Formation of the N2-acetyl-2,6-diaminopurine oligonucleotide impurity caused by acetyl capping

Abstract The acetyl ‘capping’ reaction routinely employed during phosphorothioate oligonucleotide synthesis has been implicated in the formation of an impurity species with a mass 41 amu greater than the expected oligonucleotide molecule. The impurity has been found to arise by conversion of a protected guanine nucleobase to N 2 -acetyl-2,6-diaminopurine. A two-part mechanism is proposed consisting of transamidation of the protecting group on guanine and substitution of guanine’s O 6 atom.