Timing of gametogensis and thus spawning can be inferred through changes in plasma concentrations of gonadal hormones. In preparation for ovulation and spawning, mean concentrations of 17ß-estradiol in a population of Rio Grande chub (Gila pandora) occupying the Rio Bonito, New Mexico, peaked at 37.6 ng/mL on 16 June and declined to 1.50 ng/mL by 11 August. Similarly, the gonadal somatic index (GSI) increased from 9.02 on 21 May (n = 9) to 11.85 on 16 June (n = 2) and declined to 6.10 on 11 August (n = 2). Peak concentrations of 17ß-estradiol and elevated GSI in June coincided with peak daylength for the year (14 h and 12 min) and average water temperature of 15.1°C. Concentrations of 17ß-estradiol remained low through 3 October indicating no additional spawning events in the Rio Grande chub population. We demonstrated 17ß-estradiol is a nondestructive and thus useful tool in estimating timing of spawning in a wild fish population.