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Yinhan Gong,¹ Hong Soon Chin,¹ Lis Sa Elissa Lim,² Chong Jin Loy,² Jeffrey P. Obbard,^1,3 and E.L. Yong²

¹Tropical Marine Science Institute, ²Department of Obstetrics and Gynecology, and ³Department of Chemical and Environmental Engineering, National University of Singapore, Republic of Singapore

Abstract
Abnormal sexual differentiation and other reproductive abnormalities in marine animals indicate the presence in seawater of endocrine-disrupting compounds (EDCs) that perturb the function of the sex hormone signaling pathways. However, most studies to date have reported on EDC effects in freshwater and sewage samples, and there is a paucity of bioassay data on the effects of EDCs in marine waters. Our aims in this study were to devise robust methodologies suitable for extracting potential EDCs and to measure their summated effects on activities of androgen receptors (ARs) and estrogen receptors (ER- and ER-ß) in marine samples from Singapore's coastal waters. In this study, we examined the ability of C18, hydrophilic and lipophilic balance, and diol cartridges to extract potential EDCs from seawater samples. Extracts from C18 cartridges exhibited the highest sex hormone bioactivities in reporter gene assays based on a human cell line expressing AR, ER-, and ER-ß. Examination of extracts from 20 coastal locations showed high androgenic and estrogenic agonist activities in confined clusters closest to the main island of Singapore. Sex hormone activity declined rapidly in clusters farther from the main coastline and in more open waters. Unexpectedly, surface and mid-depth samples from the confined high-activity clusters, in the presence of hormone, exhibited AR and ER- activities that were 200-900% higher than those observed for the cognate hormone alone. This enhanced sex hormone activity suggests that analyses of complex seawater mixtures may uncover unusual bioactivities that may not be obvious by studying individual compounds. Our data present a "snapshot" of the sex hormone disruptor activity in Singapore's marine environment and indicate that C18 extraction for EDCs used in conjunction with reporter gene bioassays represents a robust and sensitive methodology for measuring summated androgenic and estrogenic activities in seawater. Key words: androgen, endocrine disruptor, estrogen, reporter gene bioassay, seawater, Singapore. Environ Health Perspect 111:1448-1453 (2003) . doi:10.1289/ehp.6139 available via http://dx.doi.org/ [Online 25 April 2003]