A HPLC assay for ABA and bound ABA is described. The method uses conventional acid-base ether extractions for partial cleanup of the plant extracts followed by Sephadex G-25 column fractionation. The ABA is separated and detected by HPLC on a 3 m x 2.1 mm i.d. column of SCX-Zipax with a uv detector. The method is sensitive to less than 10 ng of ABA. HPLC analyses of ABA have been performed on extracts of leaves, stems, and roots of soybeans, pinto beans, cotton, apple seedlings, and the albedo and flavedo sections of orange peels.
Abscisic acid (ABA) is an important plant hormone which is distributed widely in the plant kingdom. Although the complete biological role of ABA in plants is not fully understood, ABA has been implicated in such physiological processes as abscission, senescence, bud dormancy and stomata1 control.
Reviews of the chemistry and physiology of ABA are given by Addicott and Lyon (1) .and Milborrow (2). Early attempts at bioassays of ABA were troubled by interference of other plant hormones and inhibitors. More recently, new physical methods have been developed for the quantitative analysis of ABA; these methods include optical rotary dispersion (ORD) (3), gas chromatographic analysis (GC) of the methyl ester and trimethyl silyl derivatives of ABA with flame ionization detectors (FID) (4,5), and the methyl ester of ABA by GC analysis with electron capture detectors (6,7).
High-performance liquid chromatography (HPLC), using efficient column packings in conjunction with sensitive uv detectors, has found increasing use for the rapid separation and quantitation of nonvolatile, sensitive biological compounds. The possible utility of HPLC for the determination of plant cytokinins was suggested in a paper by Pool and Powell (8). Only recently have preliminary results been presented on the utilization of HPLC for the separation and quantitation of plant hormones (9-11). This paper describes a convenient, sensitive assay for ABA based upon the HPLC separation of ABA in a wide variety of plant tissue extracts.