The reaction of diphenylamine with the deoxyribose moieties of DNA was described in 1930 by Dische (1) and is the basis for a quantitative calorimetric method in which DNA is subjected to acid hydrolysis in the presence of diphenylamine (2). RNA does not react with diphenylamine under the conditions of hydrolysis employed for the determination of DNA, 10 minutes at 100Β°C or several hours at 30Β°C (2, 3). This report is of a study in which the diphenylamine reaction was modified so that RNA would react. It was found that with prolonged hydrolysis conditions, 1 or more hours at 100Β°C, RNA does react with diphenylamine and that this reaction can be applied to the quantitative and qualitative analysis of RNA.
Overend, Shafizadeh, and Stacey, (4) found that the diphenylamine reaction is not specific for 2-deoxyribose but rather is a general reaction for 2-deoxypentoses. They also demonstrated that the reaction depends upon the hydrolytic conversion of 2-deoxypentoses to 5-hydroxylevulic aldehyde. The rationale for the study reported in this paper was that a similar conversion of RNA ribose might occur after prolonged hydrolysis. This was based upon known differences in the susceptibilities of deoxyribonucleotides and ribonucleotides to acid hydrolysis (5).