• Bulletin of the Korean Chemical Society
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• 제23권7호
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• pp.985-989
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• 2002

For efficient biopulping process, very active and stable lignase is essential. Laccase is one of the best enzyme in terms of environmentally benign processes, since the enzyme uses oxygen as an oxidant to degrade lignin and produces no hamful prod ucts. We could purify a laccase homogeneously from Cerrena unicolor in a very active state. It shows characteristic absorption feature with blue band at λmax = 604 ㎚. Molecular weight of the enzyme is 57,608 which could be accurately determined by MALDI/TOF MS. The enzyme has 2.8 copper ions per enzyme implying apoenzymes might exist together. The enzyme is active in lignin degradation and the activity increases 4 times in the presence of ABTS as a mediator.

• Journal of the Korean Wood Science and Technology
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• 제33권5호통권133호
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• pp.76-85
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• 2005

The degradation of pentachlorophenol (PCP) by lignin degrading fungi was performed. Several fungi, Abortiporus biennis, Cerrena unicolor and Trametes versicolor, were tested to evaluate the inhibitory effect of PCP on their growth. At the extremal concentration of PCP $(500\;{\mu}M)$, only C. unicolor showed relatively fast growth (60% within 14 days) in the comparison to the control culture. In the case of A. biennis and C. unicolor, when initial PCP concentration was $50\;{\mu}M$, about 88.2% and 79.5% of PCP degradation were achieved within 3 days, respectively. When 2,5-xylidine (0.2 mM) was added to the C. unicolor culture, as high as 98% of PCP degradation was achieved within just an hour after its addition. A. biennis removed 44% of PCP at the same condition. PCP was completely disappeared when laccase activities reached to maximum.

• Journal of the Korean Wood Science and Technology
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• 제28권4호
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• pp.29-40
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• 2000

Highly purified Cerrena unicolor laccase (benzenediol:oxygen oxidoreductase, EC 1.10.3.2) caused the demethoxylation of milled wood lignin and several lignin related substances. The constitutive form of the enzyme produced extracellularly by C. unicolor fermenter culture was isolated and purified by ion-exchange chromatography on the DEAE-Toyopearl column and by affinity chromatography on a ConA-Sepharose and Syringyl-AH-Sepharose 4B columns. The enzyme was further immobilized on functionalized porous glass (CPG) and keratin coated CPG. The demethylating activity was monitored both by estimation of released methanol and by detection of the level of methoxyl groups (also in some water miscible solvents) after incubation of lignin materials with laccase preparations (free and immobilized). The effects of the incubation time and temperature on the demethoxylating activity of immobilized laccase preparations were also studied.

• 펄프종이기술
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• 제29권4호
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• pp.7-17
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• 1997

균체가 생산하는 laccase 효소가 다방면으로 이용되면서, 이 효소를 균체로부터 대량으로 생산하고 분리.정제하여야 할 필요성이 대두되고 있다. 아울러 이 효소의 활성을 유도하기 위한 inducer 로서 2,5-xylidine 이 주로 사용되고 있는바, 이 xylidine 의 유독성이 인정되면서 사람에게 독성을 주지 않는 환경친화적 inducer의 검색이 필요하게 되었다. 본 연구에서는 백색부후균 Cerrena unicolor가 분비하는 laccase 효소의 유도를 위한 inducer로서 ferulic acid를 사용하였으며, 균체로부터 생산 및 분리된 laccase 효소의 정제특성을 구명코자 하였다. 본효소(constitutive enzyme)로서 I 및 II를, ferulic acid를 inducer로 사용한 경우 inducing 효소 III을 분리.정제하였다. 본 효소 I 및 II의 Michaelis 정수는 각각 737 M, 716 M 이었고, inducing효소 III은 167 M 로서, 기질에 대한 높은 친화성을 보여주고 있다. 분자량도 각각 65 kD, 63 kD 였으며, inducing효소 III은 59 kD 였다. 두효소 모두 15-19%의 당 및 단백질분자당 4M의 동(Cu)을 함유하고 있었다. 정제효율은 효소 I 및 II가 10.1%, 9.4% 였으며, III은 3.2% 였다. 모든 효소의 최적 pH 는 5.5였으며, 최적온도는 비교적 높은 $40^{\circ}C$였다.

• Journal of the Korean Wood Science and Technology
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• 제32권6호
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• pp.14-22
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• 2004

The effect of cadmium ions on ligninolytic and decolourizing activities in cultures of two white-rot fungi, Cerrena unicolor and Trametes versicolor, were examined. Cadmium was added to the shallow stationary cultures growing on a liquid mineral medium. Both examined strains sorbed Cd ions in the first 24 hr of incubation. An appreciable stimulation of the activity of extracellular laccase (LAC) and inhibition of the extracellular manganese-dependent peroxidase (MnP) were simultaneously observed when 25 mgL^-1 and 50 mgL^-1 of cadmium ions were added to the cultures. On the other hand, the addition of cadmium ions also resulted in stimulating the decolorization activity of C. unicolor to decolorize Remazol Brilliant Blue R (RBBR) in the cultures, but decreasing it in the culture of T. versicolor, which is compared to the inhibition of MnP activity in this fungus. Our data indicate that the presence of Cd(II) ions can affect the ligninolytic activity of white-rot fungi. It was found that C. unicolor is a strain resistant to the presence of Cd ions in the liquid culture media, and has a potential to use this strain for bioremediation of sites contaminated with both heavy metals and aromatic pollutants.

• Journal of the Korean Wood Science and Technology
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• 제29권3호
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• pp.112-122
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• 2001

Laccase enzymes from Cerrena unicolor and Trametes versicolor were immobilized on the activated glass beads (CPG), silica gel (SG) and soil (SL). The heterogeneous matrices were activated by ${\gamma}$-aminopropyltriethoxysilane (APTES) and glutaraldehyde (GA), and their surfaces were coated by keratin (KER) on activated or non-activated CPG, SG and SL. The laccase activities were tested in the aqueous solution for the native and immobilized preparations using different pH and temperature conditions. By keratin coating on supports, in the cases of CPG-KER and SL-KER, the immobilization yield was increased from about 80% to 90%. Moreover, much less protein was immobilized in keratin coated matrices than in inorganic ones alone (e.g. on CPG-KER 57.6%, whereas on CPG alone 80.6%). Laccase immobilization on keratin coated inorganic matrices was generally more effective than that of non-coated matrices. Concerned to pH dependency, the optima pH for immobilized laccases generally shifted towards to higher values, 5.5-5.8 and even 5.9 in the case of keratin for C. unicolor and from 5.3 to 5.7 for T. versicolor, respectively, and decreased less gradually both in acidic and alkaline regions. The immobilized laccase was more stable against thermal denaturation. This seems particularly true at $75^{\circ}C$ in the case of C. unicolor, where the activity of immobilized enzyme is > 50% higher than that of the free enzyme. For T. versicolor the respective values were $65^{\circ}C$, and 50%.

• Journal of the Korean Wood Science and Technology
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• 제34권4호
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• pp.46-53
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• 2006

본 연구는 Remazol Brilliant Blue R (RBBR) 염료의 탈색능을 이용, 리그닌분해능이 강한 백색부후균을 선발하고, 이들 균주들의 리그닌분해효소의 활성에 미치는 inducer, 2,5-xylidine 첨가효과를 검토하였다. 20종의 공시균주 가운데 9종이 RBBR의 탈색, glucose-1 oxidase (GOD), laccase (LAC) 및 Mn-peroxidase (MnP) 등 4가지의 활성을 모두 가지고 있었으며, 비교적 활성이 높은 5종의 균주는 Phlebia radiata, Trametes versicolor, Abortiporus biennis, Gleophyllum odoratum 및 Cerrena unicolor 등으로 확인되었다. 이들 균주 가운데 LAC 활성이 매우 높은 T. versicolor, G. odoratum 및 C. unicolor에 대한 최적 생육온도, 최적 pH 및 LAC 활성에 미치는 inducer 첨가효과를 조사한 결과, T. versicolor가 $28^{\circ}C$에서, G. odoratum 및 C. unicolor는 $25^{\circ}C$에서 가장 좋은 생장을 보였으며, 모든 균주의 배양 최적 pH는 5.5로 나타났다. 공시균주 모두 2,5-xylidine 첨가에 의하여 LAC 활성의 증가를 보였는데, T. versicolor는 $22,700nkat/{\ell}$로 대조구에 대하여 11.3배, G. odoratum은 최대 활성이 $15,400nkat/{\ell}$로 9배, C. unicolor는 $17,330nkat/{\ell}$로 5.5배의 활성증가를 나타냈다.

• 한국펄프종이공학회:학술대회논문집
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• 한국펄프종이공학회 2010년도 춘계학술발표회 논문집
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• pp.79-92
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• 2010

In the presence of laccase, generation of monomeric aromatic acids from nonphenolic lignin model dimer veratrylglycerol-$\beta$-vanillate ether (VVE) was observed. The addition of acetovanillone (AV) or acetosyringone (AS) intensified this process, i.e. transformation was more extensive than in the experiments omitting mediators. Among the products isovanillic (IA) and vanillic (VA) acids were identified.

• 펄프종이기술
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• 제38권5호
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• pp.1-8
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• 2006

This study was performed to evaluate extensive electrochemical characteristics of 23 commercially available mediators for laccase. Electrochemical properties, interactions with laccases, and ability to degrade lignin were compared for selected mediators. Among them, NNDS has very similar electrochemical properties in terms of reversibility and redox potential (about 470 mV vs. Ag/AgCl at pH=7) compared to ABTS which is a well-known mediator. Specific activity of purified laccase from Cerrena unicolor was determined by both 2,2'-azino-bis-(3-ethylbenz-thiazoline-6-sulfonic acid) (ABTS) and 1-nitroso-2-naphthol -3,6-disulfonic acid (NNDS). The specific activity of the laccase was 23.2 units/mg with ABTS and 21.2 units/mg with NNDS. The electron exchange rate for NNDS with laccase was very similar to that for ABTS, which meant that NNDS had similar mediating capability to ABTS. Determining methanol concentration after reacting with laccase compared to lignin degradation capabilities of both ARTS and NNDS. ARTS or NNDS alone cannot degrade lignin, but in the presence of laccase enhanced the rate of lignin degradation. ABTS showed better activity in the beginning, and the reaction rate of NNDS with lignin was about a half of that of ABTS at 10 minute, but the final concentration of methanol produced in 1 hour was very similar each other. The reason for similar methanol concentration for both ABTS and NNDS can be interpreted as the initial activity of ABTS was better than that of NNDS, but ABTS would be inhibited laccase activity more during the incubation.

• Journal of the Korean Wood Science and Technology
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• 제25권3호
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• pp.29-36
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• 1997

목재부후균으로 부터 락케이스 효소의 생산 및 유도를 위하여 여러가지 유도약품(inducer)을 사용하였다. 이들 가운데 ferulic acid, pentachlorophenol 및 2,5-xylidine이 매우 높은 락케이스 활성을 나타나게 하였으며, 거의 동일한 유도효과를 보여주었다. 이들 약품 이외에도 sinapic acid, syringic acid 및 coffeic acid 등도 높은 락케이스 활성을 주었는데, 산의 형태가 알데히드류보다도 높은 유도효과를 나타냈다. 그리고 실험한 48개 균주 가운데 38개 균주가 락케이스를 생산하였으며, 이 가운데 32균주가 ferulic acid에 의해 강한 효소유도 활성을 보였다. 이러한 결과는 지금까지 락케이스 효소의 검출이 어려웠던 Abortiporus biennis 및 Gleophyllum odoratum에서도 높은 락케이스 효소의 유도를 가능하게 하였다. 아울러 가장 높은 효소활성을 나타낸 균주로서는 Cerrena unicolor 였으며, 그 락케이스 효소활성이 무처리 및 inducer 첨가시 각각 40,000 및 60,000 nkat/l 정도였다.