Separated mesophyll cells from cotton (Gossypium hirsutum var. Stoneville 1613 Glandless) were isolated with pectinase and mechanical agitation. The separated cells had rates of light-dependent CO(2) fixation between 50 to 100 mumoles CO(2) per mg chlorophyll per hour. The presence of Ca(2+) in the incubation medium did not significantly affect the type of photosynthetic products formed, but 2 mm Ca(2+) did cause a 50% decrease in the appearance of photosynthetic products in the incubation medium. The movement of all types of products (sugars, organic, and amino acids) out of the cells was reduced similarly by the Ca(2+). Light had no affect on the movement of products out of the cells, whereas 1 mm ethylenediaminetetra-acetate greatly increased the movement. The addition of 1.6 mm NH(4)Cl to the cell suspensions caused a large increase in the amount of fixed (14)C appearing in the amino acid fraction and a decrease in the sugar fraction. These metabolic changes in the cells were reflected in the movement of products out of the cells so that the incubation medium also contained a larger amount of label in amino acids and a smaller amount in sucrose. Although the cell plasma membrane restricted the movement of soluble products, it did not discriminate significantly between the types of products moved.