Equine estrogens concentrations in aquatic environment and their endocrine disrupting potentials for fish

Limited data are currently available on the occurrences and toxicological implications of equine estrogens in aquatic ecosystems. In the present study, we determined concentrations of equine estrogens, including equilin, equilenin, and their 17alpha- and 17beta-dihydro analogues, in the river water collected from Hokkaido, Japan. Among six equine estrogens measured, equilin (2.72 +- 2.22 ng/L) concentrations were the highest in the river sample, followed by 17alpha-dihydroequilin (2.17 +- 2.20 ng/L). This study was the first to detect the equine estrogens in the river water collected from Hokkaido, Japan. We further investigated transcriptional profiles of estrogen-responsive genes, such as vitellogenins (Vtg1 and Vtg2), choriogenins (ChgL and ChgH) and estrogen receptor subtypes (ERalpha, ERbeta1, and ERbeta2), in the liver of male medaka (Oryzias latipes) exposed to six equine estrogens (1-300 ng/L) for 3 days. Our quantitative RT-PCR analyses revealed that the expression levels of hepatic Vtg, Chg and ERalpha genes in male medaka responded to various types and concentrations of equine estrogens. The estrogenic potencies of the chemicals were in the order of equilin > 17beta-estradiol > equilenin > 17beta-dihydroequilin > 17beta-dihydroequilenin > 17alpha-dihydroequilin > 17alpha-dihydroequilenin, suggesting the higher lowest-observed effective concentration (LOEC) of 17beta-estradiol (LOEC : 30 ng/L) than that of equilin (10 ng/L). Furthermore, we investigated the reproductive and transgenerational effects of equilin in paired medaka exposed to 10, 100, and 1000 ng/L for 21 days. The short-term reproduction assay demonstrated that equilin (100 and/or 1000 ng/L) adversely affected the reproduction (fecundity and/or fertility) of adult medaka. In Fsub(1) generation fertilized eggs, the hatching of embryos in the 100 and 1000 ng/L treatment groups also showed adverse effects, suggesting the endocrine-disrupting potential of equilin.