• Bulletin of the Korean Chemical Society
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• v.27 no.5
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• pp.642-648
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• 2006

Mushroom tyrosinase (MT) structural changes in the presence of $Cu ^{2+}$ and $Ni ^{2+}$ were studied separately. Far-UV CD spectra of the incubated MT with the either of the metal ions indicated reduction of the well-ordered secondary structure of the enzyme. Increasing in the maximum fluorescence emission of anilinonaphthalene-8-sulfonic acid (ANS) was also revealing partial unfolding caused by the conformational changes in the tertiary structure of MT. Thermodynamic studies on the chemical denaturation of MT by dodecyl trimethylammonium bromide (DTAB) showed decrease in the stability of MT in the presence of $Cu ^{2+}$ or $Ni ^{2+}$ using their activation concentrations. Both activities of MT were also assessed in the presence of different concentrations of these ions, separately, with various monophenols and their corresponding diphenols. Kinetic studies revealed that cresolase activity on p-coumaric acid was boosted in the presence of either of the metal ions, but inhibited when phenol, L-tyrosine, or 4-[(4-methylphenyl)azo]-phenol was substrate. Similarly, catecholase activity on caffeic acid was enhanced in the presence of $Cu ^{2+}$ or $Ni ^{2+}$, but inhibited when catechol, L-DOPA, or 4-[(4-methylbenzo)azo]-1,2-benzenediol was substrate. Results of this study suggest that both cations make MT more fragile and less active. However, the effect of the substrate structure on the MT allosteric behavior can not be ignored.

• YAKHAK HOEJI
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• v.45 no.4
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• pp.387-396
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• 2001

The effects of ascorbic acid, thiols such as cysteine, n-acetyl-ι-cyteine, glutathione, thiourea, 2-mercaptoethanol and dithiotreithol and organic acids such as magic acid, citric acid, glycolic acid, taurine and kojic acid on polyphenol oxidate (PPO) activity were studied in order to establish if it reacts with oxidized product and/or directly inhibits the enzyme. To investigate the mechanism, the quantification of t-butylcatechol and 4-methylcatechol (phenolic compounds) as substates, their oxidized product and sulphydryl colorless additional compounds were determined by high performance liquid chromatograph (HPLC) method. Chromatographic results indicate that ascorbic acid, organic acids and lower level of cysteine reduced oxidized product of substrates back to their respective positions uf ο-diphenols. On the other hand, other thiols and high level of cysteine reacted with oxidative product of ο-diphenols and then produced sulphydryl colorless compounds. Cysteine apperars to have two types of mechanism of actions in the formation of oxidative products of substrates depending on its concentration; ascorbic acid-type and other thiols-types. The effect of ascorbic acid with thiols on polyphenol oxidase was determined by same method. Chromatographic results indicate that ascorbic acid was more reactive with oxidized product of substrates than thiols.

• Journal of the Korean Society of Industry Convergence
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• v.5 no.2
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• pp.147-152
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• 2002

Because of the difference of the relative reactivity between two hydroxyl groups of the hydroquinone due to the steric hindrance of the substituent, many combinations of the substituent direction in the polyesters derived from asymmetrical diphenols such as monosubstituted hydroquinones was expected. It was studied how the mode of the direction affected the properties of the resulting polyesters in terms of the transition temperatures of the thermotropic polyesters prepared from terephthalic acid, 2,4-dichloroterephthalic acid, and phenylhydroquinone by the reaction using p-Toluenesulfonylchloride in pyridine. The direction was tried to control the relative reactivity by changing the reaction temperature and addition time of the hydroquinone, and by modifying it through an association of the hydroquinones with DMF.

• KSBB Journal
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• v.29 no.1
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• pp.1-8
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• 2014

Tyrosinases catalyze the hydroxylation of monophenolic compounds and the conversion of o-diphenols to oquinones. The enzymes are mainly involved in the modification of tyrosine into L-3,4-dihydroxyphenyl-alanine (L-DOPA) and DOPA/DOPAquinone-drived intermolecular cross-linking, which play the key roles of pigmentation to the cells. It is ubiquitously distributed in microorganisms, plants, and animals all around the nature world. They are classified as copper- containing dioxygen activating enzymes; two copper ions are coordinated with six histidine residues in their active sites and they are distinguished as met-, deoxy-, and oxy-form depending on their oxidative states. Natural extraction and recombinant protein approaches have been tried to obtain practical amounts of the enzymes for industrial application. Tyrosinases have been widely applied to industrial and biomedical usages such as detoxification of waste water containing phenolic compounds, L-DOPA as a drug of Parkinson's disease, biomaterials preparation based on the cross-linking ability and biosensors for the detection of phenolic compounds. Therefore, this review reports the mechanism of tyrosinase, biochemical and structural features and potential applications in industrial field.

• Applied Biological Chemistry
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• v.24 no.3
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• pp.167-173
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• 1981

Crude enzyme of polyphenol oxidase was obtained from garlic. This enzyme actively oxidized triphenols such as pyrogallol and gallic acid, although it showed very weak activity on diphenols such as catechol and chlorogenic acid among the phenolic compounds tested. The optimum pH of the enzyme was 6.5 which was slightly higher than that of the garlic itself (pH 6.0). The enzyme was stable relatively at heat treatment. Sodium metabisulfite inhibited the enzyme activity at the concentration of 1mM, and KCN, L-ascorbic acid and thiourea also inhibited the enzyme action. $Mg^{++}$ activated the enzyme activity. $Cu^{++}$ activated slightly the enzyme action at low concentration (1mM), but inhibited at high concentration (10mM).

• Applied Biological Chemistry
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• v.26 no.1
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• pp.41-46
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• 1983

The characteristics of crude polyphenol oxidase extracted from mushroom[Tricholoma matsutake(S. Ito et Imai) Sing.] were investigated. The enzyme showed highest affinity to pyrogaroll among trihydroxyphenols. Except for o-diphenols the enzyme was inactive in di-and monophenols. The optimum pH was about 4 and the optimum temperature ranged from 45 to $55^{\circ}C$. The enzyme activity was completely inhibited by $70^{\circ}C$ heat treatment for 2 min. $Cu^{++}$, $Fe^{++}$ and $Mg^{++}$ activated the enzyme at low concentration($10^{-1}mM$), but inhibited at high concentration (1mM). The most potent inhibitors were Na-diethyldithiocarbamate, L-ascorbic acid, L-cysteine and NaCl. The Km value with pyrogaroll was 0.88mM.

• Clean Technology
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• v.21 no.2
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• pp.102-107
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• 2015

Rhodamine dyes are widely used as fluorescent probes because of their excellent photophysical properties, such as high extinction coefficients, excellent quantum yields, great photostability, relatively long emission wavelengths. A great synthetic effort has been focused on developing efficient and practical procedures to prepare rhodamine derivatives, because for most applications the probe must be covalently linked to another (bio)molecule or surface. Sulforhodamine B is one of the most used rhodamine dyes for this purpose, because it carries two sulfoxy functions which can be easily utilized for binding with other molecules. Recently, we needed an expedient, practical synthesis of sulforhodamine derivatives. We found the existing procedure for obtaining those compounds unsatisfactory, particularly, with the cyclization process of the dihydroxytriarylmethane (1) to produce the corresponding xanthene derivative (2). We report here our findings, which represent modification of the existing literature procedure and provide access to the corresponding xanthene derivative (2) in a high yield. Use of methanol as a co-solvent was found quite effective to prohibit the water molecule produced during the cyclization reaction from retro-cyclizing back to the starting dihydroxytriarylmethane and the yield of the cyclization was increased (up to 84% from less than 20%). The reaction temperature was significantly lowered (80 vs. 135 ℃). Thus, the reaction proceeds in a higher yield and energy-saving manner where the use of reactants and the production of chemical wastes is minimized.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.2
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• pp.205-213
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• 2011

The Folin-Ciocalteu (F-C) reagent has been extensively used for quantifying total phenolic contents in many different types of food materials. Since several different procedures of the assay methods using the F-C reagent have been applied, we investigated changes in reactivity of various phenolic compounds with the F-C reagent under three different assay conditions and factors affecting reactivity. Among 10 standard compounds tested, compounds with high hydroxyl density (number of -OH/molecular weight) showed a largely different response according to addition sequence of the F-C reagent or $Na_2CO_3$. Preincubation in $Na_2CO_3$ significantly reduced the reactivity of the phenolic compounds bearing galloyl moiety (e.g. gallic acid, tannic acid, and epigallocatechin-3-gallate) with the F-C reagent, while monophenol compounds including ferulic acid and sinapinic acid were more stable as compared to diphenols. There was little change in response to the F-C reagent of all phenolic compounds incubated in acidic pH; their reactivity except ferulic acid was reduced significantly when incubated in neutral or alkaline pH. Changes in reactivity of gallic acid incubated in $Na_2CO_3$ or neutral/alkaline pH conditions were the most prominent. $H_2O_2$ generated from phenolic compounds did not affect the reaction with the F-C reagents. The present results suggest that reactivity of different phenolic compounds with F-C reagent was affected considerably by different procedures of the assay, and the total phenolic contents could be fluctuated according to standard compounds and assay scheme.

• Current Research on Agriculture and Life Sciences
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• v.9
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• pp.51-59
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• 1991

This study was conducted to understand browning characteristics of Chinese Chestnut during processing and storage. For this, the isolation and identification of polyphenolic compounds and the biochemical characteristics of polyphenol oxidase(PPO) were investigated. The content of total phenol was $6.5{\mu}g/g$ and it was consisted of ferulic acid, caffeic acid, synapic acid, p-coumaric acid, gallic acid and salicylic acid in order. PPO was purified 11.7 fold through ammonium sulfate fractionation, DEAE-cellulose column chromatography and Sephadex G-200 column chromatography. Purified enzyme showed single protein and activity band by polyacrylamide gel electrophoresis. The optimum pH and temperature of PPO were 5.9 and $45^{\circ}C$, respectively. The activity of PPO was lost 93% by exposing at $80^{\circ}C$ for 15minutes. $Mg^{{+}{+}}$, $Ca^{{+}{+}}$, $Zn^{{+}{+}}$ increased the activity of PPO, but $Fe^{{+}{+}}$, $K^+$, $Hg^{{+}{+}}$ inhibited PPO at 10mM concentration. L-ascorbic acid, thiourea, sodium chloride and L-cysteine were effective inhibitors of PPO. The activity of PPO was higher for o-diphenols than other polyphenols. The Km value of PPO for catechol was 5mM.

• Journal of the Korean Electrochemical Society
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• v.2 no.1
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• pp.31-37
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• 1999

Copper-containig enzyme, laccase (Rhus vernicifera) was immobilized onto gold electrode using self-assembly technique and its surface properties and catalytic activities were examined. Laccase is an oxidoreductase capable to oxidize diphenols or diamines by 4-electron reduction of molecular oxygen without superoxide or peroxide intermediates. The electrode surface were modified by $\beta-mercaptopropionate$ to have a net negative charge in neutral solution and positively charged laccase (pI=9) was immobilized by electrostatic interaction. The successful immobilization was confirmed by cyclic voltammograms which showed typical surface-confined shapes and behaviors. The amount of charge to reduce the surface was similar to the charge calculated assuming the surface being covered by monolayer. The activity of the immobilized enzyme was tested by the capbility of oxidizing a substrate, ABTS (2,2-azine-bis-(3-ethylbenzthioline-6-sulfonic acid) and it was maintained for $2\~3$ days at $4^{\circ}C$. The immobilzed laccase showed about $10\~15\%$ activity compared to that in solution. The laccase-modified electrode showed the activity of elefoocatalytic reduction of oxygen in the presence of mediator, $Fe(CN)_6^{3-}$ The addtion of azide which is an inhibitor of laccase compeletly eliminated the catalytic current.