180 J. N. Davidson 



it is necessary first to isolate the various components, 

 known and unknown, of the ribonucleotide fraction. Existing 

 methods for the separation of ribonucleotides by ion-exchange 

 chromatography or by paper chromatography all suffer from 

 certain disadvantages, and an attempt has therefore been 

 made to utilize the mobility of nucleotides in an electric 

 field. R. M. S. Smellie has succeeded in developing a method 

 for the separation of ribonucleotides by ionophoresis either 

 on agar gel or preferably on paper, using a technique similar 

 to that described by Durrum (1950) for amino-acids. The 

 nucleotides separate in the order: uridylic acid, guanylic acid, 

 adenylic acid and cytidylic acid, in decreasing order of 

 mobility, and their positions after separation can easily be 

 located in ultraviolet light by the procedure of Holiday and 

 Johnson (1949). If a permanent record is desired, a photo- 

 graph may be made by the method of Markham and Smith 

 (1949). The nucleotides can be removed quantitatively on 

 elution by the technique of Consden, Gordon and Martin 

 (1947) and can be identified by their ultraviolet absorption 

 spectra and estimated by determination of phosphorus. 

 When the method is applied to a mixture of nucleotides to 

 which radioactive inorganic phosphate has been added, the 

 nucleotides on isolation show no contamination with radio- 

 active material. 



The ionophoresis method can be applied to the analysis 

 of a purified ribonucleic acid, but it can also be used with 

 the mixture of ribonucleotides obtained in the Schmidt- 

 Thannhauser separation procedure (Smellie and Davidson, 

 1951). In this case it is preferable to acidify the KOH 

 digest with perchloric acid rather than with trichloracetic 

 acid. When the ribonucleotide fraction is submitted to 

 ionophoresis, uridylic acid is found to be preceded by a fast 

 moving spot showing up faintly in ultraviolet light. This 

 unknown component accounts for 10-20 per cent of the 

 total phosphorus in the fraction (Table VI) when liver tissue 

 is employed, and consists of a small amount of inorganic 

 phosphate (derived from "phosphoprotein") and a larger 



