Stachyose synthesis in isolated mesophyll cells of Cucurbita pepo

A pure preparation of isolated mesophyll cells and a preparation of minor veins heavily contaminated with mesophyll and epidermal tissue were obtained from leaf strips of Cucurbita pepo by enzymatic digestion. The preparations were incubated for 10 min in sodium [ 14 C]bicarbonate in the light to determine their ability to synthesize the major translocated sugar stachyose and its precursor galactinol. Stachyose, galactinol, sucrose, and hexoses accounted for ca. 0.5, 2.5, 55, and 30% respectively of the total 14 C in the neutral sugar fraction in both preparations. The close similarity was believed due to the heavy contamination of the minor vein preparation with mesophyll tissue. Intact leaf tissue incubated under similar conditions incorporated an approximately equal amount of 14 C into the sugar fraction but with a highly dissimilar distribution. Stachyose, galactinol, sucrose, and hexose accounted for up to 50, 20, 13, and 0.2% respectively of the total 14 C activity. Plasmolysis of intact tissue caused a more than 50% decrease in the 14 C incorporation into stachyose and galactinol and a large increase into sucrose and particularly hexoses. The enzymes synthesizing galactinol and stachyose were extracted from the mesophyll preparation but their specific activities on a total chlorophyll basis were 10-fold lower than when extracted from intact tissue. It was concluded that the procedure for isolating free cells, while not affecting sucrose synthesis, was highly detrimental to the synthesis of galactinol and stachyose, and that synthesis of the latter two compounds was most probably confined to the mesophyll.