• The Korean Journal of Pharmacology
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• v.29 no.2
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• pp.275-282
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• 1993

Liver microsomal epoxide hydrolase (mEH) is active in the detoxification of epoxide-containing reactive intermediate. Previous studies in this laboratory have shown that thiazole and pyrazine are efficacious inducers of mEH in rats with large increases in mEH mRNA levels (Carcinogensis, Kim et al, 1993). mEH was purified to electrophoretic homogeneity from thiazole-induced rat hepatic microsomes using DEAE-cellulose column chromatography whereas another protein $({\sim}43\;kDa)$ was co-purified with mEH from pyrazine-induced rat hepatic micrsomes (200 mg/kg body weight/day, ip, 3d). The antibody raised from a rabbit against mEH protein purified from thiazole-induced rat hepatic microsomes appeared to specifically recognize mEH protein in rat hepatic microsomes, as assessed by immunoblotting analysis. Immunoblotting analyses revealed a 10- and 7-fold increase in mEH levels in the hepatic microsomes isolated from thiazole- and pyrazine-treated rats, respectively. Moreover, immunoblotting analysis showed cross-reactivity of the mEH antibody with a 43 kDa protein in pyrazine-induced rat hepatic microsomes and with co-purified 43 kDa protein in purified fractions. The ratio between the 43 kDa protein and mEH in pyrazine-induced rat microsomes or in purified fractions was ${\sim}1$ to 15. N-terminal amino acid sequence analysis of both purified rat mEH and 43 kDa protein revealed that 10 out of 12 amino acids in N-terminus of the 43 kDa protein were identical with the mEH sequence with two amino acid residues of the 43 kDa protein undetermined. Either thiazole or pyrazine treatment, however, failed to increase the levels of mEH protein in rabbits while pyrazine caused elevation of the 43 kDa protein in this species, as determined by irnrnunoblotting analysis. These results demonstrated that treatment of rats with either thiazole or pyrazine causes elevation in hepatic mEH expiession whereas pyrazine treatment results in induction of another mEH-related 43 kDa protein and that a distinct species difference exists between rats and rabbits in the induction of mEH by these xenobiotics.

• The Korean Journal of Mycology
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• v.31 no.2
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• pp.59-66
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• 2003

Laccase produced by Pycnoporus cinnabarinus SCH-3 isolated from Korea was partially purified using ultrafiltration, anion exchange chromatography and affinity chromatography, The laccase was produced as the predominant extracellular phenoloxidase during primary metabolism. Neither lignin peroxidase nor manganese-dependent peroxidase were detected in the culture fluid. In order to examine the effect of inducers in laccase production, 2,5-xylidine was added in the culture of Pycnoporus cinnabarinus SCH-3. Addition of 2,5-xylidine enhanced 25-fold laccase production. Purified laccase was a single polypeptide having a molecular mass of approximately 66 kDa, as determined by SDS-polyacrylamide gel electrophoresis, and carbohydrate content of 9%. $K_{m}\;and\;V_{max}$ values for laccase with ABTS [2,2-azinobis (3-ethylbenzthiazoline 6-sulfonic acid)] as a substrate (Lineweaver-Burk plot) was determined to be $44.4{\mu}M\;and\;56.0{\mu}mole$, respectively. The optimal pH for laccase activity was found to be 3.0. The enzyme was very stable for 1 hour at $60{\circ}C$. Half-life ($t_{1/2}$) of the enzyme was about 10 min at $80{\circ}C$. Spectroscopic analysis of purified enzyme indicated that the enzyme was typical of copper-containing protein. Substrate specificity and inhibitor studies for laccase also indicated to be a typical fungal laccase. The N-terminal amino acid sequence of the P. cinnabarinus SCH-3 laccase showed 94% of homology to the N-terminal sequences of laccases from P. cinnabarinus PB and P. coccineus.

• Korean Journal of Environmental Biology
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• v.18 no.3
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• pp.331-336
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• 2000

In order to examine the effect of SO$_2$, which is the major component of acid rain, on the peroxidase activity, rice (Oryza sativa) seedlings were grown on the media containing various concentrations of Na$_2$SO$_3$. Na$_2$SO$_3$ concentrations needed for the 50% inhibition of rice seed germination were determined to be 300$\mu\textrm{g}$/ml at pH 7, 8$\mu\textrm{g}$/ml at pH 5 and 2$\mu\textrm{g}$/ml at pH 3. Notably, about 8 fold and 4 fold increase of the specific activity of the enzyme were observed with the seedlings treated with 8$\mu\textrm{g}$/ml Na$_2$SO$_3$ at pH 5 and 2$\mu\textrm{g}$/ml Na$_2$SO$_3$ at pH 3, respectively. The effects of Cd and Pb on the peroxidase activities and chlorophyll contents were also examined. About 3.9 fold higher peroxidase activities were found at 0.03mM Cd, and the chlorophyll contents were reduced to 63% of the control seedlings. At 0.04mM Pb, 2.5 fold higher enzyme activities were found and the chlorophyll contents were reduced to 72%. Therefore, the increases of rice peroxidase activities might be involved in the defense mechanism of the cell against various environmental stresses such as Na$_2$SO$_3$, Cd and Pb. The effects of Cu and Fe, which are the inducers of oxidative stresses by the generations of reactive oxygen species, on the peroxidase activities were also investigated. About 57% and 65% activity losses were found at 0.5mM CuSO$_4$ and 0.5mM FeSO$_4$, respectively, and radical scavenger ethanol almost completely protected both inactivations. However, dimethyl sulfoxide, mannitol, thiourea and histidine showed different radical scavenging effects one another against Cu and Fe inactivation.

• The Korean Journal of Pesticide Science
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• v.19 no.1
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• pp.64-70
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• 2015

In order to control garlic white rot (Sclerotium cepivorum), which threatens garlic production in farmers fields, soil solarization (solar sterilization), sclerotia germination inducers and effective microorganisms as biological control agents, and chemical fungicides have been used. Among them, fungicide has been largely used to reduce garlic white rot. In this study, the antifungal activities of five fungicides, prochloraz(a.i. 25%, EC), tebuconazole (a.i. 25%, WP), flutolanil (a.i. 15%, EC), iminoctadine tris-albesilate (a.i. 40%, WP) and isoprothiolane (a.i. 40%, EC) with different mode of action, in mycelial growth, sclerotia germination and sclerotia production, were tested. The inhibitory effects of the 5 fungicides on the mycelial growth, and sclerotia germination and production of garlic white rot pathogen (S. cepivorum T11-2) were investigated on potato dextrose agar (PDA) and their control efficacies were evaluated on garlic flakes. There was no mycelial growth of S. cepivorum T11-2 on PDA amended with $0.8{\mu}g\;mL^{-1}$ of prochloraz or $100{\mu}g\;mL^{-1}$ of tebuconazole. Also prochloraz and tebuconazole inhibited perfectively the sclerotia germination of the pathogen at 10 and $1.0{\mu}g\;mL^{-1}$, respectively. In spite of a very low activity of isoprothiolane in mycelial growth and sclerotia germination of S. cepivorum T11-2, it showed a good inhibitory activity against sclerotia production of S. cepivorum T11-2 on PDA amended with $1.67{\mu}g\;mL^{-1}$. Prochloraz, tebuconazole and flutolanil showed above 70% of control value when they were treated at $100{\mu}g\;mL^{-1}$ using the garlic flake cutting-method.

• Journal of Life Science
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• v.26 no.2
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• pp.226-233
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• 2016

Vascular smooth muscle cell (VSMC) apoptosis has been identified in various vascular diseases, including atherosclerosis and restenosis after angioplasty, and has been known to precipitate atherosclerotic plaque instability and rupture. Oxysterols are known as inducers of apoptosis in VSMC, and 7-ketocholesterol (7KC) is the major nonenzymically formed oxysterol in atherosclerotic lesions. The precise mechanism underlying VSMC apoptosis is still poorly understood. In this study, we investigated whether 7KC causes apoptosis, and characterized its apoptotic mechanisms in primary cultured rat aortic VSMC. Cell viability was assessed by MTT assay and trypan blue assay. Apoptosis was assessed by flow cytometry, immunofluorescence, immunoprecipitation, and Western blot analyses. 7KC markedly decreased the VSMC viability in a time- and concentration-dependent manner, and increased the production of 4-hydroxynonenal (HNE), a major end-product of lipid peroxidation, which also decreased the VSMC viability. Pretreatment with 2,4-dinitrophenylhydrazine, a well-known reagent of lipid peroxidation-derived aldehydes, significantly restored the 7KC-decreased viability of VSMC. Furthermore, HNE, as well as 7KC, reduced the level of total Akt, a major mediator of cell survival. The 7KC-decreased level of total Akt was significantly restored by pretreatments with 2,4-dinitrophenylhydrazine and N-acetylcysteine. Lactacystin, a proteasome inhibitor, protected VSMC against apoptosis and Akt degradation, but did not inhibit HNE production. In the immunoprecipitation assay, 7KC increased HNE-modified Akt. From the results, it seems that, in atherosclerotic lesions, 7KC induces HNE production in VSMC, and this HNE binds to Akt, proceeding to proteasomal degradation of Akt, through which mechanism the atherosclerotic plaque instability may be facilitated.