Chromatographic determination of nucleotides in the suspending medium from irradiated yeast.

Yeast cells suspended in distilled water, when damaged by prolonged sublethal dosages of ultraviolet radiation, liberate substances into the suspending medium capable of producing marked proliferation of normal yeast cells (1). This effect is encountered without appreciable killing of the irradiated cells. Previous communications have presented evidence that members of the vitamin B complex and amino acids are released from the irradiated cells in considerable quantities (2, 3). These substances account for a large part, but not all, of the proliferation-promoting effect. Release of nucleotides and nucleosides into the suspending medium has also been reported (4). The presence of these substances is indicated by the following facts. (1) After irradiation the suspension medium is found to have an absorption spectrum with a maximum at 2600 A, characteristic of nucleic acid derivatives (5). (2) Chemical analyses of the suspension medium by the method of Kerr and Blish (6) and Kerr (7) showed large amounts of nucleotide and nucleoside and free purine nitrogen compared to unirradiated controls. The importance of these compounds is indicated by the following evidence. (1) Th e c h emically isolated nucleotide fraction when combined with amino acids and vitamin B factors duplicated the proliferation-promoting effect of the original material. (2) The amount of growth is proportional to the concentration of nucleic acid derivatives as measured by the absorption at 2600 A. It seemed desirable to separate quantitatively the nucleotide-like components of the irradiated suspension medium and evaluate their r&e in the proliferation-promoting effect. The recent application of the anion exchange column to the separation of nucleotides by Cohn (8) seemed very suitable for this purpose. Material prepared in this way is free of toxic heavy metals which may contaminate chemical preparations.