Non-Destructive Collection of Apoplast Fluid From Developing Tomato Fruit Using a Pressure Dehydration Procedure

Abstract

Pressure dehydration techniques were evaluated for the collection of samples of apoplast fluid from the outer pericarp tissues of developing tomato fruit. Sap was expelled under pressure through a cannula inserted into the selected tissue of a fruit sealed in a Scholander pressure bomb. Osmolality and concentrations of K⁺ and hexoses were significantly lower and pH higher in sap exudates than in bulk fruit tissues. After infusion of fruit with trisodium 3-hydroxy-5,8,10-pyrenetrisulfonate as an apoplastic marker, the dye was confined to the same fruit compartment as that which yielded sap under pressure. During extended periods of sap exudation at 24ºC under 1.4 MPa or more, the initial 100 μL of exuded sap was obtained without contamination from protoplasmic contents followed by a proportionate increase in sap osmolality with the aggregate volume. However, when the exuded sap was collected at 4ºC under the applied pressure of 0.6-1.0 MPa, the composition of the sap remained constant with time and was independent of the applied pressure. This permits the collection of a relatively large volume of exuded fluid, up to about 500 μL sap per 70 g fruit, at an initial flow rate of 100-150 μL h-1. We conclude that the pressure dehydration technique under the described conditions allows the collection of uncontaminated apoplastic fluid from the pericarp of intact developing tomato fruit.