CHAPTER 7 



Separation of Nucleic Acid Components by 

 Chromatography on Filter Paper* 



G. R. WYATT 



Page 



243 

 I. Introduction 



II General Technique of Paper Chromatography ^** 



III. Detection of Purine and Pyrimidine Derivatives on Filter Paper .... 246 



1. Purine and Pyrimidine Bases 



2. Nucleosides and Nucleotides 



IV. Solvent Systems 



1 General and Theoretical Considerations ^ 



2. Separation of Purine and Pyrimidine Bases and Nucleosides 250 



3. Separation of Nucleotides ;\t ' , • ' » -j oi=;7 



V. Quantitative Estimation of the Nitrogenous Components of Nucleic Acids 257 



1. Hydrolysis of Deoxypentose Nucleic Acids 257 



2. Hydrolysis of Pentose Nucleic Acids 25J 



3. Quantitative Technique 



VI. Chromatography of Nucleic Acid Sugars 



VII. Addendum 



I. Introduction 



The feasibility of separating nucleic acid components by chromatography 

 on filter paper was first demonstrated in 1947-1948 by Vischer and Char- 

 gaffi and by Hotchkiss.^ Since then progress has been rapid and paper 

 chromatography as a quantitative technique has now attained more suc- 

 cess with this group of compounds than with any other. This is due largely 

 to the intense absorption of ultraviolet light by purine and pynmidme 

 derivatives, which facilitates their detection on paper and makes possible 

 their direct estimation, once separated, by spectrophotometry. The nitrog- 

 enous bases may be accurately estimated by ordinary methods of paper 

 chromatography from less than 0.5 mg. of nucleic acid, or, with special re- 

 finements of technique, from as little as a few micrograms. Application of 

 these methods has led to knowledge of the quantitative composition of 

 nucleic acids from a variety of sources, to recognition of two pyrimidine 

 bases not previously known to occur in nucleic acids, and, together with 

 * Contribution No. 116, Forest Biology Division, Science Service, Department of 

 Agriculture, Ottawa, Canada. 



1 E. Vischer and E. Chargaff, /. Biol. Chem. 168, 781 (1947). 



2 R. D. Hotchkiss, J. Biol. Chem. 175, 315 (1948). 



243 



