• Journal of Microbiology and Biotechnology
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• v.26 no.9
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• pp.1570-1578
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• 2016

Commercial application of laccase is often hampered by insufficient enzyme stocks, with very low yields obtained from natural sources. This study aimed to improve laccase production by mutation of a Coriolopsis gallica strain and to determine the biological properties of the mutant. The high-yield laccase strain C. gallica TCK was treated with N-methyl-N-nitro-N-nitrosoguanidine and ultraviolet light. Among the mutants isolated, T906 was found to be a high-production strain of laccases. The mutant strain T906 was stabilized via dozens of passages, and the selected ones were further processed for optimization of metallic ion, inducers, and nutritional requirements, which resulted in the optimized liquid fermentation medium MF9. The incubation temperature and pH were optimized to be 30℃ and 4.5, respectively. The mutant strain T906 showed 3-times higher laccase activity than the original strain TCK under optimized conditions, and the maximum laccase production (303 U/ml) was accomplished after 13 days. The extracellular laccase isoenzyme 1 was purified and characterized from the two strains, respectively, and their cDNA sequence was determined. Of note, the laccase isoenzyme 1 transcription levels were overtly increased in T906 mycelia compared with values obtained for strain TCK. These findings provide a basis for C. gallica modification for the production of high laccase amounts.

• 한국신재생에너지학회:학술대회논문집
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• 2011.11a
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• pp.114.1-114.1
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• 2011

Bioenergy production from lignocellulosic biomass and agriculture wastes have been attracted because of its sustainable and non-edible source. Especially, canola is considered as one of the best feedstock for renewable fuel production. Oil extracted canola and its agriculture residues are reuseable for bioethanol production. However, a pretreatment step is required before enzymatic hydrolysis to disrupt recalcitrant lignocellulosic matrix. To increase the sugar conversion, more efficient pretreatment process was necessary for removal of saccharification barriers such as lignin. Alkaline pretreatment makes the lignocellulose swollen through solvation and induces more porous structure for enzyme access. In our previous work, aqueous ammonia (1~20%) was utilized for alkaline reagent to increase the crystallinity of canola residues pretreatment. In this study, significant factors for efficient soaking in aqueous ammonia pretreatment on canola residues was optimized by using the response surface method (RSM). Based on the fundamental experiments, the real values of factors at the center (0) were determined as follows; $70^{\circ}C$ of temperature, 17.5% of ammonia concentration and 18 h of reaction time in the experiment design using central composition design (CCD). A statistical model predicted that the highest removal yield of lignin was 54% at the following optimized reaction conditions: $72.68^{\circ}C$ of temperature, 18.30% of ammonia concentration and 18.30 h of reaction time. Finally, maximum theoretical yields of soaking in aqueous ammonia pretreatment were 42.23% of glucose and 22.68% of xylose.

• Journal of the Korea Organic Resources Recycling Association
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• v.16 no.1
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• pp.79-88
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• 2008

The production of bioethanol, which is one of the alternative fuel, cause the various problem such as agflation in human society. As a substitute for the feedstock, lignocellulosic biomass have a big potential. However, bioethanol production with cellulosic material is not commercialized due to high cost. Thermochemical pretreatment to improve the rate of enzyme hydrolysis and increase the recovery of fermentable sugar, is required in order to achieve the cost down in bioethanol production. In this study, various problems and technologies for pretreatment is introduced. Acid hydrolysis, alkali hydrolysis, steam explosion, organosolv process, ammonia explosion, and wet oxidation pretreatment remove lignin and hemicellulose, and reduce cellulose crystallinity. Optimization of pretreatment process on various sources of lignocellulosic biomass such as softwood, hardwood, and straw should be performed.

• Microbiology and Biotechnology Letters
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• v.40 no.3
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• pp.243-249
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• 2012

This study was conducted to screen an alginate-degrading microorganism and to investigate the characteristics of the alginate-degrading activity of its crude enzyme. A marine bacterium which produces extracellular alginate-degrading enzymes was isolated from the brown alga Sargassum thunbergii. 16S rRNA sequence analysis and physiological profiling resulted in the bacterium's identification as a Vibrio crassostreae strain, named Vibrio crassostreae PKA 1002. Its optimal culture conditions for growth were pH 9, 2% NaCl, $30^{\circ}C$ and a 24 hr incubation time. The optimal conditions for the alginate degrading ability of the crude enzyme produced by V. crassostreae PKA 1002 were pH 9, $30^{\circ}C$, a 48 hr incubation time and 8% alginic acid. The alginate degrading crude enzyme produced 3.035 g of reducing sugar per liter in 4% (w/v) alginate over 1 hr.

• Korean Journal of Microbiology
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• v.40 no.1
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• pp.54-59
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• 2004

$\beta$-Glucan has been efficiently produced with higher yield by the optimization of liquid cultivation conditions. The optimal composition of medium for batch culture was 5% (w/v) of glucose as a carbon source, 0.5% (w/v) of yeast and 0.5% (w/v) of malt extract as a nitrogen source, 0.1% (w/v) of $KH_2PO_4$ and 0.05% (w/v) $MgSO_4{\cdot}7H_2O$, which had been the base medium for determination of other conditions. The set-up conditions are pH 5.0, $28^{\circ}C$, 1 vvm for aeration and 300 rpm for agitation. In order to minimize the inhibition effect of glucose on the initial growth of mycelia and to maximize the production of extracellular $\beta$-glucan, we have reduced the initial glucose feed to 4% and added 2nd feed at the point of 70 hr from the initial feed. The 2nd feed was composed of glucose 3%, yeast extract 0.1 % and malt extract 0.1 %. It improved the $\beta$-glucan yield upto 5.2 g/L in comparison with 2.8 g/L resulted from batch cultivation. Moreover, the serial treatment of a cell wall lytic enzyme and bromelain to the mycelia was effective for extraction of the cell wall bound $\beta$-glucan. The yield of $\beta$-glucan extraction by the enzyme treatment was 3.5 g/L, which was almost 4 times higher than that by hot-water extraction.

• Food Science and Preservation
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• v.24 no.1
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• pp.60-67
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• 2017

In this study, we tried to establish the best method for fresh blueberry beverage production using enzyme treatment as well as low temperature extraction. During extraction of physiologically functional materials, we used low temperature to prevent nutritional loss by heat. In addition, we investigated optimal blueberry extraction conditions using various enzyme treatments (cellulase, pectinase, cellulase:pectinase (1:1) mixture) to increase extraction efficiency and reduce turbidity. A variety and ratio of enzymes, extraction temperature, extraction time, and shaking speed were considered for the best extraction efficiency rate. We observed high extraction efficiency rates of 85.72-86.55% and 87.06-87.93%, respectively, upon cellulase or pectinase treatment. In addition, a mixture of cellulase:pectinase (1:1) showed an extraction efficiency rate of 86.84-88.14%. The best extraction efficiency rate was observed when crude blueberry was treated at $45^{\circ}C$ (87.91%), for 3 h (87.88%), in a 90 rpm shaker (89.19%). Sugar content and acidity of blueberry extract were not affected by the various treatments. However, total phenolic compounds were detected upon pectinase treatment (18.62 mg/g). Only fructose and glucose as free sugars were found in all samples regardless of treatments and extraction conditions.

• Journal of Microbiology and Biotechnology
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• v.22 no.2
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• pp.199-206
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• 2012

Naturally immobilized tannase (tannin acyl hydrolase, E.C. 3.1.1.20) has many advantages, as it avoids the expensive and laborious operation of isolation, purification, and immobilization, plus it is highly stable in adverse pH and temperature. However, in the case of cell-associated enzymes, since the enzyme is associated with the biomass, separation of the pure biomass is necessary. However, tannic acid, a known inducer of tannase, forms insoluble complexes with media proteins, making it difficult to separate pure biomass. Therefore, this study optimizes the production of cell-associated tannase using a "protein-tannin complex" free media. An exploratory study was first conducted in shake-flasks to select the inducer, carbon source, and nitrogen sources. As a result it was found that gallic acid induces tannase synthesis, a tryptose broth gives higher biomass, and lactose supplementation is beneficial. The medium was then optimized using response surface methodology based on the full factorial central composite design in a 3 l bioreactor. A $2^3$ factorial design augmented by 7 axial points (${\alpha}$ = 1.682) and 2 replicates at the center point was implemented in 17 experiments. A mathematical model was also developed to show the effect of each medium component and their interactions on the production of cell-associated tannase. The validity of the proposed model was verified, and the optimized medium was shown to produce maximum cell-associated tannase activity of 9.65 U/l, which is 93.8% higher than the activity in the basal medium, after 12 h at pH 5.0, $30^{\circ}C$. The optimum medium consists of 38 g/l lactose, 50 g/l tryptose, and 2.8 g/l gallic acid.

• Food Engineering Progress
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• v.21 no.4
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• pp.383-390
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• 2017

Glucansucrase is an enzyme classified as a glycoside hydrolase (GH) 70 family, which catalyzes the synthesis of glucooligosaccharides with a low molecular weight using sucrose as a donor of D-glucopyranose and maltose as a carbohydrate acceptor. In this study, glucansucrase-producing lactic acid bacteria strain was isolated from the fermented foods collected in traditional markets, and the optimum conditions for the oligosaccharide production were investigated. The strain CCK940 isolated from Chinese cabbage kimchi was selected as an oligosaccharide-producing strain due to its high glucansucrase activity, with 918.2 mU/mL, and identified as Leuconostoc lactis. The optimum conditions for the production of oligosaccharides using Leu. lactis CCK940 were to adjust the initial pH to 6.0, add 5% (w/v) sucrose and 10% (w/v) maltose as a donor and acceptor molecules, respectively, and feed 5% (w/v) sucrose at 4 and 8 h of cultivation. When Leu. lactis CCK940 was cultured for 12 h at optimum conditions, at least four oligosaccharides with a polymerization degree of 2-4 were produced.

• Journal of Microbiology and Biotechnology
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• v.25 no.12
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• pp.2110-2115
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• 2015

A liquid-based colorimetric assay using a pH indicator was introduced for high-throughput monitoring of lysine decarboxylase activity. The assay is based on the color change of bromocresol purple, measured at 595 nm in liquid reaction mixture, due to an increase of pH by the production of cadaverine. Bromocresol purple was selected as the indicator because it has higher sensitivity than bromothymol blue and pheonol red within a broad range and shows good linearity within the applied pH. We applied this for simple determination of lysine decarboxylase reusability using 96-well plates, and optimization of conditions for enzyme overexpression with different concentrations of IPTG on lysine decarboxylase. This assay is expected to be applied for monitoring and quantifying the liquid-based enzyme reaction in biotransformation of decarboxylase in a high-throughput way.

• Korean Journal of Food Science and Technology
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• v.29 no.6
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• pp.1125-1130
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• 1997

The effects of enzyme combination, pH, acid washing and enzyme treatment sequence were investigated in the hydrolysis of soy protein. Comparing Alcalase vs. Neutrase/Alcalase, it appeared that Neutrase/Alcalase was more efficient than Alcalase alone, as the highest degree of hydrolysis (DH) was seen in Neutrase/Alcalase. A surprisingly high DH (more than 60%) was observed with Flavourzyme in the second hydrolysis. The separation of insolubles from the first hydrolysis had little effect on the second hydrolysis. When the washing water from the first hydrolysis was reused in the next hydrolysis, the DH and protein recovery were increased. The addition of calcium ion showed not so much positive effects by the stabilization of Neutrase on the Protein hydrolysis. The use of carbohydrase and repeated acid washing gave positive effects on DH. The simultaneous treatment using endoprotease and exoprotease with pH adjustment improved DH significantly.