Search

The possibility of using the pyrene metabolite 1-hydroxypyrene as a biomarker of polycyclic aromatic hydrocarbons (PAHs) exposure was investigated by exposure of the marine polychaete Nereis diversicolor to several PAHs in the laboratory. Animals were exposed to pyrene alone and to five different PAHs - phenanthrene, anthracene, pyrene, benzo[a]pyrene and benzo[k]flouranthene. After five days of exposure the concentrations of parent PAHs and 1-hydroxypyrene were identified using three different analytical methods, high-performance liquid chromatography with fluorescence detection (HPLC/F), synchronous fluorescence spectroscopy (SFS) and gas chromatography with mass spectrometric detection (GC/MS). The SFS measurements of 1-hydroxypyrene were validated by the more sensitive method HPLC/F. The positive correlation between total PAHs and 1-hydroxypyrene concentrations in the polychaete tissues observed in experiments, suggests the feasibility of 1-hydroxypyrene as a suitable biomarker for total PAH exposure assessment. Furthermore, the possibility of employment of the simple and rapid SFS method instead of HPLC/F for biomarker analysis has been confirmed by the positive and significant correlation between results achieved by these two analytical methods.

We compared microbial mineralization of [4,5,9,10-14C]pyrene and its eukaryotic [4,5,9,10-14C]pyrene metabolites in estuarine sediments. Metabolites were obtained by exposing the estuarine deposit-feeding polychaete Nereis diversicolor to sediment-associated 14C-pyrene, followed by homogenization of the worms and extraction of the pyrene-metabolites. In sediment from a pristine Danish Fjord only 2.6% of the added metabolite-label and 1.7% of the pyrene-label were mineralized to 14CO2 during 175 days incubation. Pre-exposure of the pristine sediment to unlabelled pyrene for 60 days increased the mineralization potential for 14C-pyrene substantially, as 81.2% was mineralized to 14CO2 during 95 days incubation, whereas 14C-pyrene metabolite label was unaffected by pre-exposure to pyrene. In comparison, naturally aged bunker-oil contaminated sediment did not show elevated potentials for mineralization of neither 14C-pyrene nor 14C-metabolites. Six bacterial strains of known pyrene degraders were tested for growth on crystalline 1-hydroxypyrene. 1-Hydroxypyrene is the only intermediate eucaryotic metabolite of pyrene. The results indicate that 1-hydroxypyrene was not utilized as a sole source of carbon and energy by any of them. In addition, respiration was depressed in all six strains when exposed to crystalline 1-Hydroxypyrene, demonstrating an acute toxic effect of 1-hydroxypyrene. The results presented here suggest that microbial degradation of pyrene is not enhanced by release of aqueous and polar metabolites by marine invertebrates.

The uptake of polycyclic aromatic hydrocarbons (PAHs) by marine deposit-feeding invertebrates can be determined by screening for PAH-derived metabolites. We identified 1-hydroxypyrene as the only intermediate metabolite in tissue of four species of deposit-feeding polychaetes, Nereis diversicolor, Nereis virens, Arenicola marina, and Capitella sp. I exposed to pyrene spiked sediment. Synchronous fluorescence spectroscopy (SFS) provides a fast and simple method for both qualitative and quantitative analysis of 1-hydroxypyrene in all four species. The SFS assay was validated using HPLC with ultraviolet detection. A good correlation between 1-hydroxypyrene concentrations determined by the two methods was observed. We used HPLC with fluorescence detection combined with enzymatic hydrolysis of conjugated metabolites to investigate species specific metabolite patterns. A tentative aqueous metabolite identification scheme indicates that Nereid polychaetes predominately make use of glucuronide conjugation whereas Capitella sp. I. and Arenicola marina appear to utilize predominantly sulfate and/or glucoside conjugation. The usefulness of 1-hydroxypyrene as a biomarker for PAH exposure in deposit-feeding invertebrates is discussed.

Both 1-hydroxypyrene and 1-hydroxypyrene glucuronide are identified as the primary phase I and phase II metabolites of the four-ringed polycyclic aromatic hydrocarbon (PAH) pyrene in the marine deposit-feeding polychaete Nereis diversicolor. Identification of pyrene and primary metabolites was performed using high-pressure liquid chromatography (HPLC) with diode-array detection and fluorescence detection (HPLC/DAD/F) and an ion-trap mass spectrometer for positive identification of 1-hydroxypyrene glucuronide. Besides 1-hydroxypyrene and 1-hydroxypyrene glucuronide, the HPLC/F trace of tissue samples from pyrene-exposed worms showed three additional low-intensity peaks that may be related to pyrene metabolism based on similar excitation/emission wavelengths. The peaks were all too low in intensity to be positively identified. Of the total PAH in tissue, 1-hydroxypyrene glucuronide, 1-hydroxypyrene, and pyrene constituted 73%, 2%, and 25% respectively. Gut elimination of metabolic products is supported by the identification of 1-hydroxypyrene and 1-hydroxypyrene glucuronide in both gut fluid and defecation water. Being the only phase I metabolite of pyrene, 1-hydroxypyrene becomes a useful marker for PAH exposure, and it may serve as a valuable model compound for assessing species-specific PAH metabolic capabilities.

1-Hydroxypyrene glucuronide is identified as the single major aqueous metabolite of the tetracyclic aromatic hydrocarbon pyrene, in tissue from a deposit-feeding marine polychaete, Nereis diversicolor. Identification was performed using an ion trap mass spectrometer fitted with an atmospheric pressure chemical ionization (APCI) probe and connected to a high-performance liquid chromatography/diode array detector (HPLC/DAD) system. Besides 1-hydroxypyrene, the 339-nm UV trace of tissue samples from pyrene-exposed worms showed only one dominant peak that could be related to pyrene metabolism. Negative APCI-MS of this supposed 1- hydroxypyrene conjugate gave a characteristic signal at m/z 429 corresponding to the molecular ion of 1-hydroxypyrene glucuronide plus eluent adducts ([M - H + 2H(2)O](-)). Fragmentation pathways were studied by isolating the abundant ion at m/z 429 in the ion trap and performing multiple mass spectrometric experiments (MS(n)). The fragmentations observed were consistent with the proposed identification. Two low intensity LC peaks that could be related to pyrene metabolism by their DAD absorption spectra were also present in the 339-nm UV chromatogram of tissue samples. However, these peaks could not be identified by their mass spectra in negative ion mode due to ion suppression by very abundant co-eluting impurities. The present method shows that LC/MS(n) is a fast and useful analytical tool for identification of aqueous polycyclic aromatic hydrocarbon biotransformation products in samples from relatively small marine invertebrates with limited sample preparation., (Copyright 2002 John Wiley & Sons, Ltd.)