primary source of IPP that is utilized for the synthesis of carotenoids in plants.Feeding studies with carotenoids are complicated by their hydrophobic nature and susceptibility to a variety of co-oxidation reactions. As an alternative, labeling studies with 18O2 have proved a valuable tool for studying carotenoid metabolism and ABA biosynthesis. In these experiments, intact plant tissue with a high rate of ABA biosynthesis was placed in an atmosphere of 18O2 and the incorporation of oxygen at the different positions was analyzed by mass spectrometry. Using negative chemical ionization (NCI), the molecular ion of the methyl ester of ABA (Me-ABA) M- at m/z 278 is the most abundant peak (Fig. 6A). When leaves are stressed and placed in an 18O2 atmosphere, the majority of the ABA is labeled with a single 18O and M- at m/z
280 (Fig. 6B). This indicates that ABA is derived from a precursor that already contains oxygen at two of the four positions. Based upon the fragmentation pattern, the position of the 18O was found to be primarily in the carboxyl group
(Fig. 6B). By H2 18O labeling, it was shown that the oxygen introduced in the final step of the pathway, the oxidation of AB-aldehyde to ABA, is derived from water. These labeling patterns are consistent with a derivation from epoxy-carotenoids. The 18O incorporation would most likely result from an aldehyde introduced by the oxidative cleavage of the carotenoid. The hydroxyl of the epoxy-carotenoid precursor is converted to the 4'-keto and the 1'-hydroxyl of ABA is derived from an epoxy group. Hence, there was noincorporation at these positions. Longer incubations in 18O2 do result in a