Benzo{a}pyrene-induced DNA damage in Mytilus galloprovincialis: measurement of bulky DNA adducts and DNA oxidative damage in terms of 8-oxo-7,8-dihydro-2 '-deoxyguanosine formation

Bulky DNA adducts and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) were measured in gill DNA of benzo[a]pyrene (B[a]P)-exposed mussels (50 mg kg(-1 ) dw day(-1)), respectively by the P-32-post-labelling technique and high p erformance liquid chromatography coupled to electrochemical detection assay . A time-course study was performed for both biomarkers and their potential use for marine biomonitoring discussed for the sentinel species studied. I n gills, B[a]P-related DNA adducts were positively correlated with B[ a] P concentration in whole mussel, and were produced in a time-dependent manner relative to exposure. Comparison of adduct levels recorded in this paper i n gills (0.149 +/- 0.079 (standard deviation) to 0.480 +/- 0.139 adduct per 10(8) normal nucleotides) with previous measures carried out in the digest ive gland of the same animals (0.010 +/- 0.005 to 0.251 +/- 0.062 adduct pe r 10(8) normal nucleotides) (Akcha et al. in press) showed higher levels in the former tissue (p < 0. 001). As gills appeared more sensitive to B[a]P effects than the digestive gland, applying the post-labelling technique in gill DNA may allow an earlier detection of pollutant genotoxic effects. 8-O xodGuo formation was assessed as a measure of oxidative DNA damage. No incr ease in the level of 8-oxodGuo by B[a]P exposure was recorded in gill DNA. A methodological study clearly demonstrated the effect of DNA extraction pr ocedure on 8-oxodGuo measurement. No difference in 8-oxodGuo levels was obs erved between the chloroform/isoamyl alcohol and the phenol/chloroform DNA extraction protocols for digestive gland (34.8 +/- 9.3 and 25.6 +/- 4.8 8-o xodGuo per 10(5) dGuo) and gill (6.6 +/- 0.8 and 7.3 +/- 2.4 8-oxodGuo per 10(5) dGuo) tissues (p > 0.05), whereas by the chaotropic method lower 8-ox odGuo levels (0.02 p < 0.05) were measured for both tissue (8.3 +/- 2.0 and 4.8 +/- 1.1 8-oxodGuo per 10(5) dGuo respectively). Contributory factors t o the lack of observed increase in gill 8-oxodGuo level by B[ a] P exposure could be due to the selected way of exposure (via the feed supply) for whi ch gills were not the target tissue of exposure and artifactual DNA oxidati on during sample processing that could have masked the possible B[a]P oxida tive DNA damage.