Alıç (crataegus monogyna) meyvesinden polifenol oksidaz enziminin kısmi saflaştırılması ve karakterizasyonu

Abstract

Bu çalışmada, Alıç (crataegus monogyna) meyvesinden polifenol oksidaz (PFO) enzimi, soğuk aseton çöktürmesi metodu kullanılarak kısmi olarak saflaştırılmış ve karakterize edilmiştir. Enzimin optimum pH ve optimum sıcaklık değerleri sırasıyla 5.0 ve 30°C olarak belirlenmiştir. Katekol substratı için Lineweaver-Burk eğrisi ile maksimum reaksiyon hızı (V_maks) ve Michaelis–Menten sabiti (K_m) değerleri sırası ile 5507 U/mL.dak ve 12.72 mM olarak hesaplanmıştır. İnhibisyon çalışmaları polifenol oksidazın yaygın inhibitörleri olan askorbik asit, sodyum metabisülfit ve benzoik asit ile yapılmış ve IC₅₀ değerleri sırasıyla 0.012 mM, 0.099 mM, 2.21 mM olarak tespit edilmiştir. Farklı fenolik substratlarla yapılan substrat spesifikliği çalışmasında enzimin katekol substratına duyarlılığının 4-metil katekol ve L-tirosin substratlarına göre daha fazla olduğu tespit edilmiştir. 

Keywords

• crataegus monogyna,polifenol oksidaz,soğuk aseton çöktürmesi,karakterizasyon,inhibisyon

The partial purification and characterization of polyphenol oxidase from hawthorn (crataegus monogyna)

Abstract

In this study, polyphenol oxidase from crataegus monogyna fruit was partially purified using cold aceton precipitation and characterized. The optimum pH and optimum temperature of the enzyme were found to be 5.0 and 30°C, respectively, in the presence of catechol as a substrate. The maximum reaction rate (V_max) and the Michaelis-Menten constant (K_m) values for the catechol substrate were calculated to be 12.72 mM and 5507 U/mL.dak, respectively, using the Lineweaver-Burk plot. Inhibition studies were performed for ascorbic acid, sodium metabisulphite and benzoic acid, which are common inhibitors of the enzyme, and the IC₅₀ values for each inhibitor were determined to be 0.012 mM, 0.099 mM, 2.21 mM, respectively. The PPO were tested on different phenolic substrates, including 4-methylcatechol, catechol, L-tyrosine. The highest activity was observed in the presence of catechol for enzyme. 

Keywords

• crataegus monogyna,polyphenol oxidase,cold acetone precipitation,characterization,inhibition

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