DETERMINATION OF IAA AND ABA IN THE SAME PLANT-SAMPLE BY A WIDELY APPLICABLE METHOD USING GC-MS WITH SELECTED ION MONITORING

A method for the purification and subsequent quantification of indole- 3-acetic acid (IAA) and abscisic acid (ABA) from the same sample of hi ghly pigmented green tissue has been developed and tested in several s pecies. Solvent partitioning and high-performance liquid chromatograph y (HPLC) were used for purification. Separate fractions from HPLC-cont aining IAA and ABA were analyzed by gas chromatography-mass spectromet ry (GC-MS) using selected-ion monitoring (SIM). Isotope dilution was u sed to correct for incomplete recovery. Results are presented for tiss ue samples from 11 different species and five different plant organs. The method can be completed, for both IAA and ABA, for two samples in 8 h by an experienced technician. IAA and ABA were the dominant peaks in the gas chromatograms from HPLC-purified samples, and amounts of ab out 1 ng can be detected. The extract was partitioned into an aqueous solution of pH 9.5, a step suspected of ester hydrolysis. By analyzing samples known to contain esters of IAA and ABA and comparing the resu lts with methods which excluded this step, we have shown that this par titioning does not result in erroneously high values due to ester hydr olysis. A direct comparison of the method with one in which HPLC was n ot employed indicates that our method measures IAA and ABA in samples in which these compounds are not detectable when HPLC is omitted. Thus , HPLC is an essential purification step for samples where contaminati ng compounds co-purify with IAA and ABA through the solvent-partitioni ng steps.