Among the several xenobiotics that human activities have produced in the last decades, endocrine disruptors (EDs), including hormones, have been drawing the attention of researches due to the effects they can cause in animals. Those effects may result in hermaphrodite characteristics in fishes and amphibians, testicular growth inhibition, spermatogenesis inhibition, eggs fertilization capacity decrease, and changes in the reproductive behavior of living beings. Concentrations of only 10 ng L-1 of hormones in the aquatic medium are capable of causing endocrine effects in organisms. In this study, the natural hormone 17β-estradiol and the synthetic ones levonorgestrel and 17α-ethinylestradiol were determined in the waters of the São José do Rio Preto (SP) dam. The first step of this study was the validation of the methods according to ANVISA's Resolution 899. The detection, quantification, and lower quantification limits of the method for determining 17α-ethinylestradiol were, respectively, 25, 100, and 100 ng L-1. The linearity, relative standard deviation, accuracy, and average recovery of the method for determining 17α-ethinylestradiol were, respectively, R equal to 0.98, 3.23%, 100.53%, and 89.95%. The detection, quantification, and lower quantification limits of the method for determining 17βestradiol were, respectively, 100, 150, and 150 ng L-1. The linearity, relative standard deviation, accuracy, and average recovery of the method for determining 17β-estradiol were, respectively, R equal to 0.99, 3.43%, 106.16%, and 89.05%. For levonorgestrel, the detection, quantification, and lower quantification limits of the method were, respectively, 50, 150, and 150 ng L-1. The linearity, relative standard deviation, accuracy, and average recovery of the method for determining levonorgestrel were, respectively, R equal to 0.98, 3.48%, 105.15%, and 86.45%. In the second step of this research, samples collected in the São José do Rio Preto (SP) dam were analyzed regarding the presence of hormones. For extracting the hormones, SPE cartridges were used followed by HPLC/FLU/DAD. The results indicated the absence of the studied hormones down to the detection limits of the methods employed. Vitellogenin (VTG) analyses were performed in the blood plasma of fishes captured in the beforementioned dam, of the species Geophagus brasiliensis (pearl cichlid or pearl eartheater), Satanoperca pappaterra (Pantanal eartheater or Paraguay River eartheater), and Tilapia rendalli (redbreast tilapia). It was observed that male fishes had VTG concentrations between 152.4 and 2,841.8 ng mL-1. That indicates that there are substances with endocrine effect in the dam water, although one cannot say the studied hormones are among them.
