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Volatile terpenoid compounds, potentially derived from carotenoids, are important components of flavor and aroma in many fruits, vegetables and ornamentals. Despite their importance, little is known about the enzymes that generate these volatiles. The tomato genome contains two closely related genes potentially encoding carotenoid cleavage dioxygenases, LeCCD1A and LeCCD1B. A quantitative reverse transcriptase‐polymerase chain reaction analysis revealed that one of these two genes, LeCCD1B, is highly expressed in ripening fruit (4 days post‐breaker), where it constitutes 0.11% of total RNA. Unlike the related neoxanthin cleavage dioxygenases, import assays using pea chloroplasts showed that the LeCCD1 proteins are not plastid‐localized. The biochemical functions of the LeCCD1 proteins were determined by bacterial expression and in vitro assays, where it was shown that they symmetrically …