Detection of phosphodiester adducts formed by the reaction of benzo[a]pyrene diol epoxide with 2'-deoxynucleotides using collision-induced dissociation electrospray ionization tandem mass spectrometry

Abstract

In this study, we investigated the products formed following the reaction of benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (B[a]PDE) with 2′-deoxynucleoside 3′-monophosphates. The B[a]PDE plus 2′-deoxynucleotide reaction mixtures were purified using solid phase extraction (SPE) and subjected to HPLC with fluorescence detection. Fractions corresponding to reaction product peaks were collected and desalted using SPE prior to analysis for the presence of molecular ions corresponding to m/z 648, 632, 608 and 623 [M−H]− consistent with B[a]PDE adducted (either on the base or phosphate group) 2′-deoxynucleotides of guanine, adenine, cytosine and thymine, respectively, using LC-ESI-MS/MS collision-induced dissociation (CID). Reaction products were identified having CID product ion spectra containing product ions at m/z 452, 436 and 412 [(B[a]Ptriol+base)−H]−, resulting from cleavage of the glycosidic bond between the 2′-deoxyribose and base, corresponding to B[a]PDE adducts of guanine, adenine and cytosine, respectively. Further reaction products were identified having unique CID product ion spectra characteristic of B[a]PDE adduct formation with the phosphate group of the 2′-deoxynucleotide. The presence of product ions at m/z 399 and 497 were observed for all four 2′-deoxynucleotides, corresponding to [(B[a]Ptriol+phosphate)−H]− and [(2′-deoxyribose+phosphate+B[a]Ptriol)−H]−, respectively. In conclusion, this investigation provides the first direct evidence for the formation of phosphodiester adducts by B[a]PDE following reaction with 2′-deoxynucleotides.

Ultra-performance liquid chromatography–tandem mass spectrometry for rapid and highly sensitive analysis of stereoisomers of benzo[a]pyrene diol epoxide–DNA adducts

“…are consistent with previous work [22,23], further validating the assigned chirality and stereochemistry of the stereoisomers of anti-BPDE-N 2 dG. Fig.…”

“…A number of methods have been developed for detection of BPDE-DNA adducts, including capillary electrophoresis (CE)-laserinduced fluorescence (LIF) [13][14][15][16], accelerator mass spectrometry [17,18], liquid chromatography or capillary electrophoresis-mass spectrometry [19][20][21][22][23][24][25][26], and 32 P-postlabelling [27]. However, only few methods demonstrate the capability of resolving the stereoisomers of BPDE-DNA adducts.…”

Preparation, identification and analysis of stereoisomeric anti-benzo[a]pyrene diol epoxide–deoxyguanosine adducts using phenyl liquid chromatography with diode array, fluorescence and tandem mass spectrometry detection

“…The measured CD spectra of the purified (−)-trans-, (+)-trans-and (−)-cisanti-BPDE-N 2 -dGs are consistent with previous work [19,25], further validating the assigned chirality and stereochemistry. The purified amount of (+)-cis-anti-BPDE-N 2 -dG is not enough to measure their CD spectrum due to its low yield (Table 1, less than 3% in the total adducts).…”

“…To produce enough anti-BPDE-dG standards, a large amount of highly carcinogenic anti-BPDE is often required (1-10 mg) [19,20,23]. An alternative is to use the reaction of dGMP and anti-BPDE, which only requires one-step enzymatic digestion for preparation of anti-BPDE-dGs [25]. However, only two optically active anti-BPDE-dGMPs were observed in our work (unpublished data).…”

Use of DNA adducts to identify human health risk from exposure to hazardous environmental pollutants: The increasing role of mass spectrometry in assessing biologically effective doses of genotoxic carcinogens

“…These compounds include N-alkylnitrosoureas, dialkylsulfates, alkyl methanesulfonates, cyclophosphamide and cyanoethylene oxide. [73]. Further development of such approaches to make them applicable for human monitoring seems desirable.…”