Mushroom polyphenol oxidase

Polyphenol oxidase Mushroom, grape, pear, kiwi, sago, tea, potato, strawberry, olive, beet, cocoa Enzymatic browning (pig-ments),ripening of fruits, flavour formation, colour e.g. strawberries) formation... 

RODRIGUEZ LOPEZ J, FENOLL L G, TUDELA J, DEVECE C, SANCHEZ HERNANDEZ D, DE LOS REYES E and GARCIA CANOVAS F (1999) Thermal inactivation of mushroom polyphenol oxidase employing 2450 MHz microwave radiation , J Agric Food Chem, 47 (8) 3028-35. 

Y) and avidin (from egg white) were purchased from Sigma. Polyphenol oxidase (PPO) (EC 1.14.18.1, from mushroom, 4800 U/mg), was purchased from Sigma. Biotin-labeled polyphenol oxidase (PPO-B) has to be synthesized following the procedure [1] ... 

Enzymes as different as yeast alcohol oxidase, mushroom polyphenol oxidase, and horse-liver alcohol dehydrogenase demonstrated vastly increased enzymatic activity in several different solvents upon an increase in the water content, which always remained below the solubility limit (Zaks, 1988b). While much less water was required for maximal activity in hydrophobic than in hydrophilic solvents, relative saturation seems to be most relevant to determining the level of catalytic activity. Correspondingly, miscibility of a solvent with water is not a decisive criterion upon transition from a monophasic to a biphasic solvent system, no significant change in activity level was observed (Narayan, 1993). Therefore, the level of water essential for activity depends more on the solvent than on the enzyme. 

A single enzyme is sometimes capable of many various oxidations. In the presence of NADH (reduced nicotinamide adenine dinucleotide), cyclohexanone oxygenase from Acinetobacter NCIB9871 converts aldehydes into acids, formates of alcohols, and alcohols ketones into esters (Baeyer-Villiger reaction), phenylboronic acids into phenols sulfides into optically active sulfoxides and selenides into selenoxides [1034], Horse liver alcohol dehydrogenase oxidizes primary alcohols to acids (esters) [1035] and secondary alcohols to ketones [1036]. Horseradish peroxidase accomplishes the dehydrogenative coupling [1037] and oxidation of phenols to quinones [1038]. Mushroom polyphenol oxidase hydroxylates phenols and oxidizes them to quinones [1039]. 

Another route to hydroxylated phenols is the application of mushroom polyphenol oxidase in chloroform. The primary products of oxidation. 

A portable disposable bioprobe for detection and semiquantitative determination of phenols consists of a mushroom polyphenol oxidase immobilized on a nylon membrane, acting in the presence of 3-methyl-2-benzothiazolinone hydrazone. Maroon to orange colored dyes of (138) are developed, as illustrated for phenol (equation 6), of intensity proportional to the concentration of the substrate, down to 0.05 mgL. Enzyme activity remained unscathed in the pH range 4 to 10, in the presence of various concentrations of salt and metal ions and at temperatures from 5 to 25... 

Generally, the regioselective formation of o-quinones has been known to be accomplished by using polyphenol oxidase in chloroform and not in water, because of rapid inactivation of the enzyme in water. However, catechol underwent mushroom tyrosinase-catalyzed oxidation in phosphate buffer (pH 6.8) containing 4-hydroxycoumarin (322) to afford 321 in 96% yield, as shown in Scheme 69. 

Oxidation of a number of p-substituted phenols to the corresponding o-benzoquinones was first performed by Kazandjian and Klibanov, using mushroom polyphenol oxidase and a quantitative conversion was achieved in CHCI3 as a solvent. Other hydrophobic solvents such as methylene chloride, carbon tetrachloride, benzene, toluene, hexane and butyl acetate can be used, whereas the enzyme is inactive in more hydrophilic solvents such as ether, acetone, ethyl acetate, acetonitrile and other solvents. In addition, an immobilized enzyme on glass powder or beads is more efficient than a free enzyme. 

Polyphenol Oxidase Activity and Enzymatic Browning in Mushrooms... 

Glucose oxidase Polyphenol oxidase from mushroom Organoclay natural Cameroonian smectites grafted with either aminopropyl or trimethylpropylammonium groups. Modified electrodes [40]... 

Further validation of the mechanism proposed for the catecholase activity of the dicopper complexes [Cu2(L66)]" , [Cu2(L55)], and [Cu2(EBA)]" " (Scheme 17) has been obtained investigating the inhibitory effect of kojic acid [5-hydroxy-2-(hydroxymethyl)-y-pyrone] (154). This fungal metabolite is one of the most efficient inhibitors of mushroom tyrosinase and other polyphenol oxidases (160,161). When the catalytic oxidation of DTBCH2 was studied in the presence of kojic acid, strong competitive inhibition was observed in the steps exhibiting substrate concentration dependence,... 

Devece, C. et al. Enzyme inactivation analysis for indushial blanching appheations Comparison of microwave, conventional, and combination heat treatments on mushroom polyphenol oxidase activity, J. Agric. Food Chem., 47, 4506, 1999. 

Ballestra, P. et al. Effect of high-pressure treatment on polyphenol oxidase activity of the Agaricus bispo-rus mushroom. High Press. Res., 22, 677, 2002. 

Kermasha, S., Goetghebeur, M., and Monfette, A. Studies on inhibition of mushroom polyphenol oxidase using chlorogenic acid as substrate, J. Agric. Food Chem., 41, 526, 1993. 

Polyphenol Oxidase. Peroxidase, and Lipoxygenase in Resistance. Polyphenol oxidase in conjunction with chlorogenic acid as a substrate has the potential to reduce the ability of larval H. zea and S. exigua to utilize dietary protein. For example, alkylation of casein in artificial diet (at 1.0% wwt) by PPO (from mushroom or tomato plant) and chlorogenic acid, at levels commensurate with that found in tomato foliage, inhibits the growth of both larval species by up to 70% (Table 1). Rutin is a very poor substrate for mushroom tyrosinase and tomato PPO (79 unpubl data), and hence,... 

Golan-Goldhirsh and colleagues (71,72) report that sodium bisulfite, reduced glutathione, and ascorbic acid caused a 50% inactivation of mushroom polyphenol oxidase after 28, 106, and 130 min, respectively, at 5 mM concentration. Dithiothreitol was a more effective inhibitor, causing 50% inactivation at 0.1 mM after only 70 min. 

These authors suggest that (a) the observed effect of reductants on mushroom polyphenol oxidase is dependent on whether a polarographic or spectroscopic method is used to assay the enzyme (b) the spectrophotometric method is useful to determine the optimum reductant concentration required to inhibit browning and (c) the polarographic method is useful to establish whether the inactivation of the enzyme is irreversible. 

Oxidations now known to be catalyzed by copper-containing enzymes were noticed over a century ago, when Schoenbein observed that oxidation of natural substrates resulted in pigment formation in mushrooms. Individual enzymes were gradually identified laccase by Yoshida in 1883 and tyrosinase by Bertrand in 1896. However, it was not imtil potato polyphenol oxidase was isolated in 1937 by Kubowitz that the role of copper was defined. The family of copper oxidases includes a number of enzymes of both plant and animal origin that may very probably be found to react through similar mechanisms, but which exhibit a number of individual characteristics. The enzymes to be described in this section include potato phenol oxidase, mushroom polyphenol oxidase (tyrosinase), laccase, mammalian and insect tyrosinase, and ascorbic acid oxidase. Each of these differs in certain respects from the others, and undoubtedly other related enzymes will be described from other sources that resemble these, but also display individualities. In these cases, identities in nomenclature must not be extended to imply identities in enzyme structure or activity. 

Laccase. A polyphenol oxidase has been purified from the sap of the lac tree by Keilin and Mann. Laccase differs from the potato and mushroom enzyme in several respects. With regard to substrate specificity, it oxidizes p-phenylenediamine more rapidly than catechol. p-Phenylene-diamine is not a substrate for the other polyphenol oxidases described. Laccase apparently is inert with p-cresol. It is not inhibited by carbon monoxide. Unlike the other phenol oxidases, this enzyme is not a pale yellow, but is blue, as is ascorbic acid oxidase (see below). This enzyme, however, is not an ascorbic acid oxidase. 

Polyphenol oxidases and ascorbic acid oxidase, which occur in food, are known to have a Cu /Cu redox system as a prosthetic group. Polyphenol oxidases play an important role in the quality of food of plant origin because they cause the enzymatic browning for example in potatoes, apples and mushrooms. Tyrosinases, catecholases, phenolases or cresolases are enzymes that react with oxygen and a large range of mono and diphenols. 

Polyphenol oxidases (cf. 0.3.3) in mushrooms and potatoes require pressures of 800-900 MPa for inactivation. The addition of glutathione (5 mmol/1) increases the pressure sensitivity of the mushroom enzyme. In this case, the inactivation is obviously supported by the reduction of disulfide bonds. 

Merely reducing the temperature reduces the reaction rate, but the colour changes are quite rapid even at 0 °C. This means that sensitive products that have not been pre-treated should be frozen as quickly as possible, for example mushrooms and sliced peaches. Rapid intensive browning occurs during defrosting, when the activity of polyphenol oxidases increases due to disruption of cellular structures by ice crystals. 

This work, some aspects of which are outlined below, has served to show that melanins are in fact highly irregular polymers in which several different types of monomer unit are linked in a variety of ways. From the results of an extensive series of experiments in which D L-dopa deuteriated separately at the 1, 2, 2, 5 and 6 positions and D L-l-[ C]-dopa were converted into dopa-melanin autoxid-atively or by enzymic oxidation in the presence of mushroom polyphenol oxidase. Swan and his collaborators have conclude that the principal structural fragments in the polymer are probably (0 uncyclised units of dopa (68, 0.1), (ii) indoline carboxylic acid units (69, 0.1), ( /) indole units (70, 0.65) and (iv) pyrrole carboxylic acid units (71, 0.15). The indoline (69) and indole (70) units are presumed to exist in the polymer half in the phenolic and half in the quinonoid forms. It was also suggested that the pyrrole carboxylic unit (71) is probably derived by oxidative fission, during the synthesis of the dopa-melanin, of a benzenoid unit such as (70). 

Kermasha, S Goetghebeur, M Monfette, A. Studies on Inhibition of Mushroom Polyphenol Oxidase using Ghlorogenlc Acid as Substrate. Journal of Agricultural Food Chemistry, 1993 41, 526-531. 

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