Characterization of N-1- and N-6-adenosine adducts and N-1-inosine adducts formed by the reaction of butadiene monoxide with adenosine: Evidence for the N-1-adenosine adducts as major initial products

1,3-Butadiene is a known human mutagen and possible human carcinogen; however, the molecular mechanisms of its activity are poorly understood. We have previously shown that the primary metabolite, butadiene monoxide (BM), reacts with guanosine to form N-1-, N-2-, and N-7-guanosine adducts. In this study we characterize the reaction of BM with adenosine; ten adducts identified as diastereomeric pairs of N-1-(1-hydroxy-3-buten-2-yl)adenosine; N-1-(2-hydroxy-3-buten-1-yl)adenosine, N-6-(1-hydroxy-3-buten-2-yl)adenosine, N-6-(2-hydroxy-3-buten-1-yl)adenosine, and N-1-(1-hydroxy-3-buten-2-yl)inosine are characterized. The N-6-adenosine and N-1-inosine adducts were characterized by their UV spectra, H-1 NMR, FAB/MS, and stability studies. The N-6-adenosine and N-1-inosine adducts were stable for up to 168 h at 37 degrees C in phosphate buffer (pH 7.4). The N-1-adenosine adducts, which were unstable at pH 7.4 at 37 degrees C (half-life of 7 and 9.5 h for the two regioisomers), were characterized by their UV spectra and their ability to undergo the Dimroth rearrangement to yield the corresponding N-6-adenosine adducts, or undergo deamination to yield the corresponding N-1-inosine adducts. Upon the reaction of BM with adenosine in phosphate buffer (pH 7.4) at 37 degrees C, the N-1-adenosine adducts were the first to be detected, with the N-6-adenosine and N-1-inosine adducts showing a lag in formation possibly due to the time needed for rearrangement/deamination. Reaction of adenosine with an excess of BM in phosphate buffer (pH 7.4) at 37 degrees C, followed by extraction of the reaction mixture with ethyl ether to remove excess unreacted BM and incubation at 80 degrees C for 1 h, resulted in complete conversion of N-1-adenosine adducts to the corresponding N-6-adenosine and N-1-inosine adducts. Under these conditions, adduct formation exhibited pseudo-first-order kinetics, with the combined N-6-adenosine adducts being formed 3-fold more favorably than the combined N-1-inosine adducts. When incubations were carried out at lower BM concentrations, the N-6-adenosine adducts remained the major detectable adducts at all concentrations. These results show that adenosine, in addition to guanosine, can lead to multiple adducts when incubated with BM, and may be useful in development of biomarkers for exposure to 1,3-butadiene. Characterization of the N-1-adenosine adducts and their rearrangement/deamination products may also contribute to the understanding of mutagenic and carcinogenic mechanisms of 1,3-butadiene.