COLOR REACTIONS OF NUCLEIC ACID COMPONENTS 303 



minutes after addition of cysteine when the maximum absorption produced 

 by pentoses is observed. The peak of the absorption curve is at 390 mn. 

 Reaction of nucleotides and nucleosides of PNA : — ^Only the purine nucleo- 

 tides of PNA react significantly in the cysteine reaction. The molar ex- 

 tinction coefficient of adenosine-3-phosphate does not differ significantly 

 from that of pure ribose, and is twice as great as that of PNA. 



QuaJititalive determination of PNA and its constituents in presence of DNA and 

 hexoses: — The optical density in the cysteine reaction is proportional to the concen- 

 tration of pentose in the range of concentration between 2 and 40 /xg- per cc. of ribose. 

 The density decreases slowly after maximum absorption is reached, and for this 

 reason the readings of the optical density of the unknown must be carried out simul- 

 taneously with the readings of the standard solution. The purest preparation of DNA 

 showed an optical density at 390 m^, which corresponded to no more than 1.5% of 

 that of equivalent amounts of PNA. As this ratio was obtained for several prepera- 

 tions of DNA, it is probably not due to contamination of the prepartion with 

 small amounts of PNA. Aldohexoses produce with the cysteine reaction a 

 yellow color with an absorption maximum at 412 m/i- This color interferes with the 

 determination of the pentose. This interference, however, can be eliminated by di- 

 chromatic readings. The latter are carried out by finding, around 424 m/x, a wavelength 

 at which a standard of glucose or another aldohexose shows the same optical density 

 as at 390 m/n. D390 — Dm, therefore, is zero for the hexose. It is only slightly lower 

 than Z)39o for the pentose. In this way adenosine -3-phosphate can be still determined 

 in presence of a 5-fold amount of glucose or a 10-fold amount of mannose or galactose. 

 £^390 — D424 , however, is not zero for ketohexoses. Hexuronic acids show an identi- 

 cal absorption curve as pentose, although the molar extinction coefficient is far 

 smaller. When present in large quantities, however, they will interfere with the re- 

 action, and the same is true of methylpentose. It is therefore necessary to correct for 

 the presence of these two classes of saccharides as in the case of the orcinol reaction. 



Evaluation of the PNA reactions: — -Of the three reactions, that with Bial's 

 reagent appears most sensitive and the only one sufficiently investigated as 

 far as the use for quantitative determinations in tissues is concerned. The 

 procedure according to Dische and Schwarz is more sensitive than the other 

 modifications and therefore allows a shorter heating time. This in turn 

 decreases the interference from other sugars. As regards interference from 

 proteins, the orcinol reaction appears to be more influenced than the cyste- 

 ine-H2S04 reaction. The latter reaction is also more suitable for the de- 

 tection of small amounts of PNA in DNA. The reaction with phloroglucinol 

 of Euler and Hahn is much less sensitive, appears more laborious, and has 

 not been sufficiently investigated as far as interference from other sub- 

 stances is concerned. 



II. Determination of Purine and Pyrimidine Bases of 

 Nucleic Acids by Characteristic Color Reactions 



The discovery that the molar ratios of individual purines and pyrimidines 

 in nucleic acid varies from one preparation of nucleic acid to another de- 



