Fecal glucocorticoid metabolite (FGM) analysis has received considerable attention in conservation biology because it has potential to be used as a noninvasive measure of stress in animals. There has been a recent and extensive literature describing the importance of technical, physiological, and biological validations of this technique, yet surprisingly little is known about how FGM concentrations change during chronic stress. Therefore, we experimentally induced chronic stress in both captive and free-living European starlings (Sturnus vulgaris). Chronic stress was elicited using a rotation of four different 30 min acute stressors for 16 days in the laboratory and 8 days in the field. Exogenous ACTH, the primary glucocorticoid secretagog, significantly increased FGM concentrations in approximately 2 h, and our assay detected endogenous diel glucocorticoid rhythms similar to those of other birds. Thus, our assay was both physiologically and biologically validated. However, experimentally induced chronic stress did not alter daytime or nighttime FGM concentrations in captive starlings. In contrast, chronically stressed adult female starlings had higher FGM concentrations than unstressed female starlings in the field. Our field data support the general assumption that higher FGM concentrations indicate chronic stress, but our captive data do not. Overall, our results suggest that more research is need before FGM analysis can be used as a reliable measure of stress in animals, especially those kept in captivity.