Biochemical properties of partially purified polyphenol oxidase and phenolic compounds ofPrunus spinosaL. subsp.dasyphyllaas measured by HPLC-UV

Abstract

Polyphenol oxidase (PPO) enzyme was extracted from Prunus spinosa L. subsp. dasyphylla plum, partially purified by acetone precipitation, and its biochemical properties were investigated. Different substrates (p-coumaric acid, L-tyrosine, p-hydroxyphenyl propionic acid (PHPPA), catechol, 4methylcatechol (4-MTC), hydrocaffeic acid, gallic acid, quercetin, catechin, and epicatechin) were analysed to determine their affinities with Prunus spinosa PPO (PsPPO). The substrate specificity was in the following order: 4-MTC > catechol > hydrocaffeic acid > catechin > epicatechin. 4-MTC was the most suitable substrate (K m = 0.97 mM and V max = 4753 U/mg protein). The optimal pH values were 7.0 for 4-MTC and catechol, 5.0 for catechin and epicatechin, and 4.0-6.0 for hydrocaffeic acid. Optimal temperatures were 40°C for 4-MTC, 30°C for catechol, and 60°C for catechin, epicatechin, and hydrocaffeic acid. In the inhibition tests, the most potent inhibitor was found to be sodium metabisulphite (IC 50 = 0.01 mM), followed by ascorbic acid, thiourea, benzoic acid, L-cysteine, and sodium azide. Approximately 80 and 75% of the diphenolase activity was conserved at pH 5.0 and 7.0, respectively, at 4°C after 7 d incubation. Moreover, native polyacrylamide gel electrophoresis (Native-PAGE) of the enriched extract revealed the presence of at least six bands with PPO activity, suggesting the existence of different PPO isoforms. However, the oxidation of diphenol related to browning significantly, so the data obtained in this research provided a basis for the prevention of enzymatic browning of Prunus spinosa during processing. High-performance liquid chromatography (HPLC) revealed the plum extract contained protocatechuic acid, p-OH benzoic acid, vanillic acid, syringic acid, epicatechin, p-coumaric acid, and luteolin.

Exploring the Promotive Effects and Mechanisms of Different Polyphenolic Extracts from Prinsepia utilis Royle Seed Shell on Tyrosinase

“…It can be seen from Figure 5 A and Table 2 that Km showed a decreasing trend and Vm showed an increasing trend as the concentration of L-DOPA increased, indicating that the binding affinity force and the catalytic rate of TE to tyrosinase increased, and the latter may be affected by the former. For (+)-catechin or (−)-epicatechin, as shown in Figure 5 B,C and Table 2 , the Km value decreased with decreasing L-DOPA concentration, indicating that (+)-catechin or (−)-epicatechin increased the binding affinity to tyrosinase as reported in the previous study [ 51 ], which may be because all (+)-catechin, (−)-epicatechin and L-DOPA can act as substrates for tyrosinase [ 13 ], and (+)-catechin or (−)-epicatechin compete with L-DOPA for the substrate binding site of tyrosinase. Meanwhile, (+)-catechin or (−)-epicatechin showed an increasing trend in the catalytic rate, consistent with the above result of TE, which further indicated that the binding affinity affects the catalytic rate of tyrosinase.…”

Inhibitiory effect of garlic extracts on polyphenoloxidase

“…IC50 values were 0.011, 0.013, 0.087 mM for 4-methyl catechol, pyrogallol, catechol, respectively. In literature, it was found that ascorbic acid was the most effective inhibitory for Prunus spinosa PPO when using 4-methyl catechol as a substrate, and its IC50 value was 0.04 mM [17]. It was determined that glutathione and L-cysteine show less inhibitory effect for PPO, than the ascorbic acid.…”

Unveiling the Dual Nature of Heavy Metals: Stressors and Promoters of Phenolic Compound Biosynthesis in Basilicum polystachyon (L.) Moench In Vitro

“…A 20 µL sample was injected, and the flow rate was set to 1 mL/min. The detection wavelength was 280 nm [ 59 ].…”