Handling, including blood collection, has often been discouraged in molting penguins because it is considered an additional stress imposed on birds already experiencing major physiologic stress associated with molting. To evaluate the degree of physiologic stress posed by molting, we compared the hematologic and plasma biochemical values and hormone levels of molting and nonmolting African penguins, Spheniscus demersus. Five male and 5 female penguins randomly chosen were given complete physical examinations, were weighed, and blood samples were taken at 7 time points before, during, and after the molt. Data were analyzed by linear mixed-model analysis of variance. Throughout the study, behavior and appetite remained normal. Catecholamine levels were highly variable within and among subjects, whereas mean corticosterone levels were significantly different between baseline, molt, and postmolt values. Significant differences from baseline values were observed in many of the hematologic analytes; however, only decreases in hematocrit and red blood cell count values were considered clinically significant. Anemia due to experimentally induced blood loss as a possible cause of the significant hematologic changes was ruled out based on results of a follow-up control study during the nonmolt season, which showed no significant changes in hematocrit level or total red blood cell counts when using similar sampling protocols, which indicates that these changes were associated with molt.