HPLC DETERMINATION OF BENZYLADENINE RESIDUES IN MICROPROPAGATED APPLEEXPLANTS

A specific reversed-phase high-performance liquid chromatographic (HPL C) method has been developed to determine the concentration of benzyla denine (BA) [N-(phenylmethyl)-1H-purin-6-amine; CAS 1214-39-7] in micr opropagated apple explants. BA is extracted into 95% ethanol, the etha nol evaporated, and the resulting extract residue redissolved in ammon ium formate buffer (pH 3.7), which is then washed with petroleum ether . Further cleanup is performed with exchange chromatography (DEAE-cell ulose) before the application of the solid-phase extraction procedure. Methanolic eluates of BA are submitted to HPLC determination using a C-8 column and an isocratic system of 30:70 (v/v) water-methanol conta ining 0.05% acetic acid. Recovery and reproducibility assessments indi cate good accuracy (overall mean relative recovery of 87%) and precisi on (coefficient of variation from 0.4 to 7.7) over the BA concentratio n range 0.5-300 ppm, with a limit of quantitation of 0.5 ppm.