Two new products from the incubation of beta-carotene with intestinal mucosa homogenates of human, monkey, ferret, and rat were isolated using high-performance liquid chromatography (HPLC). Identification by comparing retention times in HPLC, by monitoring ultraviolet/visible spectra, by reduction to corresponding alcohol, by oxime formation, and by mass spectrometry demonstrated that they are beta-apo-13-carotenone and beta-apo-14'-carotenal. These compounds were not found in incubations done without intestinal homogenates or with disulfiram as an inhibitor. Under standard incubation conditions, these products increased linearly for 60 min and up to a protein concentration of 1.5 mg/mL and increased along with increasing concentrations of beta-carotene. Therefore, they are enzymatic cleavage products from beta-carotene. The formation of the beta-apo-13-carotenone and beta-apo-14'-carotenal provides direct evidence for an enzymatic excentric cleavage mechanism.