Wells–Dawson phosphotungstates as mushroom tyrosinase inhibitors: a speciation study

In order to elucidate the active polyoxotungstate (POT) species that inhibit fungal polyphenol oxidase (AbPPO4) in sodium citrate buffer at pH 6.8, four Wells–Dawson phosphotungstates [α/β-PV2WVI18O62]6− (intact form), [α2-PV2WVI17O61]10− (monolacunary), [PV2WVI15O56]12− (trilacunary) and [H2PV2WVI12O48]12− (hexalacunary) were investigated. The speciation of the POT solutions under the dopachrome assay (50 mM Na-citrate buffer, pH 6.8; L-3,4−dihydroxyphenylalanine as a substrate) conditions were determined by 183W-NMR, 31P-NMR spectroscopy and mass spectrometry. The intact Wells–Dawson POT [α/β-PV2WVI18O62]6− shows partial (~ 69%) disintegration into the monolacunary [α2-PV2WVI17O61]10− anion with moderate activity (Ki = 9.7 mM). The monolacunary [α2-PV2WVI17O61]10− retains its structural integrity and exhibits the strongest inhibition of AbPPO4 (Ki = 6.5 mM). The trilacunary POT [PV2WVI15O56]12− rearranges to the more stable monolacunary [α2-PV2WVI17O61]10− (~ 62%) accompanied by release of free phosphates and shows the weakest inhibition (Ki = 13.6 mM). The hexalacunary anion [H2PV2WVI12O48]12− undergoes time-dependent hydrolysis resulting in a mixture of [H2PV2WVI12O48]12−, [PV8WVI48O184]40−, [PV2WVI19O69(H2O)]14− and [α2-PV2WVI17O61]10− which together leads to comparable inhibitory activity (Ki = 7.5 mM) after 48 h. For the solutions of [α/β-PV2WVI18O62]6−, [α2-PV2WVI17O61]10− and [PV2WVI15O56]12− the inhibitory activity is correlated to the degree of their rearrangement to [α2-PV2WVI17O61]10−. The rearrangement of hexalacunary [H2PV2WVI12O48]12− into at least four POTs with a negligible amount of monolacunary anion interferes with the correlation of activity to the degree of their rearrangement to [α2-PV2WVI17O61]10−. The good inhibitory effect of the Wells–Dawson [α2-PV2WVI17O61]10− anion is explained by the low charge density of its protonated forms Hx[α2-PV2WVI17O61](10−x)− (x = 3 or 4) at pH 6.8.