decreased with increasing phenol concentration, while torulene content was almost constant.

Enzymatic oxidation of aqueous pentachlorophenol

The influence of the nonionic surfactant Tween 80 on pentachlorophenol (PCP) oxidation catalyzed by horseradish peroxidase was studied. The surfactant was tested at concentrations below and above its critical micelle concentration (CMC). Enhancement of PCP removal was observed at sub-CMCs. The presence of Tween 80 in the reaction mixture reduced enzyme inactivation which occurred through a combination of free radical attack and sorption by precipitated products. A simple first-order model was able to simulate time profiles for enzyme inactivation in the presence or absence of Tween 80. At supra-CMCs, the surfactant caused noticeable reductions in PCP removal, presumably through micelle partitioning of PCP which precluded the hydrophobic PCP molecule from interacting with the enzyme.

Removal of aqueous pentachlorophenol by horseradish peroxidase in the presence of surfactants

An important issue in the oxidation of pentachlorophenol (PCP) by the enzyme horseradish peroxidase (HRP) is enzyme inactivation during the reaction. This study was initiated to investigate the ability of two nonionic surfactants (Tween 20 and Tween 80) to mitigate HRP inactivation. The surfactants were tested at concentrations below and above their critical micelle concentrations (CMCs). Enhancement of PCP oxidation was observed at sub-CMCs, indicating effective protection of HRP by the two surfactants. Maximum levels of PCP removal were observed when the concentrations of Tween 20 and Tween 80 were 40 and 50% of the CMCs, respectively. At supra-CMCs, both surfactants caused a noticeable reduction in the extent of PCP removal.

Improved immobilization yields by addition of protecting agents in glutaraldehyde-induced immobilization of protease

The effects of peptides and polyethylene glycol (PEG) on the immobilization of protease using glutaraldehyde-activated Lewatit R258-K resin were investigated. When soy peptide was added at 0.1 g/l, the activity recovery and effectiveness factor increased by 20% and 38%, respectively. When PEG4000 (0.5 g/l) was used as a protecting agent, the activity recovery and effectiveness factor were improved by 35% and 60%, respectively, leading to an increased specific activity of the immobilized enzyme. The addition of PEG4000 is suggested to improve the cost-effectiveness for the production of immobilized enzymes.

Removal of Ammonia from a Biologically Treated Pesticide Wastewater by Struvite Precipitation

The aim of this study was to investigate ammonia removal from the effluent of a full-scale biofilm reactor treating pesticide wastewater through the formation of struvite (MgNH4PO4•6H2O). The requirement of magnesium and phosphate ions for precipitation was met by adding MgO and Na2HPO4. Different NH4+:Mg2+:PO43- molar ratios were tested in batch reactors to optimize the precipitation conditions. Ammonia was reduced to 25 mg/L from an initial concentration of 210 mg/L at the highest ratio tested (1:1.2:1.3), equating to 88% removal. However, the extent of ammonia removal was only slightly affected when the N:Mg ratio was reduced from 1:1.2 to 1:0.6; dropping to 83% from 88% on a percentage basis. Struvite precipitation can be easily incorporated within the existing post-treatment section of the full-scale plant, providing the pesticide manufacturing facility with a reliable backup process for its nitrification system.

Biopolymer Production by the Yeast Rhodotorula glutinis

In this work, exopolysaccharide (EPS) production by the yeast Rhodotorula glutinis was investigated. The results suggest that the synthesis of EPS in batch fermentations was enhanced by using a low C/N ratio to stimulate fast cell growth during the exponential phase and by adding glucose to stationary phase cultures to provide excess carbon for EPS formation. The apparent viscosity and molecular weight of EPS were found to decrease with increasing fermentation time due to the partial hydrolysis of EPS by the acidic fermentation broth (pH 1.8). Maintaining stationary phase cultures at pH 4 was found to be effective in minimizing acid hydrolysis and producing EPS with high molecular weight and apparent viscosity.