The estrogenic activity of phthalate esters in vitro

A large number of phthalate esters were screened for estrogenic activity using a recombinant yeast screen. A selection of these was also tested for mitogenic effect on estrogen-responsive human breast cancer cells. A small number of the commercially available phthalates tested showed extremely weak estrogenic activity. The relative potencies of these descended in the order butyl benzyl phthalate (BBP)>dibutyl phthalate (DBP)>diisobutyl phthalate (DIBP)>diethyl phthalate (DEP)>diisononyl phthalate (DINP). Potencies ranged from approximately 1 x 10(6) to 5 x 10(7) times less than 17 beta-estradiol. The phthalates that were estrogenic in the yeast screen were also mitogenic on the human breast cancer cells. Di(2-ethylhexyl) phthalate (DEHP) showed no estrogenic activity in these in vitro assays. A number of metabolites were tested, including mono-butyl phthalate, mono-benzyl phthalate, mono-ethylhexyl phthalate, mono-n-octyl phthalate; all were found to be inactive. One of the phthalates, ditridecyl phthalate (DTDP), produced inconsistent results; one sample was weakly estrogenic, whereas another, obtained from a different source, was inactive. Analysis by gel chromatography mass spectrometry showed that the preparation exhibiting estrogenic activity contained 0.5% of the ortho-isomer of bisphenol A. It is likely that the presence of this antioxidant in the phthalate standard was responsible for the generation of a dose-response curve-which was not observed with an alternative sample that had not been supplemented with o,p'-bisphenol A-in the yeast screen; hence, DTDP is probably not weakly estrogenic. The activities of simple mixtures of BBP, DBP, and 17 beta-estradiol were assessed in the yeast screen. No synergism was observed, although the activities of the mixtures were approximately additive. In summary, a small number of phthalates are weakly estrogenic in vitro. No data has yet been published on whether these are also estrogenic in vivo; this will require tests using different classes of vertebrates and different routes of exposure.