P. Bishop, C. P. WHITTINGHAM
Mar 1, 1963
THE oxygen production resulting from flash illumination of chloroplast fragments was determined, using the method previously described1. Spinach leaves were ground in 0.35 M sodium chloride, 0.037 M magnesium chloride, 0.01 M ascorbate and 0.1 M tris buffer adjusted to pH 7.6. After centrifugation, the chloroplasts were washed once in the grinding medium but without ascorbate and broken in the same medium diluted ten times. The reaction mixture contained, in a final volume of 2 ml., chloroplast fragments containing 0.15 mg chlorophyll and the following (in µmoles): potassium ferricyanide, 3; sodium chloride, 70; magnesium chloride, 10; adenosine triphosphate, 1.5; inorganic phospate, 30; tris buffer, 50, adjusted to pH 7.4. Throughout the measurements the reaction mixture was maintained at 5° ± 1° C and was flushed with nitrogen gas containing less than 1 p.p.m. oxygen. Flashes, from Philips photoflux bulbs, were of sufficient intensity to saturate the reaction (2.8 × 10−6 lumens, 35-msec duration). The change of oxygen yield per single flash was used to determine the rate of decline of activity of the chloroplasts with time.