Fig. 1. Structures of (±)-anti-BPDE enantiomer (top) and the moieties of four BPDE–N²–dG stereoisomers in oligonucleotides (bottom).

Fig. 2. Chromatograms obtained from HPLC analysis of the reaction products of the (±)-anti-BPDE and 16mer using a Capcell Pak C18 MG column (column 2). The analytes were simultaneously monitored by DAD (200–400 nm) and fluorescence (Ex 343 nm, Em 400 nm). The eluted species including unmodified DNA (8.996 min) and four stereoisomers of anti-BPDE–16mer (20.820, 22.200, 24.960, and 26.327 min). Peaks 1, 2, 3, and 4 represent cis-anti-BPDE–N²–dG-16mer A, trans-anti-BPDE–N²–dG-16mer A, trans-anti-BPDE–N²–dG-16mer B, and cis-anti-BPDE–N²–dG-16mer B. For the conditions of HPLC analysis see Section 2.

Fig. 3. HPLC analysis of the reaction products of the (±)-anti-BPDE and unmodified 16mer oligonucleotide using the uncoated ODS column (column 1). The analytes were monitored by DAD and fluorescence detection as described in Fig. 2. Peaks 1 and 2 are predominantly trans-anti-BPDE–N²–dG-16mer A and trans-anti-BPDE–N²–dG-16mer B, respectively. For the HPLC conditions see Section 2.

Fig. 4. HPLC analysis of fraction 1 (left) and fraction 2 (right) using Capcell Pak C18 MG column (column 2). The fractions 1 and 2 were collected from HPLC preparation using the uncoated ODS column (column 1) as shown in Fig. 3. For fraction 1, an isocratic elution with a mobile phase of 25.7% solvent B and 74.3% solvent A was used for the first 5 min; then a gradient elution of solvent B increased from 25.7% to 35.1% for the next 20 min. For fraction 2, an isocratic elution with a mobile phase of 27% solvent B and 73% solvent A was used for the first 5 min; then a gradient elution of solvent B increased from 27% to 35.1% for the next 25 min. Under such conditions, the fraction 1 can be separated into two peaks (14.52 and 16.37 min), and fraction 2 was separated into two peaks (17.12 and 19.70 min). The analytes were monitored by both DAD (200–400 nm) and fluorescence detection (Ex: 343 nm; Em: 400 nm). For other HPLC conditions see Section 2.

Fig. 5. HPLC analysis of the reaction products of the (±)-anti-BPDE and unmodified 16mer oligonucleotide using a Capcell Pak C18 SG column (300 Å pore size) showing peak tailing of DNA and DNA adducts. The analytes were monitored by DAD (200–400 nm) and fluorescence detection. For the HPLC conditions see Section 2.

Fig. 6. Chromatograms obtained from HPLC analysis of the reaction products of the (±)-anti-BPDE and 16mer using a Capcell Pak Ph UG column (column 4). The analytes were simultaneously monitored by DAD (200–400 nm) and fluorescence (Ex: 343 nm; Em: 400 nm). Eluted species include unmodified DNA (6.51 min) and four stereoisomers of anti-BPDE–16mer (17.21, 18.11, 20.40, and 21.40 min). Peaks 1, 2, 3, and 4 are cis-anti-BPDE–N²–dG-16mer A, trans-anti-BPDE–N²–dG-16mer A, trans-anti-BPDE–N²–dG-16mer B, and cis-anti-BPDE–N²–dG-16mer B, respectively. For the conditions for HPLC analysis see Section 2.

Fig. 7. HPLC preparation of four adducts using an analytical Capcell Pak C18 column (column 2, A) and an analytical Capcell Pak Ph group column (column 4, B).

Fig. 8. HPLC analysis of the three single adducts purified by the Capcell Pak C18 column (column 2, top) and the corresponding the absorption spectra (bottom). Peaks 2, 3, and 4 are corresponding to trans-anti-BPDE–16mer A, trans-anti-BPDE–16mer B, and cis-anti-BPDE–16mer B, respectively.