Analysis of growth components in Allium roots

Vacuoles were prepared from cultured parsley cells by polyamine-induced rupture of protoplasts. Acid-phosphatase activity, associated exclusively with the vacuoles, served for determination of vacuole yield in subsequent transport studies. Isolated vacuoles rapidly accumulated [2-¹⁴C]apigenin 7-O-(6-O-malonylglucoside) or 2-¹⁴C]-methyl D-6-O-malonylglucoside added at approximately 20 nM and 1.5 M concentration, respectively, to the incubation mixture. The accumulation was linear with time and strongly dependent on alkaline buffer conditions as well as on the age of the vacuole preparation. Subsequent addition of a malonic hemiester esterase did not relase the label from the vacuoles. Moreover, neither [2-¹⁴C]apigenin 7-O-glucoside or [2-¹⁴C]malonic acid accumulated in the vacuoles under any assay conditions, nor did such compounds or -methyl D-glucopyranoside, a malonic diester, and a succinic monoester inhibit transport of the acylated flavonoid. Transport was, however, inhibited by -methyl D-6-O-malonylglucopyranoside. Vacuoles which had been incubated for more than 40 min at pH 8.0 did not stain any more with neutral-red dye and concomitantly lost the previously accumulated acylated glucoside. Our data confirm that malonylglucoside uptake by parsley vacuoles involves selective transport sites. It is suggested that changes in the molecular symmetry of the malonylglucosides are responsible for vacuolar trapping of flavonoids in parsley.Abbreviation DEAE diethylamionethyl