Monitoring contaminants from oil production at sea by measuring gill EROD activity in Atlantic cod (Gadus morhua)

An ex vivo gill EROD assay was applied in Atlantic cod (Gadus morhua) as a biomarker for waterborne CYP1A-inducing compounds derived from oil production at sea. Exposure to nominal concentrations of 1 ppm or 10 ppm North Sea crude oil in a static water system for 24 h caused a concentration-dependent gill EROD induction. Further, exposure of cod for 14 days to environmentally relevant concentrations of produced water (PW, diluted 1:200 or 1:1000) from a platform in the North Sea using a flow-through system resulted in a concentration-dependent induction of gill EROD. Crude oil (0.2 ppm) from the same oil field also proved to induce EROD. Finally, gill EROD activity in cod caged for 6 weeks at 500-10 000 m from two platforms outside Norway was measured. The activities in these fish were very low and did not differ from those in fish caged at reference sites.     

Quantitative determination of de-conjugated chrysene metabolites in fish bile by HPLC-fluorescence and GC--MS

Two analytical methods have been evaluated for quantitative determination of de-conjugated chrysene metabolites in fish bile. High performance liquid chromatography-fluorescence (HPLC-F) and gas chromatography-mass spectrometry (GC--MS) were compared regarding instrumental and overall limits of detection (LOD) as well as recoveries for the following nine chrysene compounds: 1-, 2,- 3-, 4- and 6-hydroxychrysene (1-, 2-, 3-, 4- and 6-OH-chr), 1,2-dihydroxy-1,2-dihydrochrysene (1,2-DHD-chr), 3,4-dihydroxy-3,4-dihydrochrysene (3,4-DHD-chr), 5,6-dihydroxy-5,6-dihydrochrysene (5,6-DHD-chr) and chrysene. Instrumental LODs were comparable for the two methods whereas the overall LOD was better for HPLC-F. Recoveries varied per chrysene compound for both HPLC-F (62-107%) and GC-MS (48-124%). In vivo formed chrysene metabolites were studied in the bile of Atlantic cod (Gadus morhua) exposed to chrysene (1 mg/kg) via intra-peritoneal (i.p.) and inter-muscular (i.m.) injection. Total amounts of chrysene metabolites were three times higher in i.p. compared to i.m. exposed cod bile, but the relative distribution of determined metabolites was very similar. 1,2-DHD-chr was the most prominent metabolite in de-conjugated bile and constituted more than 88% of the total chrysene metabolites. Additional chrysene metabolites formed were 3,4-DHD-chr and 1-, 2-, 3- and 4-OH-chr. K-region chrysene metabolites (oxidation at carbons 5 and 6) were not detected and seem to be a less favoured biotransformation route. The two methods were applied and evaluated for analysis of chrysene metabolites in two bile reference materials (BCR 720 and 721) and a limited number of field exposed cods.     

An evaluation of two fluorescence screening methods for the determination of chrysene metabolites in fish bile

Two screening methods, synchronous fluorescence spectrometry (SFS) and high performance liquid chromatography with fluorescence detection (HPLC-F), have been evaluated for their suitability in determining chrysene metabolites in fish bile. The optimal wavelength pair, excitation/emission 272/374 nm, for SFS measurements of chrysene metabolites was identified by analysis of bile taken from fish exposed to the pure compound. This analysis revealed in addition some information about the metabolite pattern. However, when bile from fish exposed to complex and environmentally relevant mixtures of polycyclic aromatic hydrocarbons (PAHs) was analysed using these methods, identification of the chrysene metabolites was poor. Analysis of bile taken from fish exposed to single PAHs identified other three- and four-ring aromatics as the main interfering compounds. Both methods were equally able to discriminate between impacted and reference sites by determination of relative concentrations of fluorescent three- and four-ring aromatics.