In vitro screening assay for detecting aromatase activity using rat ovarian microsomes and estrone ELISA.

摘要

Aromatase, a key steroidogenic enzyme that catalyses the conversion of androgens to estrogens, present a target for endocrine disrupting chemicals. However, little is known about the effect of pollutants on aromatase enzymes. In this study, we first optimized a non-radioisotope aromatase assay using rat ovarian microsomes in vitro and measuring the estrone level with an enzyme-linked immunosorbent assay (EIA method). The sensitivity of the EIA method was about ten times as high as that of the radioisotope (RI) method. A significant positive correlation was indicated between EIA and RI method. We used this EIA assay system to investigate the effects of aromatase activity on 45 chemicals that had previously been reported to act as endocrine disruptors or to have the possibility of having such an effect. Six of the chemicals, rose bengal, erythrosine, phloxine, allura red, gallic acid, and tributyltin, inhibited aromatase activity. The inhibitory effect of rose bengal was the strongest (IC50=2.4 x 10(-8) M), and its strength of inhibition was about 100 times that of a known aromatase inhibitor, 4-hydroxy-androstenedione (IC50=2.6 x 10(-6) M) but was about 1/25 that of fadrazole (IC50=1.0 x 10(-9) M). It is thought that this EIA method would be useful for measuring the aromatase activity of microstructures.