PHOTOCHEMICAL ACTIVITY OF ISOLATED SPINACH CHLOROPLASTS IN RELATION TO REACTION CONDITIONS

Abstract

Oxygen production by illuminated chloroplasts in p-benzoquinone solution proceeds most rapidly at neutrality, is more rapid with sorbitol borate than phosphate buffer, is insensitive to buffer molarity, and does not require carbon dioxide. In 0.08% quinone solution, the oxygen yields are 65 to 75% of the theoretical with crude spinach chloroplasts and 85 to 95% with separated chloroplasts. At concentrations above 0.08%, quinone strongly inhibits the photochemical reaction in which it serves as hydrogen acceptor, decreasing the total yield of oxygen as well as the initial rate of its production. Separation of the cell sap and cytoplasm from the chloroplasts by centrifuging and washing treatments largely eliminates dark reactions but also decreases photochemical activity in Hill and quinone solutions. Centrifuged chloroplasts have similar activities when resuspended in cell sap, distilled water, phosphate buffer, sucrose solution, or phosphate–sucrose solution. The supernatant fraction obtained by high speed centrifuging shows higher photochemical quotients in quinone solution and lower photochemical quotients in Hill solution than the chloroplast fraction diluted to the same chlorophyll concentration. The photochemical activity of crude and separated chloroplast suspensions in Hill and quinone solutions is inhibited to various degrees by 10−3M azide and hydroxylamine, but not by 10−3M fluoride. The presence of tannins or 2 × 10−3M adenosine triphosphate has no effect, but 1/10th saturated vitamin K weakly inhibits photochemical activity in quinone solution. The constant relation which was observed between the photochemical yields of H+ and oxygen in Hill solution, employing different reaction periods, chloroplast concentrations, or Hill solution minus certain components, indicates that these products have a common source, and that illuminated chloroplasts effect a photodecomposition of water.