Evaluation of p-phenylene-bis and phenyl dithiocarbamate sodium salts as inhibitors of mushroom tyrosinase.

Abstract

Two structurally related compounds, phenyl dithiocarbamate sodium salt (I) and p-phenylene-bis (dithiocarbamate) sodium salt (II) were prepared by reaction of the parent aniline and p-phenylenediamine with CS₂ in the presence of sodium hydroxide. These water soluble compounds were characterized by spectroscopic techniques, IR, ¹H NMR and elemental analysis. The inhibitory effects of both compounds on both activities of mushroom tyrosinase (MT) from Agricus bisporus were studied at two temperatures, 27°C and 37°C. L-3, 4-dihydroxyphenylalanine (L-DOPA), and l-tyrosine were used as natural substrates for the catecholase and cresolase enzyme reactions, respectively. Kinetic analysis confirmed noncompetitive inhibition mode of I and mixed type of II on both activities of MT; I and II inhibit MT with inhibition constants (K(I)) of 300 µM and 4 µM, respectively. Analysis of thermodynamic parameters indicated predominant involvement of hydrophobic interactions in binding of I and electrostatic ones in binding of II to MT. It seems that II is a more potent MT inhibitor due to its two charged head groups able to chelate copper ions in the enzyme active site. Intrinsic fluorescence studies as a function of concentrations of both compounds showed unexpectedly quenching of emission intensity without any shift of emission maximum. Extrinsic ANS-fluorescence indicated that only binding of I induces limited changes in the tertiary structure of MT, in agreement with the postulated hydrophobic nature of the binding mechanism.