Synthesis And Inhibitory Effect Of Aromatic Heterocycle Thiosemicarbazones On Tyrosinase

Tyrosinase(EC 1.14.18.1), is a kind of oxidoreductase with dinuclear copper ions, widely exists in plants, animals and microorganisms, which is the key enzyme in the synthesis of melanin. Heterocyclic compounds are widely exist in natural and drug molecules, such as chlorophyll, hemoglobin, morphine, berberine, camptothecin, vitamins, antibiotics etc, many heterocyclic compounds have important physiological activity. It was reported that furfural could inhibit the activity of tyrosinase, but the inhibition mechanism of its analogues and chemical modifications on tyrosinase, is rarely reported in the previous literature.Three structurally similar aromatic heterocyclic compounds 2-thiophenecarboxaldehyde(a, d, e), 2-furaldehyde(b), 2-pyrrolecarboxaldehyde(c) were chosen and a series of their thiosemicarbazone derivatives(1a-3a, 1b-3b, 1c-3c, 1d and 1e) were synthesized. Enzyme kinetics showed that thiosemicarbazone derivatives have been found exhibiting more remarkable inhibition than their indexcompounds on mushroom tyrosinase;compound 1a-1c have the better inhibition activity than kojic acid;Among then,the compound 1a(IC50 = 0.43 μM)has the strongest inhibitory ability and about 42 times stronger than that of kojic acid(IC50 = 17.94); The hydrogen atom of N-terminal on thiourea groups replaced by methyl or phenyl, the inhibitory activity of compounds on tyrosinase was significantly reduced;The compounds with sulfur hetero atoms in the heterocyclic ring have the strongest inhibitory ability, followed by compounds with oxygen hetero atoms in the heterocyclic ring, compounds with nitrogen hetero atoms in the heterocyclic ring.Using enzyme kinetics method, ultraviolet spectroscopy, fluorescence spectroscopy, nuclear magnetic titration method to research the inhibitory mechanisms, and reveal the relationship between structure and activity, the results showed that the compound 1a-1c were the reversible and mixed-type inhibitors on tyrosinase;Copper ion chelating experiments show that compound 1a-1c and copper ions formed 1.5:1 complex; In fluorescence quenching experiments, compound 1a-1c could quench tyrosinase intrinsic fluorescence and results were consistent with inhibitory activity of tyrosinase., compound 1a behaved stronger fluorescence quenching on tyrosinase towards 1a-Cu2+ complex. Nuclear titration show that compounds could combine with copper ions in the enzyme and form hydrogen between inhibitor and enzyme. ANS fluorescence probe revealed compounds has not change the process of inhibition enzyme hydrophobic environment.This paper not only introduced a series of new, efficient, high-quality tyrosinase inhibitors, but also summed up the underlying structure-activity relationship, providing the theoretical basis for further search of novel, efficient and safe tyrosinase inhibitors. Research on inhibition mechanism and structure-activity relationship of inhibitors, provide a more scientific theory for the application of the compounds in whitening cosmetics.