Investigation of extracellular tyrosinase isolated from Ralstonia solanacearum and screening of tyrosinase inhibitors from some plant extracts

The crude extracellular tyrosinase enzyme of Ralstonia solanacearum is partially purified by fraction precipitation with ammonium sulfate and combined with Amicon membranes to concentrate and remove salts and molecules <10 kDa.At the fraction of 80% saturation, the tyrosinase enzyme of R. solanacearum exhibited the strongest activity with both oxidative activities of monophenolase and diphenolase in the reaction with 2 substrates L-tyrosine and L-3,4-dihydroxyphenylalanine (L-DOPA).Compared with the tyrosinase of the fungus Agaricus bisporus, the activity and V max of R. solanacearum reached 89.5 and 86.7%, respectively.The IC 50 values of kojic acid with tyrosinase of A. bisporus in reaction with L-tyrosine and L-DOPA were 0.0139 and 0.0128 mM respectively and of R. solanacearum were 0.0181 and 0.0191 mM respectively.Zerumbone from Zingiber zerumbet and flavonoid fractions from Artocarpus heterophyllus leaves, Styphnolobium japonicum, and Cirsium japonicum inhibited the activity of both enzymes.The flavonoid fraction from leaves of A. heterophyllus showed higher enzyme inhibition activity than that of S. japonicum and C. japonicum.Compared to kojic acid, the flavonoid fraction of leaves of A. heterophyllus showed an inhibitory effect of 81.76% at a concentration of 0.1 mg/ml (p<0.05).The initial results show that the extracellular tyrosinase of R. solanacearum can be applied to screen for natural compounds inhibiting tyrosinase enzymes.