Evaluation of individual Benzo(a)Pyrene Metabolites as environmental exposure markers for Polycyclic Aromatic Hydrocarbons (PAHs) in marine fish using areolated grouper (Epinephelus areolatus) as a model

Abstract

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous contaminants in the environment. Their contamination of the coastal aquatic environment is of concern because many of these compounds are mutagenic and carcinogenic. Sources of PAHs to the coastal aquatic ecosystems are anthropogenic activities, oil spills and atmospheric fallout of airborne particles. Although they are hydrophobic and have strong binding affinity to particulate matters, they are also known to be readily bioavailable to aquatic organisms. They can be easily absorbed by marine fish and increase their risk of tumors development. Within this class of pollutants, benzo[a]pyrene (BaP) is the most carcinogenic compound. Numerous literatures have reviewed its harmfulness on marine mammals. On the other hand, enzymatic systems in higher marine vertebrates can bio-transform PAHs and thus only small amount of original BaP could be found even in highly exposed marine fish. Therefore, instead of the parent PAH compounds, their bio-transformed metabolites may be better environmental exposure markers to reflect their contamination and health risk to marine fish. In this study, a novel high performance liquid chromatography–programmed wavelength fluorescent detection method was developed to quantify the concentration of a series of BaP metabolites in different tissues of marine fish in a single sample injection. The selected metabolites can be separated by using isocratic elution and detected by using sensitive fluorescence detection by a carefully optimized excitation – emission wavelength switching programme using a new programmable wavelength HPLC fluorescent detector. Method detection limits of 0.7 – 1.6 g kg-1 in muscle and liver, and 1.1 - 5.0 g L-1 in bile were achieved with the various BaP metabolites. Laboratory study on the metabolic fate of BaP after oral administration of BaP-injected pellets was carried out on areolated groupers (Epinephelus areolatus). The model fish were subjected to a series of different BaP exposure scenarios, e.g. force-fed with BaP, force-fed with BaP and kept under hypoxic conditions, kept under hypoxic conditions without exposure to BaP, and control. At days 3, 7, 14, 21 and 28, fish were dissected and liver and muscle were collected for analysis. Two metabolites were found in both tissues. In general, higher amount of metabolites were found in fish tissues which were exposed to BaP and kept under hypoxic conditions. Besides the oral administration route of exposure, intraperitoneal injection of BaP to fish was adopted as another route of intoxication. Areolated groupers (Epinephelus areolatus) were divided into five groups. Three groups were subjected with BaP of different concentrations (6, 12.5, and 25 mg-BaP/kg-fish weight), one group with canola oil and the rest was acted as control. At fixed time intervals, fish were sacrificed and their liver, muscle and bile were collected for qualitative and quantitative analysis of BaP metabolites. The distribution patterns of various BaP metabolites were studied and their correlations with the dosage of BaP that they have been exposed to were discussed.

Date of Award	2 Oct 2007
Original language	English
Awarding Institution	City University of Hong Kong
Supervisor	Hon Wah Michael LAM (Supervisor)