Estrogenic activity of tissue conditioners in vitro

Objectives. In order to assess the estrogenic activities of plasticizers used in tissue conditioners and four commercial tissue conditioners, we carried out in vitro tests. Methods. Seven plasticizers and two metabolites were diluted to concentrations ranging from 10−9 to 10−4 M. Four commercial tissue conditioners were also diluted to concentrations ranging from 2×10−8 to 2×10−3 g/ml. Estrogenic activities were tested by the E-screen test using MCF-7 cells. When estrogen is present, the cells proliferate. Instead of counting the cells or nuclei directly, cell numbers were assessed by measurement of the total protein content using the sulforhodamine B assay. The liquid compositions of the four commercial tissue conditioners were examined by high-performance liquid chromatography. Results. n-Butyl benzyl phthalate, dibutyl phthalate, n-butyl phthalyl n-butyl glycolate, di-2-ethylhexyl phthalate and benzyl salicylate significantly increased proliferation of MCF-7 cells. The remaining two plasticizers, di-2-ethylhexyl adipate and benzyl benzoate, as well as two metabolites of dibutyl phthalate and di-2-ethylhexyl phthalate, i.e. monobutyl phthalate and mono-2-ethylhexyl phthalate, respectively, did not increase proliferation of MCF-7 cells at the concentrations tested. Four commercial tissue conditioners, Coe comfort (CC), Tissue Conditioner (TC), Hydro Cast (HC) and Denture Soft (DS) II, significantly increased proliferation of MCF-7 cells. HPLC data revealed the commercial products contained plasticizers: benzyl benzoate and dibutyl phthalate in CC, dibutyl phthalate in TC, n-butyl benzyl phthalate in HC and n-butyl phthalyl n-butyl glycolate in DS II. Significance. Except for benzyl benzoate and di-2-ethylhexyl adipate, the plasticizers tested showed estrogenic activity. The four commercial tissue conditioners tested also showed estrogenic activity, and HC showed especially strong estrogenicity.