The deoxyribose in DNA in the presence of acid forms β-hydroxylevulinaldehyde which reacts with diphenylamine to give a blue color with a sharp absorption maximum at 595nm.

In DNA, only the deoxyribose of the purine nucleotides react with Diphenylamine, so that the value obtained represents half of the total deoxyribose present.

Why is this the case?

  • $\begingroup$ Why not start with researching what happens when DNA (not just the deoxyribose part) is placed in acidic solution? $\endgroup$
    – Armand
    Commented Aug 16, 2021 at 19:05

1 Answer 1


As you have correctly stated, the DNA cannot directly react with diphenylamine. First, it has to be cleaved so that the deoxyribose is exposed. In acidic solution and at high temperature, a depurination occurs several orders of magnitude faster than depyrimidination, because N-7 of the purine can be protonated and assist the cleavage.

See this mechanism of the acid-catalyzed depurination:

Acid-catalyzed depurination


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