L-DOPA assay of polyphenol oxidase (PPO) for varietal identification in rice (Oryza sativa L.)

Abstract

A phenol colour response test on the grain is used to identify varieties of wheat and rice. It is a harmful, damaging procedure due to the carcinogenic properties of phenol. The activity of polyphenol oxidase (PPO) provides the basis for the phenol colour reaction. The goal of this research was to assess a method similar to the phenol colour reaction, by determining where the polyphenol oxidase activity threshold lies for classifying rice types into different groups. The new PPO assay was applied to commercially available rice varieties with the intention of classifying those that ranged from extremely high to extremely low PPO levels into manageable categories. Substrates such as phenol, L-tyrosine, catechol and 3,4-dihydroxyphenylalanine (L-DOPA) were tested. Twenty cultivars cultivated under identical conditions were put through a screening using a standard assay [1.5 mL of 10 mM L-DOPA in 50 mM 3-(N-morpholino) propane sulphonic acid (MOPS) buffer, pH 6.5, with 3 to 5 seeds constantly rotated in a 2-mL microcentrifuge tube for 0.5 or 1 hour at room temperature]. At 30 minutes, PPO levels were assessed. Based on the PPO values, the varieties belonging to phenol colour groups such as black, dark brown, brown, light brown or no colour matched perfectly. Considering easiness in estimation and other advantages of PPO activity, L-DOPA is proposed as a means of identifying rice varieties instead of the phenol colour reaction test.