Investigation of some kinetic properties of polyphenol oxidase from parsley (Petroselinum crispum, Apiaceae)

Polyphenol oxidase was extracted and purified from parsley by a procedure that included (NH4)2SO4 precipitation followed by dialysis and gel filtration chromatography. These procedures led to 26.92-fold purification with 26.46% recovery. Optimum pH, temperature, substrate concentration, and ionic strength were determined with six substrates. Some kinetic properties of the enzyme such as Vmax, KM, and kcat were calculated for the substrates. The kcat/KM values of the enzyme for catechol, 4-methyl catechol, progallol, l-dopa, dopamine, and catechin were 24,937, 2,680, 48.65, 10,000, 3.04, and 206.3 mM/min, respectively. The best substrate of the enzyme was found to be catechol. The native molecular weight of the enzyme was estimated to be 237 kDa based on its mobility in gel filtration column. The inhibitory effects of sodium azide, ascorbic acid, l-cysteine, and glutathione on the enzyme activity were tested, and IC50 values were estimated to be 23 mM, 51 mM, 62 mM, and 240 mM, and Ki constants were also calculated as 6.06 ± 3.06 mM, 7.75 ± 1.65 mM, 15.75 ± 6.23 mM, and 23.60 ± 8.25 mM, respectively by means of Lineweaver-Burk graphs. All inhibitors inhibited the enzyme noncompetitively and the most effective of them was sodium azide.

► Parsley PPO was isolated 26.92-fold purification. ► The best substrate of parsley PPO was catechol. ► The native molecular weight of parsley PPO was 237 kDa. ► Sodium azide, ascorbic acid, L-cysteine, and glutathione inhibit parsley PPO. ► The most effective of these inhibitors was sodium azide.