DISTRIBUTION OF MICRONUTRIENT METALS IN LEAVES AND CHLOROPLAST FRAGMENTS

Abstract

An investigation of the extracellular reactions of chloroplasts in the light (4) and in the dark (5) has focused attention on the possible catalytic role of one or more of the micronutrient metals: iron, manganese, copper, zinc, and molybdenum in the cellular activities of these cytoplasmic bodies. The consideration of a possible participation of one or more of the micronutrient metals involves the knowledge of their relative distribution in the leaf and chloroplasts. This communication reports results of analyses undertaken with this purpose in view. Methods Sugar beet and chard plants were grown in a greenhouse in aerated nutrient solutions of the composition previously described (2), supplemented with the micronutrients B, Mn, Cu, Zn and Mo (1). Large leaves from plants approximately 12 wTeeks old were used for chemical analysis and the isolation of chloroplast fragments. The chloroplast fragments were prepared by trituration of the leaves in the cold without dilution in a (3) and subsequent differential centrifugation as previously described (2), except that the final centrifugation was carried out in a Sorvall Type SS-1 centrifuge, for two hours in the cold, at the maximum speed corresponding to approximately 20,000 g. (A special test disclosed that the Vitajuicer could contribute only insignificant amounts of metal contamination to the analysis.) The chloroplast fragments were dried in an all-glass lyophilizer, and the dried material was kept in the cold in screw-capped bottles. The analyses of the whole leaves were made on samples of fresh leaves which were heated in the autoclave at 5 lbs. pressure for five minutes to stop enzymatic activity and were subsequently dried at 70? C. The dry material was ground in a porcelain mortar and redried at 70? C prior to analysis. The analytical determinations were made on aliquots of the ground leaves and of the lyophilized chloroplast fragments. Chlorophyll was measured as previously described (2). The total nitrogen, found by the micro-Kjeldahl method, in the trichloracetic acid precipitate of a water suspension of the dried material wTas designated as protein nitrogen and was multiplied by the factor 6.25 to give protein values. The metals were determined by the procedures of Parks et al. (15) modified as follows: in the preparation of the ash solution the hydrofluoric acid step was omitted and sulphuric acid was used in the perchloric acid digestion;