Previous studies have demonstrated that ovulatory goldfish synthesize and release a variety of steroids into the water, where some of them function as sex pheromones. Among the steroids which have been measured are free androstenedione, testosterone, 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta-P), 17 alpha,20 beta,21-trihydroxy-4-pregnen-3-one (17,20 beta,21-P), 17 alpha-hydroxy-4-pregnene-3,20-dione (17-P), 11-deoxycortisol (17,21-P), and 17 alpha,20 alpha-dihydroxy-4-pregnen-3-one (17,20 alpha-P), glucuronidated testosterone, 17,20 beta-P, and 17,20 beta,21-P, and sulfated 17,21-P, 17,20 beta-P, and 17,20 beta,21-P. This study reports the changes in the amounts of these steroids released into the water, at 3-hr intervals, by 12 female goldfish injected with human chorionic gonadotrophin (HCG). Eight fish ovulated. The amounts of steroids released into the water were significantly higher in the fish which ovulated than in those which did not. The peak rate of release (ng per hour) of free androstenedione (91), glucuronidated testosterone (143), and sulfated 17,21-P (33) occurred at ca. 4 hr postinjection; free 17-P (163) and free 17,21-P (217) at ca. 6 hr.; free 17,20 beta-P (58) and free 17,20 beta,21-P (58) at ca. 7.5 hr; and glucuronidated 17,20 beta-P (96), glucuronidated 17,20 beta,21-P (264), sulfated 17,20 beta-P (64) and sulfated 17,20 beta,21-P (153) at ca. 9 hr. Amounts of all of the steroids were at their lowest between 12 and 15 hr and, in the ovulated fish, had risen significantly again between 18 and 21 hr. The amounts of glucuronidated testosterone, 17,20 beta-P and 17,20 beta,21-P, and of sulfated 17,20 beta-P and 17,20 beta,21-P, in water exceeded those of free steroid. Elevated levels of free and glucuronidated steroids were also found in the blood plasma of HCG-injected fish. Considerably more androstenedione than testosterone was released into the water. Assessment of the "pheromonal effectiveness" of each steroid indicated that the free and sulfated forms of 17,20 beta-P and 17,20 beta,21-P, and androstenedione were the most important pheromonal steroids released by female goldfish.