Improved rates of CO2-fixation by intact chloroplasts isolated in media with KCl as the osmoticum.

Intact chloroplasts were isolated from spinach leaves using media with either 330 mM sorbitol or 200 mM KCl as the osmoticum. Chloroplasts isolated in KCl exhibited higher rates of CO2-dependent oxygen evolution in nine out of ten experiments, the average increase being 43%. Chloroplasts isolated in KCl routinely achieved rates of CO2-dependent oxygen evolution of 200-300 μ mol·mg chlorophyll(-1)·hour(-1) at 20°C. Intact chloroplasts were also isolated in media with 200 mM NaCl or choline chloride but the rates of CO2 fixation were not superior to those isolated in sorbitol media. The K(+) content of chloroplasts isolated in KCl media was higher than for chloroplasts isolated in sorbitol. It is suggested that the use of KCl as an osmoticum prevents the loss of chloroplast K(+) which can occur during isolation in sorbitol media. Chloroplasts isolated in KCl lost, on average, 36% of the initial CO2 fixation activity after storage for four hours on ice, compared to 24% loss of activity for chloroplasts isolated in sorbitol. This increased loss of activity was not observed if KCl was used in the grinding medium and sorbitol or glycinebetaine in the resuspension media. For measurement of the maximum photosynthetic capacity in vitro, the use of KCl in the grinding medium may be better than sorbitol.