• Food Science and Preservation
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• v.12 no.5
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• pp.489-495
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• 2005

Polyphenol oxidase (PPO) from Burdock (Arctium lappa L.) was purified and characterized. Purification of polyphenol oxidase was achieved by ammonium sulfate precipitation, Phenyl-sepharose CL-4B hydrophobic chromatography and Sephadex G-100 gel filtration chromatography. The molecular mass of the purified PPO was estimated to be 30 kDa by SDS polyacrylamide gel electrophoresis. In a substrate specificity, maximum activity was achieved with chlorogenic acid, followed by catechol and catechin. Whereas, there was low activity with hydroquinic acid, resorcinol or tyrosine. The optimum pH and temperature for enzyme activity were 7.0 and 35$\circC$ with catechol, respectively. The enzyme was most stable at pH 7.0 while unstable at acidic and alkaline pH. The enzyme was stable when heated to 40$\circC$. But heating at 50$\circC$ for more than 30 min caused 50% loss of activity. Ascorbic acid, L-cystein and $Cu^{2+}$ inhibited the activity of pholyphenol oxidase.

• Korean journal of food and cookery science
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• v.10 no.4
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• pp.339-345
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• 1994

Polyphenol oxidase in Yam was partially purified through ammonium sulfate fractionation, DEAE-cellulose column chromatography. The specific activity of purified PPO was 138.22 unit/mg protein. The optimum pH and temperature of purified PPO were respectively 7.0 and 30$^{\circ}C$. The heat treatment at 80$^{\circ}C$ for 6 min, decreased PPO activity to 50%. The enzyme showed high substrate specificity toward catechol. The Km value for catechol was 5 mM. In the Ames test using S. typhimurium TA98 and TA100 catechol-YEBRP, pyrogallol-YEBRP, chlorogenic-YEBRP showed strong antimutagenicity on sodium azide and MECF excepting hydroquinone-YEBRP showed killing effect on both strains.

• YAKHAK HOEJI
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• v.60 no.2
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• pp.78-84
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• 2016

N-Phenylthiourea derivatives and catechol oxidase receptor complex was studied using molecular mechanics method. The starting structure was adopted from the protein databank and the calculation of energy minimization and molecular dynamics was performed with AMBER package. The molecular dynamics showed that the simulation time span of 20 ns was long enough to observe the interaction profile and stationary ligand-receptor configuration in the complex. The conformation of the ligand was related to the interaction to the receptor and the efficacy was also interpreted in this context.

• Applied Biological Chemistry
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• v.34 no.1
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• pp.49-53
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• 1991

Three polyphenol oxidase(polyphenol oxidase I, II and III ) were isolated from the crude extract of a Spuriopimpinella bracycarpa by $(NH_4)_2SO_4$ precipitation and subsequent Sephadex G-150 chromatography. The final preparation thus obtained showed three peaks of enzyme activity. Optimum pH and temperature for the activity of polyphenol oxidase were 7.5 and $30^{\circ}C$, respectively. The enzyme was completely inactivated when i4 was treated at$70^{\circ}C$ for 30min and at $80^{\circ}C$ for 5min at pH 6.5. The enzyme was partially inactivated by ascorbic acid, glutathione and potassium cyanide (0.1mM), and was completely inhibited by L-cysteine, ascorbic acid, glutathione and potassium cyanide(0.5 and 1.0mM). The enzyme has good activity on catechol and 3,4-dihydroxytoluene but was strongly inactivated on pyrogallol, dopamine and DL-dopa. The Michaelis cons4ant of the enzyme was 86.5mM with catechol as a substrate.

• Applied Biological Chemistry
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• v.31 no.4
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• pp.331-338
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• 1988

Acetone powder was prepared from raw arrowroots and the polyphenol oxidases of crude enzyme prepared from acetone powder were identified 5 isoenzymes by staining with catechol containing 0.05% phenylene diamine. The crude enzyme was passed through the columns of ion exchangers and gel permeation to fractionate the polyphenol oxidases. The main fraction of polyphenol oxidase appeared to be purified by 94-fold, with the activity yield of 45.4%, and its molecular weight was determined as 38,500 by poly acrylamide gel electrophoresis. The optimal pH and temperature for the enzyme activity were pH 7.5 and $50^{\circ}C$, respectively. The purified enzyme showed a high affinity for catechol and pyrogallol. The Michaelis constant for catechol was calculated to be 16.67mM according to the Lineweaver-Burk method. The enzyme activity was strongly inhibited by L-ascorbic acid, sodium bisulfite, EDTA and KCN, and totally inhibited, by $Fe^{3+}$ at a concentration of 1mM. However the enzyme was activated by $Zn^{2+}$ approximately 1.7 times at the same concentration.

• Preventive Nutrition and Food Science
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• v.1 no.1
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• pp.111-116
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• 1996

Polyphenol oxidase(PPO) isolated from the crude extract of ascidian, Halocynthia roretzi, showed higher affinity for catechol than tyrosine or DL-DOPA. Successful enzyme assay could be performed at $25^{\circ}C$, 10min. by mixing 0.2ml of crude enzyme extract with 2.8ml of 0.13M catechol in 0.1M sodium phosphate buffer(pH 6.4). The specific activity of PPO which had been purified with a combination of ammonium sulfate treatment, ion exchange chromatography on DEAE-cellulose, and gel filtration on Sepharose 6B was 13-fold disc gel electrophoresis. The activity of PPO was stable from pH 5.0 to 8.0 and showed the peak activity at pH 6.4 .The optimum reaction temperature for PPO oxidation on catechol was 35$^{\circ}C$ and those enzyme were heat stable up to 4$0^{\circ}C$. Molecular weigth of the enzyme was estimated about 170kDa. One molecule was found to be composed of gour subunits. Two of them had molecular weigh of 55kDa and the others 30kDa. The {TEX}$K_{m}${/TEX} values, {TEX}$V_{max}${/TEX} and catalytic efficiency({TEX}$V_{max}${/TEX}/{TEX}$K_{m}${/TEX}) for catechol were 0.12mM, 2.5mM/liter/min. and {TEX}$0.18min^{-1}${/TEX} respectively. The substrate affinity and electrophorectic pattern suggested that the enzyme of ascidian was considered to be not tyosine but catechol oxidase.

• Korean Journal of Food Science and Technology
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• v.16 no.4
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• pp.413-417
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• 1984

To examine enzymatic browning of apple wine, changes of active bands of polyphenol oxidase (EC 1.10.3.1) as well as polyphenol substances related to browning of apple wine were investigated during wine brewing. The decrease of total phenol was remarkably inhibited by the addition of sodium metabisulfite. In the meantime, auto-oxidation of catechol in a model system increased proportionally as the reaction pH and temperature increased. Catechol oxidation, however, was not detected at $4^{\circ}C$ below pH 5.0. Polyacrylamidegel electrophoretic patterns showed that the apple (Jonathan) indicated 4 bands with polyphenol oxidase activity, designated a, b, c and d whose Rm were 0.21, 0.30, 0.41 and 0.51, respectively. Among these, 2 bands, a and c remained until 5th day fermentation and only c band after 6th day fermentation. After pasteurization of apple wine at $60^{\circ}C$ for 30min, c band also remained.

• Applied Biological Chemistry
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• v.35 no.3
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• pp.170-172
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• 1992

Four types of polyphenol oxidase were isolated from the crude extract of a Ligularia fischeri by gel filtration on Sephadex G-150. Optimum pH and temperature for the activity of partially purified enzyme were 7.5 and $25^{\circ}C$, respectively. It was stable at temperature $40^{\circ}C$ when examined at pH 7.5 for 5 min, and lost 90% of its activity at $70^{\circ}C$ in 30 min at pH 7.5. The enzyme has good activity on catechol and chlorogenic acid but was inactive on dopamine.

• BMB Reports
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• v.29 no.2
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• pp.142-145
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• 1996

To investigate the cause of increased plasma catecholamine levels in liver disease, catechol-O-methyltransferase (COMT), which provides a major route of catabolism for circulating catecholamines, was studied under the cholestasis induced by mechanical biliary obstruction in rats. Monoamine oxidase (MAO) activity and the $K_m$ and $V_{max}$ values for both enzymes were also measured. Cytosolic, microsomal, and mitochondrial COMT activities in the cholestatic liver were significantly decreased throughout the experiment. Microsomal, and mitochondrial MAO activity in the cholestatic liver were also significantly decreased. Vmax values of COMT and MAO were lower. Serum COMT and MAO activities were detected after CBD ligation. These results indicate that plasma catecholamine levels are increased in liver disease due to decreased hepatic degradation of catecholamines by decreased activities of COMT and MAO. The decreased activity of these enzymes is caused by decreased biosynthesis and by flowage into the blood from the damaged hepatocyte.

• Journal of Ginseng Research
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• v.37 no.1
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• pp.117-123
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• 2013

Polyphenol oxidase (PPO) was purified from fresh ginseng roots using acetone precipitation, carboxymethyl (CM)-Sepharose chromatography, and phenyl-Sepharose chromatography. Two isoenzymes (PPO 1 and PPO 2) were separated using an ion-exchange column with CM-Sepharose. PPO 1 was purified up to 13.2-fold with a 22.6% yield. PPO 2 bound to CM-Sepharose, eluted with NaCl, and was purified up to 22.5-fold with a 17.4% yield. PPO 2 was further chromatographed on phenyl-Sepharose. The molecular weight of the purified PPO 2 from fresh ginseng was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was about 40 kDa. The optimum temperature and pH were $20^{\circ}C$ and 7.0, respectively, using catechol as a substrate. Pyrogallol showed the highest substrate specificity. The effect of a PPO inhibitor showed that its activity increased slightly in the presence of a low concentration of citric acid. High concentrations of acidic compounds and sulfite agents significantly inhibited purified ginseng PPO 2.