Keggin-type polyoxotungstates as mushroom tyrosinase inhibitors - A speciation study

Mushroom tyrosinase abPPO4 is a commercially relevant polyphenol oxidase and has been being targeted for numerous inhibition studies including polyoxometalates (POMs). In the present work, its diphenolase activity was inhibited at pH 6.8 by a series of structurally related polyoxotungstates (POTs) of the alpha-Keggin archetype, exhibiting the general formula [Xn+W12O40]((8-n)-) in order to elucidate charge-dependent activity correlations. Kinetic data were obtained from the dopachrome assay and W-183 NMR was applied to obtain crucial insights into the actual Keggin POT speciation in solution, facilitating a straightforward assignment of inhibition effects to the identified POT species. While [PW12O40](3-) was completely hydrolyzed to its moderately active lacunary form H-x[PW11O39]((7-x)-) (K-i = 25.6 mM), [SiW12O40](4-) showed the most pronounced inhibition effects with a K-i of 4.7 mM despite of partial hydrolysis to its ineffective lacunary form H-x[SiW11O39]((8-x)-). More negative Keggin cluster charges of 5- and 6- generally resulted in preclusion of inhibitory efficacy as well as hydrolysis, but with the Ni-substituted cluster [PW11O39{Ni(H2O)}](5-) enzymatic inhibition was clearly restored (K-i = 9.7 mM). The inhibitory capacity of the structurally intact Keggin POTs was found to be inversely correlated to their net charge. The here applied speciation strategy is of utmost importance for any biological POM application to identify the actually active POM species.