Photoreduction of Chlorophyll a in the Presence of Ascorbic Acid in Pyridine Solutions.

Abstract

Over the last decade, Russian investigators?among whom Krasnovsky, Evstigneev, and Gavrilova are most prominent?have published many papers describing the photoreduction of chlorophyll in dilute solutions. Since the photoreduction has been reported to be reversible and energy-storing, the reaction is of great interest because it could be related to the primary photochemical process of photosynthesis. This paper describes experiments carried out after initial attempts by the writer failed to reproduce certain of the Russian findings. Krasnovsky and Brin (5) reported that, when an oxygen-free pyridine solution of chlorophyll a (10~5 M) and ascorbic acid was illuminated with red light, the blue-green color characteristic of chlorophyll faded away and a pink solution was formed: the absorption bands of chlorophyll disappeared and a new absorption band with a maximum at 523 ??? appeared. Lynch and French have confirmed this observation. Krasnovsky et al (3, 4, 5) found that, with a moderate concentration of ascorbic acid (0.05 M ) and in the absence of water, the bleaching was reversible?i.e., in the dark following bleaching, chlorophyll was regenerated and a blue-green color was restored. In solutions containing water or more ascorbate (0.10 M), the bleaching was irreversible since pheophytin a rather than chlorophyll was generated in the dark. The Russian workers apparently regarded the formation of pheophytin as an unimportant secondary process unrelated to the photoreduction. The results of preliminary experiments of the writer differed significantly from those of the Russians'. In dry pyridine solutions, no bleaching was detected at all. When a certain amount of water was added to the samples, a rapid bleaching resulted but colorless rather than pink solutions were formed ; moreover, no reversibility could be demonstrated because the bleached chlorophyll was quantitatively converted into pheophytin in the dark following illumination. These findings thus failed to corroborate the existence of a pink photoreduced chlorophyll and failed also to show any reversible reaction at all. Since the Russian studies were not only convincing in their extent, but also important in providing evidence for the only known, reversible, light-sensitized, energy-storing redox reaction of chlorophyll, a thorough reinvestigation was undertaken. Part of the results of this study are reported in this paper.