31P‐NMR DIFFERENTIATION BETWEEN INTRACELLULAR PHOSPHATE POOLS IN COSMARIUM (CHLOROPHYTA)1

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Abstract
31P nuclear magnetic resonance (NMR) spectroscopy of intact Cosmarium sp. cells is presented as a suitable tool for the differentiation of intracellular accumulation pools of polyphosphates.The cold trichloroacetic acid (TCA) insoluble fraction is shown to contain most of the total cellular phosphate in the phosphate rich Cosmarium cells. Moreover, evidence from a 31 P‐NMR study and electron microscopic observations of cold TCA treated Cosmarium cells indicate that this fraction consists mostly of polyphosphates which seem to retain the native morphological structure observed in the untreated cells. The determination of orthophosphate in the hot water extract of Cosmarium cells did not measure the polyphosphate pools. Determination of total phosphorus content in the hot water extract rendered a value three times higher than the frequently used orthophosphate determination procedure. However, as revealed by the 31P‐NMR spectra and the chemical analyses of the extract and of the treated cells, even total phosphorus in the extract measured only 30% of the total cellular phosphorus. 31P‐NMR enabled the unequivocal chemical identification of the major phosphate compounds in the hot water extract (“Surplus P”) as orthophosphate and polyphosphates of about 10 phosphate units chainlength. More than 70% of the accumulation pool of polyphosphates was still in the cells after extraction. However, the electron microscopy study revealed that the native granular structure of polyphosphates had been destroyed by the hot water extraction procedure.