Structural identification of the pyrimidine derivatives formed from N-(deoxyguanosin-8-yl)-2-aminofluorene in aqueous solution at alkaline pH

Abstract

The major aminofluorene-DNA derivative formed from the carcinogen N-acetyl-2-aminofluorene in vivo in rat liver is N-(deoxyguanosin-8-yl)-2-amino-fluorene. This nucleoside is hydrolyzed in aqueous solution at alkaline pH through the 7-8 guanine bond to form two pyrimidine derivatives which were separated by Sephadex LH-20 column chromatography and thin-layer chromatography on silica. From chemical, u.v., i.r., n.m.r. and mass spectral analysis the pyrimidine derivatives have been identified as 1-[6-(2, 5-diamino-4-oxopyrimidinyl-N⁶-deoxyriboside)]-3-(2-fluorenyl)ureas, which probably are stereoisomers. Similar products were isolated from enzymatic hydrolysates of DNA reacted with N-hydroxy-2-aminofluorene under mildly acidic conditions (pH 5) and subsequent treatment with 0.1 N NaOH. Kinetic studies of the hydrolysis reaction showed that it occurs already at a measurable rate at pH 9.5 and 37°C. The reaction is catalyzed by Mg²⁺ and Mn²⁺ ions and by alkaline phosphatase from E. coli.