The in vitro metabolism of ciclesonide, a novel inhaled nonhalogenated glucocorticoid for the treatment of asthma, was compared in cryopreserved hepatocytes from mice, rats, rabbits, dogs, and humans. Incubations of C-ciclesonide with individual hepatocyte suspensions revealed similar metabolite profiles in all 5 in vitro systems used. Ciclesonide was rapidly converted to its active metabolite, desisobutyryl-ciclesonide (des-CIC). Des-CIC was then extensively metabolized to pharmacologically inactive metabolites through oxidation and reduction, followed by glucuronidation. A total of 12 groups of metabolites derived from des-CIC were characterized and identified by liquid chromatography/radioactivity monitor/mass spectrometry. Oxidation occurred on both the cyclohexane ring and the steroid moiety. Hippuric acid formation by cleavage of the cyclohexylmethyl moiety of ciclesonide, followed by aromatization of the cyclohexane ring through multiple steps of hydroxylation, dehydration, and conjugation with glycine, was found in rat, rabbit, and human hepatocyte incubations. The results indicated that ciclesonide and its active metabolite, des-CIC, were extensively metabolized in vitro in animal and human hepatocytes and that the metabolite profiles in mouse, rat, rabbit, and dog hepatocytes were similar to the profiles in human hepatocytes.