We examined the efficacy of noninvasive monitoring of endocrine function via fecal steroid immunoassays in the golden eagle and peregrine falcon. High-pressure liquid chromatography analyses of fecal glucocorticoid metabolites (fGCM) revealed that minor percentages of immunoreactive fGCM co-eluted with [3H]corticosterone in both sexes of the eagle (2.5–2.7%) and falcon (7.5–11.9%). In contrast, most fecal estrogen metabolites in eagle and falcon females co-eluted with radiolabeled estradiol-17β ([3H]; 57.6, 64.6%, respectively) or estrone ([3H]; 26.9, 4.1%, respectively). Most fecal progestin metabolite immunoreactivity in the female eagle (24.8%) and falcon (21.7%) co-eluted with progesterone ([14C]). Most fecal androgen metabolite immunoreactivity in eagle (55.8%) and falcon (63.7%) males co-eluted with testosterone ([14C]). Exogenous adrenocorticotropin hormone induced increased fGCM excretion above pre-treatment in both species, but only significantly (P< 0.05) in the eagle. Both species showed increased fGCM after saline administration, suggesting the detection of ‘handling stress.’ Both species exhibited enterohepatic and renal recirculation of administered steroids as demonstrated by biphasic and triphasic excretion patterns. Thus, noninvasive fecal hormone monitoring is a valid and promising tool for assessing gonadal and adrenal status in rare and threatened birds-of-prey.