THE NUCLEIC ACID CONTENT OF TISSUES AND CELLS 5 



acid (TCA) or perchloric acid (PCA) allows the DNA and protein to be 

 centrifuged do\ATi (VI) while the PNA remains in solution as acid-soluble 

 nucleotides (IV). The two nucleic acids can be determined on the basis of 

 the P content of IV and VI, if the amount of "phosphoprotein" in the 

 former is small enough to be neglected.^ A closer estimate of PNA is ob- 

 tained by precipitating the inorganic phosphate from "phosphoprotein" in 

 IV"' 26; PNA phosphorus (PNA-P) is represented by the difference (IV-V). 

 For rat liver tissue nonnucleotide phosphorus derivatives in IV are now 

 known to represent about 25 % of the total P of the fraction." High amounts 

 of nonnucleotide P have been found by Logan et alP in the same fraction 

 from brain tissue ; these varied from 66 % of the total NAP in white matter 

 to 37.5 % in gray matter. Fraction IV from bacterial and yeast cells also 

 contains relatively large amounts of nonnucleotide P.^^-^i In Streptococcus 

 faecalis most of the DNA remains undissolved during the alkaline incuba- 

 tion. -^ 



Various modifications of the Schmidt and Thannhauser procedure have been intro- 

 duced. Schmidt et al.^^ used 1% egg albumin to produce a flocculent precipitate (VI), 

 since the relatively small amounts of DNA in Arbacia eggs were otherwise difficult 

 to precipitate after acidification of III. Davidson et al.^^ adapted the procedure to 

 the very small amounts of PNA and DNA in tissue explants growing in vitro, and 

 Scott and Fraccastoro'^ report a modification suitable for micro-amounts in tissue 

 sections. Where digestions of fractions IV and VI are carried out with concentrated 

 PCA at 210°, NaOH has to be used instead of KOH for the alkaline digest in order to 

 avoid formation of insoluble potassium perchlorate. If PNA is to be measured by 

 ultraviolet absorption, PCA is a more convenient precipitating agent than TCA 

 because of its low optical density in the wavelength region 257 to 270 m/z. If necessary, 

 it can subsequently be removed as potassium salt, as Davidson and Smellie'^ have 

 done in order to obtain solutions of nucleotides suitable for treatment by paper 

 inophoresis. This method enables the PNA-P to be determined from the summation 

 of the P content of its component nucleotides, thus eliminating errors due to the 

 concomitant P of fraction IV. 



" G. E. Delory, Biochem. J. 23, 1161 (1938). 

 26 G. C. Mathison, Biochem. J. 4, 233 (1909). 

 " J. N. Davidson and R. M. S. Smellie, Biochem. J. 52, 599 (1952). 



28 J. E. Logan, W. A. Mannell, and R. J. Rossiter, Biochem. J. 51, 470 (1952). 



29 P. Mitchell and J. Moyle, J. Gen. Microbiol. 5, 966 (1951). 

 ="> J. M. Wiame, J. Biol. Chem. 178, 919 (1949). 



=>• H. S. A. Sherrat and A. J. Thomas, /. Gen. Microbiol. 8, 217 (1953). 

 32 G. Schmidt, L. Hecht, and S. J. Thannhauser, /. Gen. Physiol. 31, 203 (1948). 

 '3 J. N. Davidson, I. Leslie, and C. Waymouth, Biocheyn. J . 44, 5 (1949). 

 'M. F. Scott and A. P. Fraccastoro, J. Natl. Cancer Inst. 13, 203 (1948). 

 " J, N. Davidson and R. M. S. Smellie, Biochem. J. 52, 594 (1952). 



