• Analytical Science and Technology
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• v.36 no.1
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• pp.44-52
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• 2023

This protocol clarifies a simple and precise method for measuring the activity of xanthine oxidase (XO) enzyme inhibitor. XO enzyme, which accelerates oxidative stress-related disorders through its capacity to generate hydrogen peroxide and superoxide anion radicals (O₂^•-), has been found to be inhibited by several plant extracts. Enzyme samples were incubated with a suitable buffer containing adequate amounts of xanthine as a substrate to determine XO activity. The method depends on direct measurements of uric acid and hydrogen peroxide production to test XO with and without interference. The CUPRAC reagent (Cu(Nc)₂²⁺) was used to inhibit enzyme reaction after incubation was complete. The generated urate and peroxide reduced the Cu(II)-neocuproine complex (Cu(Nc)₂²⁺) to a brightly colored Cu(I)-neocuproine complex (Cu(Nc)₂⁺), which was assessed with a spectrophotometer at 450 nm. XO activity was found to be directly related to the increased absorbance of the colored Cu(I)-neocuproine complex (Cu(Nc)₂⁺). To eliminate catalase enzyme interference, the proposed method used sodium azide and was validated against XO activity using the UV method in matched samples with t-test analysis. The proposed assay can determine XO activity with high precision, as indicated by the correlation coefficient (R² = 0.9935) from comparison with the reference protocol.

• Applied Chemistry for Engineering
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• v.20 no.4
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• pp.375-380
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• 2009

Polyepichlorohydrin rubber was treated with sodium azide (Na$N_3$) to replace its chlorine by azide ($N_3$). Then, the azidated polyepichlorohydrin rubber (Az-PECH) was blended with thermoplastic styrene-acrylonitrile copolymer with the rubber/plastic ratio of 80/20, 70/30 and 60/40 (wt/wt). The miscibility, mechanical and dynamic mechanical properties as well as elastic recovery properties of the blends were evaluated by DMA (Dynamic Mechanical Analyzer) and tensile tests. When azidation level in azidated PECH was upto 50%, the blends exhibited excellent miscibility, manifested by a single $T_g$, and fairly good elastic recovery. When azidation level was 75%, the blends showed phase separation. The miscible Az-PECH/SAN blends exhibited typical thermoplastic elastomer like properties, ie. melt processibility and high extensibility as well as good elastic recovery rate. It was also observed from combustion test that higher energy is released with the increase in the azidation level of the Az-PECH in the blends.

• The Korea Journal of Herbology
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• v.30 no.5
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• pp.15-21
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• 2015

Objectives : Methicillin-resistantStaphylococcus aureus(MRSA) is a human pathogen. New antibacterial agents are needed to treat MRSA-related infections. This study investigated the antibacterial activity of EtOH 70% extracts ofTonghyeonipal-dan(THD) which prescription is composed of oriental medicine against MRSA.Methods : The antibacterial activity of THD was evaluated against MRSA strains by using the Disc diffusion method, broth microdilution method, Checkerboard dilution test, and Time-kill test; its mechanism of action was investigated by bacteriolysis, detergent or ATPase inhibitors were used.Results : The minimum inhibitory concentration (MIC) of THD is 1,000~2,000 μg/mL against MRSA. In the checkerboard dilution test, fractional inhibitory concentration index (FICI) of THD in combination with antibiotics indicated synergy or partial synergism againstS. aureus. Furthermore, a time-kill assay showed that the growth of the tasted bacteria was considerably inhibited after 24 h of treatment with the combination of THD with selected antibiotics. For measurement of cell membrane permeability, THD 500 μg/mL along with concentration of Triton X-100 (TX) and Tris-(hydroxymethyl) aminomethane (TRIS) were used. In the other hand, N,N-dicyclohexylcarbodimide (DCCD) and Sodium azide (NaN3) were used as an inhibitor of ATPase. TX, TRIS, DCCD and NaN3 cooperation againstS. aureusshowed synergistic action.Conclusions : Accordingly, antimicrobial activity of THD was affected by cell membrane and inhibitor of ATPase were assessed. These results suggest that THD has antibacterial activity, and that THD extract offers great potential as a natural antibiotic against MRSA.

• KSBB Journal
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• v.27 no.4
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• pp.257-262
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• 2012

The gene encoding Thermus thermophilus HJ6 laccase (Tt-laccase) was cloned, sequenced, and comprised of 1,389 nucleotides encoding a protein (462 amino acids) with a predicted molecular mass of 51,049 Da. The deduced amino acid sequence of Tt-laccase showed 99.7% and 44.3% identities to the Thermus thermophilus HB27 laccase and Synechococcus sp. RS9917 laccase, respectively. Tt-laccase gene was expressed as a fusion protein with six histidine residues in E. coli Rosetta-gami (DE3) cells, and the recombinant protein was purified to homogeneity. UV-Vis spectrum analysis revealed that the enzyme has copper atoms, a type I Cu(II) and a type III binuclear Cu(II). The optimum pH for the oxidation of guaiacol was 5.0 and the optimum temperature was $90^{\circ}C$ The half-life of heat inactivation was about 120 min at $90^{\circ}C$ The enzyme reaction was inhibited by sodium azide, L-cystein, EDTA, dithiothreitol, tropolone, and kojic acid. The enzyme oxidized various known laccase substrates, its lowest $K_m$ value being for 4-hydroxyindole, highest $k_{cat}$ value for syringaldazine, and highest $k_{cat}/K_m$ for guaiacol.

• The Korean Journal of Food And Nutrition
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• v.18 no.1
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• pp.54-62
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• 2005

The antioxidative effect, antimutagenic capacity and inhibitory effect of cancer cell proliferation in ethanol extracts of Ganoderma lucidum were studied for suggestion of prevention and dietetic treatment of chronic diseases and development of antioxidative, antimutagenic and anticancer functional food by employing biological and biochemical assay. The IC/sub 50/ of MDA with BSA conjugation reaction, lipid peroxidation and scavenging effect on DPPH radical in ethanol extracts of Ganoderma lucidum showed 1026.21 mg/mL, 0.152 mg/mL and 0.412 mg/mL, respectively. So, the most effective antioxidative capacity in ethanol extracts of Ganoderma lucidum was the lipid peroxidation, among the method used this study. The indirect and direct antimutagenic effects of ethanol extracts of Ganoderma lucidum were examined by Ames test using Salmonella typimurium TA98 and TA100. The inhibition rates on indirect mutagenicity mediated by 2-Anthramine showed 100% in the Salmonella typimurium TA98 and 98.74% in the Salmonella typimurium TA100 and the direct mutagenicity mediated by sodium azide in Salmonella typimurium TA100 was 82.96%. But, the inhibitory effect on indirect mutagenicity mediated by 2-Nitrofluorene in Salmonella typimurium TA98 was low(7.81%). The inhibitory Effect of Ganoderma lucidum ethanol extracts on cell proliferation in Hela and MCF-7 by MTT test were 74.36% in HeLa and 73.90% in MCF-7 at the 0.50 mg/assay concentration and IC/sub 50/ were 0.163 mg/mL and 0.196 mg/mL respectively. From this result, it is suggested that Ganoderma lucidum is believed to have a possible antioxidative, antimutagenic and anticancer capacities.

• Korean Journal of Veterinary Research
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• v.48 no.1
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• pp.17-26
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• 2008

This study was carried out to evaluate the antitumor, hepatoprotective and antimutagenic activities on hot water extract of Phellinus gilvus (PGE). Growth of tumor in mice that were orally given $0.25,0.5,1.0,2.0g\;kg^{-1}$ dose of PGE was inhibited in a dose-dependent manner (p < 0.05). The hepatoprotective effect of PGE in the carbon tetrachloride ($CCl_4$)-intoxicated rats was studied. In $CCl_4$ + PGE group, PGE was orally administered with 100 mg/kg/day dose 7 days before the treatment of $CCl_4$. The serum activity of aspartate aminotransferase and alanine aminotransferase in $CCl_4$ + PGE group were decreased at a rate of 59.6% and 54.1% compared with those in $CCl_4$ group, respectively (p < 0.05). Also, total cholesterol and triglyceride in $CCl_4$ + PGE group were significantly decreased at a rate of 90% and 73.6% compared with those in $CCl_4$ group (p < 0.05). In the Ames test, we confirmed PGE doesn't have any activity as a mutant, and PGE showed inhibitory effect against mutagenesis induced by 2-amino fluride and sodium azide in Salmonella typhimurium TA98, TA100 and TA1535 in a dose-dependent manner. From the above results, we may suggest that PGE might have useful as a material for functional food and/or animal pharmaceutics.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.16
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• pp.6575-6580
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• 2014

This study aimed to evaluate the antimutagenic and anticarcinogenic activity of turmeric essential oil as well as to establish biochemical mechanisms of action. Antimutagenicity testing was accomplished using strains and known mutagens with and without microsomal activation. Anticarcinogenic activity was assessed by topical application of 7, 12 - dimethylbenz[a]anthracene (DMBA) as initiator and 1% croton oil as promoter for the induction of skin papillomas in mice. Inhibition of p450 enzymes by TEO was studied using various resorufins and aminopyrene as substrate. Turmeric essential oil (TEO) showed significant antimutagenic activity (p<0.001) against direct acting mutagens such as sodium azide ($NaN_3$), 4-nitro-O-phenylenediamine (NPD) and N-methyl-N-nitro N'nitrosoguanine (MNNG). TEO was found to have significant antimutagenic effect (>90%) against mutagen needing metabolic activation such as 2-acetamidoflourene (2-AAF). The study also revealed that TEO significantly inhibited (p<0.001) the mutagenicity induced by tobacco extract to Salmonella TA 102 strain. DMBA and croton oil induced papilloma development in mice was found to be delayed and prevented significantly by TEO application. Moreover TEO significantly (P<0.001) inhibited isoforms of cytochrome p450 (CYP1A1, CYP1A2, CYP2B1/2, CYP2A, CYP2B and CYP3A) enzymes in vitro, which are involved in the activation of carcinogens. Results indicated that TEO is antimutagenic and anticarcinogenic and inhibition of enzymes (p450) involved in the activation of carcinogen is one of its mechanisms of action.

• Journal of Microbiology and Biotechnology
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• v.22 no.11
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• pp.1540-1548
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• 2012

Manganese peroxidase (MnP) was isolated from the culture filtrate of the wood log mushroom Stereum ostrea (S. ostrea), grown on Koroljova medium, and then purified by ammonium sulfate [70% (w/v)] fractionation, DEAE-cellulose anion exchange chromatography, and Sephadex G-100 column chromatography, with an attainment of 88.6-fold purification and the recovery of 22.8% of initial activity. According to SDS-PAGE the molecular mass of the MnP was 40 kDa. The optimal pH and temperature were found to be 4.5 and $35^{\circ}C$, respectively. The enzyme was stable even after exposure to a pH range of 4.5 to 6.0, and at temperatures of up to $35^{\circ}C$ at a pH of 4.5 for 1h. The $K_m$ and $V_{max}$ values for the substrate phenol red were found to be $8{\mu}m$ and 111.14 U/mg of protein, respectively. The MnP also oxidized other substrates such as guaiacol, DMP, and veratryl alcohol. Sodium azide, EDTA, SDS, $Cu^{2+}$, and $Fe^{2+}$, at 1-5 mM, strongly inhibited enzyme activity, whereas $Ca^{2+}$ and $Zn^{2+}$ increased enzyme activity. The participation of the purified enzyme in the decolorization of dyes suggests that S. ostrea manganese peroxidase could be effectively employed in textile industries.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.21
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• pp.9517-9522
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• 2014

Purple rice (Oryza sativa L. var. indica) cv. Kum Doisaket is cultivated in northern Thailand. This study evaluated the mutagenic and antimutagenic properties of hydrophilic and lipophilic components of purple rice using the Ames test. The seed and hull of purple rice were extracted with hexane, methanol, ethanol, and water. The methanol extracts had the highest amounts of phenolic acids and flavonoids, while the hexane extracts contained large amount of tocols and ${\gamma}$-oryzanol. None of the extracts were mutagenic in Salmonella typhimurium strains TA98 and TA100. The hexane extract of rice hull and the methanol extract of rice seed were strongly effective against aflatoxin B1- and 2-amino-3, 4 dimethylimidazo (4, 5-f) quinoline-induced mutagenesis, while aqueous extracts showed weakly antimutagenic properties. All extracts with the exception of aqueous extracts enhanced the number of revertant colonies from benzo (a) pyrene induced-mutagenesis. None of the extracts inhibited mutagenesis induced by the direct mutagens 2-(2-furyl)-3-(5-nitro-2-furyl)-acrylamide and sodium azide. The hull extracts showed more potent antimutagenicity than the seed extracts. Based on a chemical analysis, ${\gamma}$-oryzanol and ${\gamma}$-tocotrienol in the hull and cyanidin-3-glucoside and peonidin-3-glucoside in the seed are candidate antimutagens in purple rice. The antimutagenic mechanisms of purple rice might be related to either modulation of mutagen metabolizing enzymes or direct attack on electrophiles. These findings supported the use of Thai purple rice as a cancer chemopreventive agent.

• The Korean Journal of Food And Nutrition
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• v.20 no.4
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• pp.341-348
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• 2007

The antioxidative effect and antimutagenic capacity in ethanol extracts of Lentinus edodes were studied for suggestion of prevention and dietetic treatment of chronic diseases and development of antioxidative and antimutagenic functional food by employing biological and biochemical assay. The $IC_{50}$ of MDA with BSA conjugation reaction, lipid peroxidation and scavenging effect on DPPH radical in ethanol extracts of Lentinus edodes showed 74.58 mg/assay, 5.747 mg/assay and 0.939 mg/assay respectively. So, the most effective antioxidative capacity in ethanol extracts of Lentinus edodes was the scavenging effect on DPPH radical, among the method used this study. The indirect and direct antimutagenic effects of ethanol extracts of Lentinus edodes were examined by Ames test using Salmonella typimurium TA98 and TA100. The inhibition rates on indirect mutagenicity mediated by 2-anthramine showed 91.67% in the Salmonella typimurium TA98 and 96.60% in the Salmonella typimurium TA100. The inhibitory effect on direct mutagenicity mediated by sodium azide in Salmonella typimurium TA100 was 22.83%. and mediated by 2-nitrofluorene in Salmonella typimurium TA98 was 5.34%. This data indicates that ethanol extracts of Lentinus edodes have more effective effects on indirect mutagenicity than direct mutagenicity. From this result, it believed to have a possible antioxidative and antimutagenic capacities, and taken for the candidate of prevention and dietetic treatment of chronic diseases and development of antioxidative and antimutagenic functional food.