226 



WALDO E. COHN 



5 7 10 



Fig. 10. Relative distribution coefficients of various nucleotides and organic phos- 

 phates relative to inorganic phosphate as a function of pH, derived from anion-ex- 

 change chromatography" (see Fig. 7). 



ammonium salt can be volatilized. Acetate was used in the first ion-ex- 

 change separations of the 2' and 3' nucleotide isomers.'^- ^"'^^ On the other 

 hand, the dilute HCl solution has an advantage, if subsequent concentra- 

 tion by resorption and re-elution is considered,'^' ^' because of the low ionic 

 strength ; the more concentrated formate or acetate solutions must usually 

 be diluted before resorption, if such is indicated. 



(2) pK's of Isomeric Nucleotides. The 5'-, 2'-, and 3'-phosphate isomers 

 of the various ribonucleosides appear in the order named upon elution un- 

 der the proper conditions for separation. It would appear, compositional 

 factors being the same, that the pK values of the phosphate and amino 

 groups, which differ slightly among the isomers, might be responsible for 

 the separations to be presented. These values are not known for all sets of 

 isomeric nucleotides; those so far reported are collected in Table II. It will 

 be seen that, in general, they vary in magnitude and in direction as one 

 might infer from the order of elution, and hence may properly be held to 

 be responsible for the separations. 



(3) Cyiidylic Acids. The cytidylic acid isomers are eluted rapidly with 

 0.001 A^ HCl solution and with 0.01 A^ formic acid;^" hence, a pH of 3 or 



'" J. X. Khym and W. E. Cohn, unpublished observations. 



31 W. E. Cohn, J. Am. Chem. Soc. 72, 2811 (1950). 



32 H. S. Loring, N. G. Luthy, H. W. Bortner, and L. W. Levy, /. Am. Chem. Soc. 

 72, 2811 (1950). 



" W. E. Cohn, /. Am. Chem. Soc. 71, 2275 (1949). 

 3* W. E. Cohn, /. Cellular Comp. Physiol. 38, Suppl. 1, 21 (1951). 

 « L. F. Cavalieri, J. Am. Chem. Soc. 74, 5804 (1952); 75, 5268 (1953). 

 3« H. Wassermeyer, Z. physiol. Chem., 179, 238 (1928); see Levene and Bass,'^ p. 231. 

 37 P. A. Levene and H. S. Simms, J. Biol. Chem. 65, 519 (1925); 70, 327 (1926); see 

 Levene and Bass,!^ pp. 223, 228. 



