Modulation of the lipophilicity and molecular size of thiosemicarbazone inhibitors to regulate tyrosinase activity

Six cresolic thiosemicarbazone derivatives bearing different tertiary amino substituents attached at 3-position of the cresolic ring through a methylene joint were prepared and evaluated as mushroom tyrosinase (TYR) inhibitors. A close correlation between their inhibition potency and molecular lipophilicity and size was observed. The hydroxyethyl substitution was beneficial to improve inhibitory competency and acted as a lipophilicity regulating group of the inhibitors. The inhibitors are worthy of evaluation as potential TYR activity regulators used for different purposes. [Display omitted] A group of 5-methylsalicylaldehyde thiosemicarbazone derivatives (HMTs) bearing different lipophilic and steric substituents attached at the 3-position of cresol ring were synthesized and investigated as mushroom tyrosinase (TYR) inhibitors. The ability of HMTs to inhibit the diphenolase activity of TYR was evaluated with L-DOPA as substrate by determining IC50 values in relation to their structure modifications. HMTs displayed distinct inhibitory competencies towards TYR activity with IC50 values in the range of 1.02–143.56 μM. A close correlation between their inhibition potency and both lipophilicity and molecular size was observed. The inhibitory effect of the hydroxyethyl-containing derivatives was much higher than the hydroxyethyl-free ones overall. Among them, HMT-NBO exhibited the most potent effect with IC50 of 5.85 μM, which was nearly 25-fold and 3.8-fold lower than its parent HMT-NBE and the control kojic acid, respectively. The hydroxyethyl clearly benefited the improvement of the inhibitory competences and acted as a regulating group of lipophilicity of the inhibitors. The kinetic analyses showed that HMTs were reversible and mixed type inhibitors against mushroom TYR. The inhibition mechanism was studied by means of fluorescence spectroscopy, FT-IR, ESI-MS and molecular docking analysis. The results indicated that the observed inhibitory effect of HMTs was accomplished by acting on the amino acid residues rather than by chelating the centre copper ions of TYR. Each of HMTs can insert the hydrophobic pocket and interact with the residues of TYR through Van der Waals forces and hydrogen bonds, with additional electrostatic interactions for HMT-NEE and HMT-NEO further strengthening the affinity. Meanwhile, the inhibitors were observed to bind with L-DOPA or/and L-DOPAquinone forming 1:1 stoichiometric complexes, probably exerting indirect inhibition against