A micropipette method for collecting vacuolar sap of intact tannin and parenchyma cells in persimmon

A method for collecting the vacuolar contents of intact tannin and parenchyma cells of persimmon (Diospyros kaki Thunb.) fruit using a micropipette was developed. Thin sections of the mesocarp tissue from mature persimmon fruit, 'Miyazaki-mukaku' and 'Hiratanenashi', were placed on a glass slide. Using a micromanipulator and an inverted microscope, a micropipette was inserted into a vacuole and its contents were withdrawn. A 5-nL sample of vacuole sap was collected per tannin cell from 'Hiratanenashi' and 7 nL from 'Miyazaki-mukaku', whereas only 2 nL was withdrawn from adjacent parenchyma cells. Analyses of the vacuolar sap revealed that the tannin cells of both cultivars contained 10% to 12% (m/v) of tannin as (+)-catechin equivalents and 10% to 13% (m/v) of soluble sugars, whereas the parenchyma cells contained trace amounts of tannins and ≃20% of soluble sugars. Tannin cells contain only a slight amount of sucrose, in contrast to a relatively large amount in parenchyma cells.