• Journal of Microbiology and Biotechnology
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• v.7 no.2
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• pp.151-156
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• 1997

Two commercially available lipases, Lipase OF (non-specific lipase from Candida rugosa) and Lipolase 100T (1, 3-specific lipase from Aspergillus niger), were immobilized on insoluble hydrophobic support HDPE (high density polyethylene) by the physical adsorption method. Hydrolysis performance was enhanced by mixing a non-specific Lipase OF and a 1, 3-specific Lipolase 100T at a 2 : 1 ratio. The results also showed that the immobilized lipase maintained its activity at broader temperature ($25~55^{\circ}C$) and pH (4-8) ranges than soluble lipases. In the presence of organic solvent (isooctane), the immobilized lipase retained most of its activity in upto 12 runs of hydrolysis experiment. However, without organic solvent in the reaction mixture, the immobilized lipase maintained most of its activity even after 20 runs of hydrolysis experiment.

• Biotechnology and Bioprocess Engineering:BBE
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• v.11 no.6
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• pp.522-525
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• 2006

Biodiesel conversion from soybean oil reached a maximum of 70% at 18 h using immobilized 1,3-specific Rhizopus oryzae lipase alone. Biodiesel conversion failed to reach 20% after 30 h when immobilized nonspecific Candida rugosa lipase alone was used. To increase the biodiesel production yield, a mixture of immobilized 1,3-specific R. oryzae lipase and nonspecific C. rugosa lipase was used. Using this mixture a conversion of greater than 99% at 21 h was attained. When the stability of the immobilized lipases mixture was tested, biodiesel conversion was maintained at over 80% of its original conversion after 10 cycles.

• BMB Reports
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• v.36 no.4
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• pp.417-420
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• 2003

The present work describes a colorimetric microplate assay for lipase activity based on the reaction between 5,5'-dithiobis(2-nitro benzoic acid) (DTNB) and the hydrolysis product of 2,3-dimercapto-1-propanol tributyrate (DMPTB). Reaction mixtures containing DTNB, DMPTB, and lipase were prepared in microplate wells, and the absorbance at 405nm was recorded after incubation at $37^{\circ}C$ for 30 min. A linear relationship was obtained in the range of 0.1-1 U of lipase activity by this method. The reaction conditions were also optimized for the range of 0.01-0.1 U or 1-10 U. When assaying crude tissue extracts, the reaction of DTNB with non-specific reducing agents created a major source of error. However, this error was corrected by the use of blank samples that did not contain DMPTB.

• Korean Journal of Microbiology
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• v.51 no.3
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• pp.187-193
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• 2015

Candida (also known as Pseudozyma) antarctica lipase B (CAL-B) has been intensely studied in academic and industrial fields. However, the research related to its homologous enzymes has been rarely reported. In the current investigation, protein sequence similarity search of CAL-B has been conducted and six homologous protein sequences were identified. After the syntheses of their codon-optimized genes, the synthetic genes have been cloned into a periplasmic expression vector to express in Escherichia coli. Among six homologous sequences, four sequences were successfully expressed in E. coli. The hydrolytic activities of the expressed proteins towards 4-nitrophenyl acetate and 4-nitrophenyl butyrate were measured and compared with those of CAL-B to identify whether the expressed proteins work as a hydrolase. It has been revealed that the expressed proteins can hydrolyze the substrates and the specific activities were determined as $(1.3-30){\times}10^{-2}{\mu}mol/min/mg$, which are lower than those of CAL-B. Among these homologous enzymes, Pseudozyma hubeiensis SY62 exhibits the comparable enantioselectivity to that of CAL-B towards the hydrolysis of (${\pm}$)-1-phenylethyl acetate.

• KSBB Journal
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• v.7 no.3
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• pp.172-178
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• 1992

A simple and effective method has been developed for the immobilization of lipase producing Rhizopus chinensis on polyurethane foam. In this method, the fungal cells with 1, 3 specific lipase in there inside are immobilized within the foam matrix. Four types of commercially available polyurethane foam were tested. The ultimate purpose of the process is to produce low-cost biocatalysts for lipase-catalyzed reactions, which are being increasingly used for industrial applications. Effects of several parameters were studied on the cell loading and the hydrolytic activity of intracellular lipase after acetone drying. These parameters were the type, size, and amount of polyurethane foam. In all the cases, the intracellular lipase activity obtained with the foam was approximately twice greater than that obtained in the absence of the foam.

• Journal of Microbiology and Biotechnology
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• v.13 no.3
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• pp.451-456
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• 2003

An expression vector system was developed for the secretory production of recombinant Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae. The mature L1 lipase gene was fused to ${\alpha}-amylase$ signal sequence from Aspergillus oryzae for the effective secretion into the culture broth and the expression was controlled under GAL10 (the gene coding UDP-galactose epimerase of S. cerevisiae) promoter. S. cerevisiae harboring the resulting plasmid successfully secreted L1 lipase into the culture broth. To examine an optimum condition for L1 lipase expression in the fed-batch culture, L1 lipase expression was induced at three different growth phases (early, mid, and late-exponential growth phases). Maximum product on of L1 lipase (1,254,000 U/l, corresponding to 0.65/1) was found when the culture was induced at an early growth phase. Secreted recombinant L1 lipase was purified only through CM-Sepharose chromatography, and the purified enzyme showed 1,963 U/mg of specific activity and thermoalkalophilic properties similar to those reported for the enzyme expressed in Escherichia coli.

• Microbiology and Biotechnology Letters
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• v.46 no.1
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• pp.51-58
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• 2018

The expression, purification, and characterization of cold-adapted lipase from the psychrophile, Janthinobacterium sp. were investigated. The gene encoding lipase from Janthinobacterium sp. PAMC 25641 was cloned into a pET28a(+) vector and heterologously expressed in Escherichia coli BL21 (DE3). The amino acid sequence deduced from the nucleotide sequence (930 bp) corresponded to a protein having 309 amino acid residues with a molecular weight of 32.7 kDa and a pI of 5.55. Recombinant E. coli harboring the Janthinobacterium lipase gene were induced by addition of isopropyl-${\beta}$-D-thiogalactopyranoside. $Ni^{2+}$-NTA affinity chromatography was used to purify the lipase, which had a specific activity of 107.9 U/mg protein. The effect of temperature and pH on the activity of lipase was measured using p-nitrophenyl octanoate as a substrate. The stability of the lipase at low temperatures indicated it is a cold-adapted enzyme. The lipase activity was increased by $Na^{2+}$, $Mg^{2+}$, and $Mn^{2+}$, and decreased by $Zn^{2+}$ and $Co^{2+}$. Analysis of the lipase activity using various p-nitrophenyl esters showed a strong preference toward short acyl chains of the esters, indicating the ability of the cold-adapted lipase to hydrolyze short-chain esters.

• Microbiology and Biotechnology Letters
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• v.23 no.6
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• pp.695-701
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• 1995

An alkaline lipase producing bacteria was isolated from soil and identified as Serratia liquefaciens AL-11. from the results of analysis of its morphological, biochemical and physiological properties. This strain showed the highest productivity of alkaline lipase when grown at pH 9.0 and 30$\circ$C for 42 hours in the medium of 1% peptone, 0.5% tryptone, 0.9% yeast extract, 1% starch, 1% tween 80, 0.05% CaCl$_{2}$ and 0.05% NaCl. The enzyme was purified by ammonium sulfate treatment, Sephadex G-100 gel filtration and DEAE-Sephadex A-50 column chromatography. The specific activity of the purified enzyme was 27 unit/mg protein and the yield of enzyme activity was 61.3%. The homogeneity of the purified enzyme was verified by polyacrylamide gel disc electrophoresis. Molecular weight of the purified enzyme was estimated about 53,000 by sodium dodecyl sulfate- polyacrylamide gel electrophoresis. This enzyme is composed of 17 amino acids of which glycine, proline and glutamic acid were three miajor acids.

• Journal of the Korean Society of Food Science and Nutrition
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• v.32 no.8
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• pp.1200-1205
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• 2003

Structured lipids (SL) were synthesized by transesterification of corn oil and conjugated linoleic acid (CLA) in the continuous type reactor using sn-1,3 specific Rhizomucor miehei lipase. The parameters of reaction were observed in terms of flow rate, temperature, and substrate molar ratios. The highest incorporation of CLA was obtained with 1 mL/min flow rate, 55$^{\circ}C$ and 1 : 3 (corn oil/CLA) molar ratio, showing 10.26 ㏖%. When different reaction temperatures and substrate ratios were studied, the highest incorporation was obtained at $65^{\circ}C$ (17.33 ㏖%) and 1 : 5 (corn oil/CLA) ratio (17.50 ㏖%), respectively. After pancreatic lipase analysis, most of all CLA were found at sn-1,3 position. The iodine values of obtained SLs ranged from 110 to 120. From the neutral lipid analysis by normal-phase HPLC, produced SLs composed of 99.35 ∼ 99.89% triacylglycerols, 0.11 ∼ 0.51% 1,2- and 1,3-diacylglycerols, and 0.06 ∼ 0.22% monoacylglycerols.

• BMB Reports
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• v.30 no.1
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• pp.7-12
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• 1997

The kinetics of the interesterification of triolein and stearic acid catalyzed by immobilized Rhizopus delemar lipase were studied in a batch operation. In order to clarify the mechanisms of this reaction, three models are discussed under various conditions in terms of the ratio of triolein and stearic acid. The rate constants involved in the proposed model were determined by combining the numerical Gauss-elemination method, and the trial-and-error method so as to fit the calculated results with the experimental data. The accuracy of the obtained rate constants was confirmed after they were substituted for simultaneous differential equations and the equations simulated using an adaptive step-size Runge-Kutta method. Finally, the model which agrees with the calculated results and the experimental data was selected.