Estrogenic Activity of Phthalate Esters by In Vitro VTG Assay Using Primary-cultured Xenopus Hepatocytes

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Estrogenic activity of phthalate esters in dental soft resins was evaluated with an amphibian system consisting of a vitellogenin (VTG)-detecting Enzyme-Linked Immunosorbent Assay and a primary-cultured hepatocyte assay using adult male Xenopus laevis. In particular, phthalate esters-Di-n-butyl phthalate (DBP), Butyl phthalyl butyl glycolate (BPBG), Benzyl butyl phthalate (BBP), and Benzyl benzoate (BB)-were investigated. Bisphenol A (BPA) was prepared for comparison with these chemicals, and 17β-estradiol (E2) was used as a positive control. The chemicals were diluted in dimethyl sulfoxide (DMSO) to obtain final concentrations ranging from 10^<-11> to 10^<-4> mol/l. BPA induced estrogenic activity at a concentration of 1.1×10^<-6> mol/l, while E2 showed at 4.1×10^<-11> mol/l. DBF, BBP, BB, and BPBG showed no estrogenic activity at concentrations between 4×10^<-7> mol/l and 1×10^<-4> mol/l. The latter result indicated that these phthalate esters might be metabolically transformed into non-estrogenic substances in Xenopus hepatocytes. Furthermore, this study demonstrated that through in vitro metabolism assessment, the estrogenic activity of chemical substances could be directly detected in terms of VTG secretion in primary-cultured Xenopus hepatocytes.
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