• Journal of Life Science
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• v.23 no.12
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• pp.1471-1476
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• 2013

The present study deals with the immobilization of Kluyveromyces lactis ${\beta}$-galactosidase on a weak ionic exchange resin (Duolite A568) as polymer support. ${\beta}$-Galactosidase was immobilized using the adsorption method. A kinetic study of the immobilized enzyme was performed in a packed-bed reactor. The adsorption of the enzyme followed a typical Freundlich adsorption isotherm. The adsorption parameters of k and n were 14.6 and 1.74, respectively. The initial rates method was used to characterize the kinetic parameters of the free and immobilized enzymes. The Michaelis-Menten constant ($K_m$) for the immobilized enzyme (120 mM) was higher than it was for the free enzyme (79 mM). The effect of competitive inhibition kinetics was studied by changing the concentration of galactose in a recycling packed-bed reactor. The kinetic model with competitive inhibition by galactose was best fitted to the experimental results with $V_m$, $K_m$, and $K_I$ values of 46.3 $mmolmin^{-1}mg^{-1}$, 120 mM, and 24.4 mM, respectively. In a continuous packed-bed reactor, increasing the flow rate of the lactose solution decreased the conversion efficiency of lactose at different input lactose concentrations. Continuous operation of 11 days was conducted to investigate the stability of a long-term operation. The retained activity of the immobilized enzymes was 63% and the half-life of the immobilized enzyme was found to be 15 days.

• Journal of Applied Biological Chemistry
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• v.66
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• pp.512-518
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• 2023

To address inherent weaknesses such as low mechanical strength and limited enzyme loading capacity in conventional chitosan or alginate beads, an additional step involving the exchange of anionic surfactants with hydroxide ions was employed to prepare porous chitosan hydrogel capsules for enzyme immobilization. Consequently, excellent thermal stability and long-term storage stability were confirmed. Furthermore, coating the porous chitosan hydrogel capsules with polydopamine not only improved mechanical stability but also exhibited remarkable enzyme immobilization efficiency (97.6% for M1-D0.5). Additionally, it was demonstrated that the scope of application for chitosan hydrogel beads, prepared using conventional methods, could be further expanded by introducing an additional step of polydopamine coating. The enzyme immobilization matrix developed in this study can be selectively applied to suit specific purposes and is expected to be utilized as a support for the adsorption or covalent binding of various substances.

• Bulletin of the Korean Chemical Society
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• v.31 no.1
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• pp.157-161
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• 2010

A sensory element against carbaryl, as a widely used pesticide was prepared based on adsorbed acetylcholine esterase (AChE) from Torpedo california. Octadecyl was substituted on macro-porous silica, confirmed by infra-red (IR) spectroscopy and quantitatively estimated through thermo-gravimetric analysis (TGA). Immobilization of the enzyme was achieved by adsorption on this support. Activity of the immobilization product was measured as a function of the loaded enzyme concentration, and maximum binding capacity of the support was estimated to be 43.18 nmol.mg-1. The immobilized preparations were stable for more than two months at storage conditions and showed consistency in continuous operations. Possible application of the immobilized AChE for quantitative analysis of carbaryl is proposed in this study.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.5
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• pp.684-689
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• 2006

The present study examined the effects of Tween 80 on the attachment and hydrolytic activity of a cellulase enzyme against ball-milled cellulose (BMC), using the whole component (native CBH I) and the catalysis module (core CBH I) of carbohydrolase I purified from Trichoderma viride (Meicelase, Meiji Seika, Tokyo, Japan). The effects were evaluated as protein concentrations in the supernatant after mixing enzyme and substrate with Tween 80 at room temperature. Tween 80 decreased the adsorption of native CBH I and core CBH I onto BMC (p<0.001) and increased the amount of reducing sugars released from BMC by native CBH I (p<0.001). However, Tween 80 did not enhance the hydrolytic activity of core CBH I. Observations using SEM revealed that Tween 80 caused cellulose filter paper to swell and enhanced surface cracks and filaments caused by native CBH I but not by core CBH I. These results suggested that Tween 80 decreases enzyme adsorption to its substrate but enhances enzymatic activity.

• Journal of Microbiology and Biotechnology
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• v.2 no.3
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• pp.197-203
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• 1992

Studies of the pretreatment of chitin and its subsequent hydrolysis by Aspergillus carneus chitinase are reported. Ball milling was found to be the most effective way among the pretreatment methods tested. Data are presented describing the effect of enzyme and substrate concentrations on the rate and extent of the hydrolysis process. It was found that the successive addition of enzyme improved the saccharification yield. Significant product inhibition of the chitinase was observed when N-acetylglucosamine concentration was 3.6％ or higher. Adsorption of enzymes to the substrate occurred during a 24 hr hydrolysis period. An initial rapid and extensive adsorption occurred, followed by gradual desorption which increased during the time of reaction. Intermediate removal of the hydrolyzate and continuation of the hydrolysis by adsorbed enzyme on the residual chitin was also investigated. A total of 75.4 g/l reducing sugars, corresponding to 69.2％ saccharificaton yield (as N-acetylglucosamine) was obtained. In addition an increase in the amount of recoverable enzymes was observed under these conditions. Evidence presented here suggests that the technique, whereby the free enzymes in the recovered hydrolyzate are re-adsorbed onto the new substrate, may provide a means of recirculating the dissolved enzymes.

• Microbiology and Biotechnology Letters
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• v.24 no.6
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• pp.678-686
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• 1996

To improve the preservation of Kimchi, we isolated Lactobacillus plantarum lytic enzyme-producing strain from soil, and the enzyme was purified and characterized. From the observation of cultural and morpho- logical characteristics, the isolated strain was identified as Metarrisium anisopliae (Metschn.) Sorok. The enzyme was purified to 75-folds with 40% yields through affinity adsorption and CM-Sephadex C-50 column chromatog- raphy. The optimum pH and temperature for lytic activity are 4.0 and 40$\circ$C, respectively, and the enzyme acitvity is stable between pH 3.0 and 9.0, and up to 50$\circ$C. The enzyme is a monomeric protein with molecular weight of 40,000 daltons by SDS-PAGE and gel filtration. The enzyme is endopeptidase which breaks the peptide linkage of Lactobacillus plantarum peptidoglycan. The lytic action spectra confirmed that Leuconostoc mesenteroides, a useful strain for the fermentation of Kimchi, is not lysed by the enzyme. The enzyme activity is inhibited by N-bromosuccinimide (NBS), which probably indicates the involvement of tryptophan residue in active site of the enzyme, and also inhibited by Ag$^{+}$. The amino acid composition analysis showed that the enzyme contains more acidic amino acids than basic ones, and composition of alanine, glycine, proline and tyrosines was very high.

• Microbiology and Biotechnology Letters
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• v.24 no.4
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• pp.445-451
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• 1996

The strain YCH-37, which produces yeast cell wall lytlc enzyme, was isolated from soil. From the microscopic observation, morphological and cultural characteristics, this strain was identified to fungus, Dicyma sp. So, we named this strain as Dicyma sp. YCH-37. The lytic enzyme effectively lysed Salmonella typhimurium among intact living bacteria and Torulopsis, Hansenula, Zygosaccharomyces among intact living yeast, as well as autoclaved yeast strains. The yeast cell wall lytic enzyme was succesively purified to 204 folds with 13% yields through yeast glucan affinity adsorption and DEAE-cellulose column chromatography. The enzyme was identified to monomeric protein with molecular weight of 25,000 daltons from the results of SDS-PAGE and gel filtration. The optimum pH and temperature for the yeast lytic activity were 8.0 and 50$\circ$C, respectively. The enzyme was stable up to 40$\circ$C, and between pH 4.0-pH 10.0.

• Microbiology and Biotechnology Letters
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• v.18 no.3
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• pp.260-267
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• 1990

The mechanism of enzymatic hydrolysis of raw corn starch by the purified glucoamylase and a - amylase in an agitated bead reaction system was studied by investigating the changes of sugar profiles produced by each enzyme, the granular structure of raw corn starch, the amount of enzyme adsorption on residual starch, and the amylose content in residual raw starch. The sugar profiles produced by the action of exo-type glucoamylase or endo-type $\alpha$ -amylase in an agitated bead system were not recognizably differed with those produced in reaction system without bead. Without enzyme the intergenic microcrystalline structure of starch granule was not changed by the simple mechanical impact of solid media, but it was cleaved. However, starch granule was fragment into large number of small particles by the synergistic action of enzyme and attrition-milling media, identified to be the major saccharification enhancing mechanism along with the increased amount of enzyme adsorption. The amylose content decreased more readily in an agitated bead reaction system, especially by $\alpha$ -amylase.

• Journal of the Korean Wood Science and Technology
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• v.29 no.4
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• pp.67-78
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• 2001

Two species(Quercus mongolica, populus euramericana) of hardwood chips were subjected to steam explosion 25 kg/$cm^2$, for 6 min. The exploded woods were treated by the single or multi-stage chemical process with sodium hydroxide, sodium hypochlorite and sodium chlorite. The multi-stage treatment of exploded wood can be successfully removed lignin. Enzymatic hydrolysis rate of substrate varied from 25% for exploded wood to about 80% for the multi-chemical treated exploded wood. The enzymatic susceptibility was different among wood species. The multi chemical treatment of the exploded wood resulted in the high rate of glucose in the enzymatic hydrolyzate. Cellulase adsorption increased at high lignin content of substrates, while crystallinity, pore area and specific surface area of substrates did not affected enzyme adsorption. According to the proposed pretreatment and saccharification process in this study, it can be acquired about 37~40 kg of glucose from 100 kg of hardwood.

• KSBB Journal
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• v.13 no.2
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• pp.147-154
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• 1998

Enzymatic hydrolysis using an immobilized enzyme was carried out to produce chitosan oligosaccharides (COSs) from chitosan effectively. Chitosanase was immobilized on eight different carriers by physical adsorption. The enzyme immobilized on chitin had higher activity than those immobilized on the other carriers in spite of its lower adsorption. The activity of chitin-immobilized enzyme was more than 90% of the original activity. Optimal temperature of the immobilized enzyme increased by about $15^{\circ}C$ and its thermostability was excellent in relatively wide range of temperature. But its effects of pH did not improve compared to the free enzyme. The immobilized enzyme produced 153 mg/g chitosan of the reducing sugar for 3hrs of hydrolytic incubation time. The total content of higher oligomers, tetramer to hexamer, among amount of total COSs obtained for 2hrs was more than 90%. In kinetic parameters for both enzymes, immobilized enzyme showed lower affinity for substrate and reaction rate than free enzyme, however, no reduction of the rate for high substrate concentrations. Consequently, chitin-immobilized could effectively hydrolyse chitosan and produce the higher COSs without activity decrease in comparison with the free enzyme.