The objectives of this project are to develop and apply methods which can be used to identify and quantify xenobiotic substances in biological samples that act through the estrogen receptor (ER) and/or the aryl- hydrocarbon receptor (AhR) to cause adverse effects on human reproductive health. Specific Aim I will utilize cell-based bioassay systems developed in the Cellular Biology Project (Project 5) as the basis for methods to detect ER- and AhR agonists in biological samples following removal of endogenous (steroidal) estrogens which compete for ER binding. We hypothesize that endogenous ER binding ligands can be removed from serum prior to assay by immuno-precipitation. The bioassays will be tested using a unique set of paired serum and urine samples from a population of male Russian children with high levels of exposure to environmental chemicals. Specific Aim II employs the non- human primate model to validate the bioassay methods using blood and urine samples recovered following experimental in vivo exposures to TCDD, t-octylphenol, fenvalerate, atrazine and diuron1. We hypothesize that bioassays for bioactivators of the ER and AhR will be as sensitive as specific immunoassays (from the Immunochemical Biomarkers Project, Project 3) for detecting exogenous ER- and AhR agonists. Specific Aim II involves the application of previously developed assays for reproductive biomarkers and bioassays of exposure for study of a population of women who eat contaminated fish from the San Francisco Bay. Our preliminary data indicate that there are abnormalities of menstrual function in this group of women, and we hypothesize that exposure to xenobiotic ER agonists and AhR agonists is responsible. Specific Aim IV will utilize the same assays for study of a population of women with suspected exposure to chlorinated hydrocarbons who are recruited in the Epidemiology Project (Project 9). Specific Aim V will adapt existing immunoassays for reproductive hormone metabolites to a format which is more efficient for population-based studies. This aim relies on the Immunochemical Biomarkers Project (Project 3) for the development of critical reagents and the Biosensors Project (Project 8) for development of assay formats.