The first genistin absorption screening into vacuoles of Trifolium pratense L.

Abstract

The determination of a transport mechanism for genistin (genistein-7-O-glucoside) across the tonoplast was performed on vacuoles from a cell culture of Trifolium pratense L. Genistin levels were examined in vacuoles as well as in an assay medium by HPLC (high-performance liquid chromatography) after treatment with various substances. MgATP increased the uptake of added genistin by 25%, but the nucleotide-free samples also contained this glycoside. Applying bafilomycin A1, an H⁺-ATPase inhibitor, indistinctly inhibited genistin absorption. However, vacuolar absorption of genistin was significantly reduced by N,N´-dicyclohexylcarbodiimide. This inhibitor can suppress both H⁺-ATPase and H⁺-PPase; the effect of pyrophosphate alone was not investigated. An increase in genistein levels, as result of genistin hydrolysis, could also affect the transport mechanism. The results investigated with inhibitors suggest the possible involvement of proton pumps; however additional research is required to confirm the participation of multidrug and toxin extrusion (MATE) proteins in genistin transport.