Early Release of ABA from Cell Walls During Rose Petal Protoplast Isolation

During the isolation of rose petal protoplasts, two successive transient abscisic acid (ABA) accumulations were detected when digest medium and tissue slices were assayed simultaneously, using polyclonal antibodies raised against ( + )-ABA. Late accumulation, occurring after more than 1 h of incubation, was observed, whether enzymes were present or not, in a hyper osmotic medium. Very early accumulation, occurring at a maximum of 15 min after the beginning of incubation, was observed only in the presence of cellulolytic and pectinolytic enzymes. The ABA level could be increased by the addition of tetcyclacis, an inhibitor of mono-oxygenase known to delay ABA oxidation. The need for cell wall digesting enzymes to observe this first ABA increase suggested that it was due to a release from the cell wall. Analysis by radioimmunoassay (RIA) and high performance liquid chromatography (HPLC) showed that ABA and another non-acidic immunoreactive compound were released from purified cell walls by the same enzymes.