Characterization of Polyphenol Oxidase from Airen Grapes

Abstract

Polyphenol oxidase (PPO) was isolated from grapes grown in Spain and its characteristics were studied. The partially purified enzyme had both cresolase and catecholase activities. Catecholase activity had a pH optimum in a range 3.5-4.5 and was characterized by a relatively high stability to heat. The apparent KM for 4-methylcatechol was 9.5 mM. Cresolase activity presents a lag period which is modulated by different factors: enzyme concentration, substrate concentration, temperature or pH. The presence of o-diphenols in the reaction medium abolishes the lag period, these acting as co-substrates. The apparent K, towards p-cresol and the activation constant for o-diphenol for cresolase activity were 0.35 mM and 1.75 CM, respectively. 4000 x g for 15 min. The precipitate was extracted for 30 min with 1.5% Triton X-100 in 100 mM phosphate buffer pH 7.3. Two percent water-insoluble polyvinyl pyrrolidone (PVP) and calcium chloride to give a final concentration of 0.05M were added to the' medium lo precipitate phenols and pectic substances (Cash et al., 1976). The mixture was centrifuged at 15000 x g for 1 hr. An (NH&SOJ fractionation was carried out and the fraction precipitating between 45% and 95% saturation collected and rcdissolvcd in 4 mL 10 mM phosphate buffer pH 7.0. This solution, after dialysis against the same buffer, was used as enzyme source.Protein concentration of the samples at different stages of purification were determined by the method of Lowry ct al. (1951), after precipitating phenols with PVP.

Uso analítico de tecidos e de extratos brutos vegetais como fonte enzimática

“…catecol, L-dopa, dopamina, adrenalina) (Eq.7) [97][98][99][100][101][102][103][104][105][106][107] . É largamente distribuída na natureza e encontrada nos tecidos de kiwi, arroz, abacate, pêra, maçã, morango, uva, folha de espinafre, palmito, banana, batata inglesa, batata doce, pêssego, manga, berinjela, inhame entre outros [47][48][49][50][108][109][110][111][112][113][114][115][116][117][118][119][120][121][122][123][124][125][126] .…”

Diphenol Oxidases, Enzyme-catalysed Browning and Plant Disease Resistance

“…Thus the problem of catecholase browning in wines may be due, in part, to the indirect oxidative effects brought on by caftaric acid quinones (formed by catecholase catalysis) rather than to direct enzyme oxidation, since Valero et al (1988) report that, even with this endogenous anti-browning system, enzymatic browning still occurs rapidly after crushing of grapes. Methods for the control of oxidation and browning during wine processing have been comprehensively reviewed by Machiex et al (1991).…”

Efeito da aplicação foliar de silício na resistência à ferrugem e na potencialização da atividade de enzimas de defesa em cafeeiro

“…As POX estão relacionadas com o processo de proteção antioxidativa catalisando a oxidação de componentes celulares como o peróxido de hidrogênio e também no aumento da síntese de lignina que fortalece a parede celular contra a ação de enzimas líticas produzidas em abundância por alguns patógenos (Silva et al, 2008). As PFO catalisam a hidroxilação de monofenóis formando orto-difenóis e também a oxidação de ortodifenóis formando orto-quinonas, as quais exibem natureza antimicrobiana superior aos compostos fenólicos que as precedem (Valero et al, 1988). Aumento ou decréscimo na atividade de enzimas relacionadas com a resposta de defesa das plantas a patógenos detectadas durante todo o curso do processo infeccioso é um fenômeno normal diante de todas as possíveis mudanças citológicas decorrentes da íntima interação patógeno-planta.…”