A SEPARATION OF THE REACTIONS IN PHOTOSYNTHESIS BY MEANS OF INTERMITTENT LIGHT

Experiments on photosynthesis in intermittent light have been made on two occasions. Brown and Escombe, in 1905, made use of a rotating sector to study the effect of light intensity on the photosynthesis of leaves. They found three-quarters of the light from a given source could be cut out in each revolution of the sector without decreasing the rate of photosynthesis. Willstatter (1918, p. 240) explains that this was probably due to the low concentration of carbon dioxide available for the leaves. The short periods of light would be sufficient to reduce all the carbon dioxide which could reach the cells by diffusion during the dark periods. In 1919-20 Warburg made experiments on Chlorella similar to those of Brown and Escombe on leaves. Instead of stating his results as amount of photosynthesis per total elapsed time, as Brown and Escombe did, he gave photosynthesis per total time during which the cells were illuminated. Since he used sectors which cut out half the incident light in each revolution, the time during which the cells were illuminated was always half of the elapsed time of an experiment. Working with a high intensity of light and a high concentration of carbon dioxide, Warburg found that a given amount of light reduced more carbon dioxide when allowed to fall on the cells intermittently than when allowed to fall on them continuously. The improvement in the yield of the intermittent over the yield in continuous light depended on the frequency of the flashing. With a frequency of four periods per minute the improvement was 10 per cent, and with a frequency of 8000 per minute it was 100 per cent. Warburg proposed two alternative explanations for the improvement in the yield of the intermittent light. Either the reduction of carbon dioxide continues in the dark, or it proceeds twice as fast during the brief light flash as during the same length of time in continuous light. He considers the latter explanation more likely, and assumes that certain steps in the photosynthetic process continue in the dark until a dark equilibrium is reached. After the dark period a short flash of light would find a higher concentration of reactive substance ready for it than is available in continuous light, and would be able to effect more decomposition than an equal amount of continuous light. The experiments described in this paper indicate, we think, that the steps in photosynthesis which proceed in the dark involve what has hitherto been known as the Blackman reaction. Probably the reduction of carbon dioxide is not completed during the photochemical part of the process. A more correct way of representing the sequence of events in intermittent light would be as follows. Two steps are involved in the reduction of carbon dioxide: a reaction in which light is absorbed, followed by a reaction not requiring light -- the so called Blackman reaction. If the light intensity is high the photochemical reaction is capable of proceeding at great speed, but in continuous light it can go no faster than the Blackman reaction. We suppose that the product formed in the photochemical reaction is converted to some other substance by the Blackman reaction, and at the same time the chlorophyll is set free to take part again in the photochemical reaction. If a green cell is illuminated, we think that the photochemical reaction proceeds rapidly until an equilibrium concentration of its product is formed. After this the photochemical reaction proceeds only as fast as the Blackmail reaction removes the intermediate product. If the cell is now darkened, the photochemical reaction stops at once, but the Blackman reaction continues until its raw material, the product formed by the photochemical reaction, is exhausted. After this nothing further happens until the cell is again illuminated. Higher efficiency of the light would be obtained if each light flash lasted only long enough to build up the equilibrium concentration of the intermediate product, and each dark period were long enough to allow the Blackman reaction time to use up all the intermediate product present at the moment the light period ended. In Warburg's flicker experiments the light and dark periods were always of equal length. He found that the amount of work done by the light could be increased by shortening both the light and the dark periods. This indicates that his light periods were too long for maximum efficiency. In the latter part of each light period the photochemical reaction must have been brought down to near the speed of the Blackman reaction. Using 133 light flashes per second, Warburg obtained an improvement of 100 per cent over the continuous light yield. We were able to improve the continuous light yield 300 per cent to 400 per cent by using only 50 flashes per second and making the light flashes much shorter than the dark periods. This opened the possibility of determining the length of the dark period necessary for the complete removal of the intermediate product formed in a light flash of given intensity and duration. Lengthening the dark period should improve the yield until there is time enough for all the intermediate product formed in each light flash to be removed before the next light flash. In this paper we describe experiments which show that the necessary dark time is about 0.03 to 0.4 of a second, depending on the temperature. Further experiments are described to show certain characteristics of the reactions taking place both in the light and in the dark.