ISOLATION AND COMPOSITION OF PENTOSE NUCLEIC ACIDS 395 



(including methylcytosine) are always unity. ^* The implications of these 

 findings are discussed in Chapter 10 and elsewhere in this book. Here the 

 results obtained by the analysis of PNA preparations will be examined 

 with the view to finding out whether similar or different generalizations 

 can be made with regard to the composition of PNA. 



2. Analytical Procedures 



The methods used for the estimation of nucleic acid components have 

 been discussed in Chapters 5 to 9. Here only a brief description will be 

 given of those methods which served to obtain most of the results pre- 

 sented in the succeeding sections. 



Method 1.^^ Hydrolysis of PNA by N HCl to a mixture of purine bases and pyrimi- 

 dine mononucleotides followed by separation of the components of the mixture by 

 paper chromatography, elution, and spectrophotometric estimation. 



Method 2^°- " Hydrolysis of PNA by dilute alkali to a mixture of mononucleotides, 

 separation of the mixture by paper chromatography, followed by elution and spec- 

 trophotometric estimation. In this method buffered isobutyric acid is used as the 

 organic phase in chromatography. Guanylic acid and uridylic acid, which occupy the 

 same position on the chromatogram, are eluted together and their respective concen- 

 trations determined from the extinction values of the mixed eluate at two different 

 wavelengths by means of simultaneous eciuations. The value for the molecular extinc- 

 tion of guanylic acid in M phosphate buffer of pH 7.0 which was used by the authors 

 to calculate the concentration of guanylic acid and of uridylic acid was obtained 

 from measurements carried out on an impure sample of sodium guanylate and cor- 

 rected according to the nitrogen content of this sample. Recent determinations of 

 the molecular extinction of guanylic acid purified bj' ion-exchange chromatography 

 have shown that the value used was too high. [Cf. Chapter 14.] Consequently the 

 results of the analysis of pentose nucleic acids by Chargaff et «/."< and by Magasanik 

 and Chargaff" report values for guanylic acid which are too low. The compositions 

 of these preparations have been recalculated using the following extinction coeffi- 

 cients taken from a recent paper of Elson et al.^^ (Ae is the difference between the 

 molecular extinction at the wavelengths indicated and that at 290 fi.) 



Nucleotide \,mn At 



Adenylic acid 260 15.12 



Guanylic acid 265 7.18 



252.5 10.74 



245 8.87 



Cytidylic acid 270 6.87 



Uridylic acid 265 9.49 



261 9.80 



245 5.47 



58 E. Chargaff, Experientia 6, 201 (1950): Federation Proc. 10, 654 (1951). 



" J. D. Smith and R. Markham, Biochem. J. 46, 509 (1950). 



60 B. Magasanik, E. Vischer, R. Doniger, D. Elson, and E. Chargaff, J . Biol. Chem. 



186, 37 (1950). 

 «' D. Elson, T. Gustafson, and E. Chargaff, J. Biol. Chem. 209, 285 (1954). 



