Polychlorinated Biphenyls in Brackish Water Fish in the River Niger, Nigeria

Background. Anthropogenic polychlorinated biphenyls (PCBs) in aquatic environments poses human and ecological health risks in Nigeria. Objectives. This study determined the concentrations of PCBs in brackish water fish in the River Niger to assess the contamination status of fish consumed by the local population. Methods. The sampled fish species included Drepane africana, Mochokus niloticus, Chrysichthys nigrodigitatus, Pristipoma jubelini, Vomer septapinis, Pseudotolithus senegalensis, Mugil cephalus, Pseudotolithus elongatus, Sphyraena piscatorum and Lutjanus goreensis, purchased from landing sites. Six fish from each species were sampled, for a total of 60 samples. Twenty-seven (27) PCB congeners, #8, #18 #28, #44, #52, #60, #77, #81, #101, #105, #114, #118, #123, #126, #128, #138, #153, #156, #157, #167, #169, #170, #180, #185, #189, #195, and #206 were screened in the fish samples using standard methods. The PCBs were identified and quantified using gas chromatography (GC) (Hewlett Packard GC 5890 series 11 with electron capture detector). Confirmation was performed using Shimadzu GCMS QP2010. Results. The sum of the National Oceanic and Atmospheric Administration Agency (ΣNOAA) PCBs occurred at the highest concentration of 1830.0±484.0 μg/kg detected in Vomer septapinis, and the lowest in Pseudotolithus senegalensis, with a mean concentration of 795±169.3 μg/kg. The concentration of dioxin-like (DL) PCBs was highest in Pristipoma jubelini (992.0±88.6 μg/kg) and lowest (285.6±81.5 μg/kg) in Drepane africana. The highest mean concentration (418.±177.6 μg/kg) of International Council for the Exploration of the Seas-7 (ICES-7) PCBs was observed in Vomer septapinis. The heavier ICES-7 congeners PCB-138, PCB-153, and PCB-180 occurred at higher concentrations compared to the lighter molecular weight ICES-7: PCB-28, PCB-52, and PCB-101. The European Union (EU) marker PCB limit of 335 μg/kg was exceeded in all the brackish water fishes with the exception of Mochokus niloticus, Pristipoma jubelini and Pseudotolithus senegalensis. Discussion. The total level of PCBs in the brackish fish samples was relatively high at >1000 μg/kg (above the World Health Organization (WHO) and Food and Agriculture Organization of the United Nations (FAO) guideline of 1000 μg/kg fresh weight). The EU guideline value for fish (220 μg/kg fresh weight) was exceeded in about 80% of the brackish fish samples studied. Conclusions. Consumption of fish from the River Niger may expose humans to polychlorinated biphenyls. In addition, since contamination of the fish samples is an indication of river contamination, river water quality is of great concern and there is a need for additional PCB data on water quality to be distributed to the community, followed by mitigation measures. Competing interests. The authors declare no competing financial interests.


Introduction
Improper waste management and pollution are environmental problems confronting developing countries such as Nigeria. The effects of pollution are seen across the various components of the environment. 1, 2 Contamination of aquatic products and food items with organic pollutants, especially organochlorine pesticides, have been reported. 3 Other organic pollutants which require assessment are polychlorinated biphenyls (PCBs) due to their historically intensive use in the region.
Manufactured in different countries under various trade names (e.g., Aroclor, Clophen, Phenoclo), PCBs were introduced in 1929. These groups of organic pollutants are chemically stable and heat resistant, and were used worldwide as transformer and Research mono, di, tri, tetra, penta, hexa, hepta, octa, nona, and deca. The different homologues have 1-46 PCB isomers with similar molecular weight. The sequence in isomeric forms are penta-46, tetra, hexa-42 each, tri, hepta-24 each, di, octa-12 each, mono, nona-3 each and deca-1, for a total of 209 isomers. 5 Out of the 209 possible PCB congeners, only about 130 are likely to occur in commercial products. However, these chemicals are being phased out of use in products because of their health effects and persistence in the environment.
The persistence of PCBs may be understood by the mode and mechanism of their degradation. Biodegradation of PCB congeners occurs by dechlorination by aerobic and anaerobic bacteria. 5 Generally, these are slow processes and hence contribute to the persistence of PCBs. The low congeners of mono, di, and tri PCBs generally biodegrade aerobically, while the heavier congeners (tetra, penta, hexa PCBs) generally biodegrade anaerobically. Biodegradation also depends on temperature and PCB concentration.
PCBs have been classified as substances for which there is evidence of endocrine disruption. 6,7 There is, therefore, the need to monitor the presence and levels of these chemicals in the environment, particularly in aquatic products through which humans may be exposed.
High levels of PCBs in the water of the River Niger have recently been reported, along with other persistent organic pollutants (POPs) such as organochlorine pesticides. 8, 9 There is a need to determine if PCBs are bioaccumulated in the fish species of the River Niger. This study, therefore, aims to determine PCB levels in the brackish water fish of the River Niger.

Methods
Appropriate locations for the collection of fish samples were selected, and sample types and species and sample numbers for each location were identified. Samples were collected during the fall when reproduction was deemed to be complete and a full dry season of exposure to potential toxins had occurred. The target species of fish in this project were representative and economically important species for the River Niger.
The sampled fish were identified as either demersal, benthic pelagic or pelagic. Sampled  Fish samples were purchased from landing sites. The method of fishing in those areas included netting, trotlines, angling, and traps. The fish were thoroughly washed with distilled water, sorted and identified. They were placed in food grade coolers, covered with ice blocks and as quickly as possible shipped to the laboratory for analysis where they were deep frozen immediately on arrival in the laboratory. In the laboratory, basic biogenic data were generated, including total length (measured to the nearest tenth of a centimeter) using a metric measuring board, and weighed to the nearest gram using a metric weighing scale. Observations were made on other conditions of the fish, e.g. tumors, lesions, collection problems, weather conditions, etc. The knife used for filleting was washed with soapy water and rinsed with analytical grade acetone (both sides of the knife blade were decontaminated), air-dried and wrapped in aluminum foil and kept inside zip closure plastic bags. The knife was decontaminated between samples.
Latex gloves were worn during the handling of the fish for fillet processing. Fish were rinsed with distilled water before scaling and filleting. The samples were carefully scaled on a fillet board lined with heavy duty aluminum foil. Both sides of the fish were filleted, including the belly portion. After filleting the fish, the fillet was wrapped in restaurant grade aluminum foil with the dull side in contact with the skin and placed in a zip closure plastic bag with the sample label dropped inside. The skin-off fillet was collected for

Extraction
The extraction columns (330 mm x 23 mm, each fitted with removable Teflon stopcock) were prepared by inserting a small glass wool (Corning Pyrex) plug into the bottom of each chromatographic column and then the column was rinsed twice with 15 mL of petroleum ether. Air was removed from the glass wool by lightly tapping it with a clean glass stirring rod. A Zymark concentration tube, 200 mL with 1 mL endpoint, was placed under each column and the appropriate labels were transferred to these tubes. The sample mixture was then poured into the column, after which 50 mL of acetone/petroleum ether was added to the sample beaker, stirred and transferred to the column. The solvent was allowed to pass through the column, but as it began to elute into the concentration tube, the stopcock was closed. At this point, the column was lightly stirred with a glass rod to remove trapped air. Elution was then continued at the rate of 1-2 mL per minute until the solvent level reached the beginning of the sample mixture. Another 50 mL of acetone/ petroleum ether was added and elution was continued at the same rate. The columns were allowed to drain completely after the second 50 mL of solvent was added. The stopcocks were rinsed with acetone/petroleum ether to wash any residue lipids or analytes into the concentration tube.

Extract Concentration
The eluent was concentrated by placing the concentrator tubes in a Kuderna-Danish TurboVap concentrator. The concentrator water bath was kept at 40 o C and the argon sweep gas (purified grade) pressure was set at 10-12 psi with the TurboVap control knobs. The sample was concentrated to 1 mL and solvent exchanged to isooctane. The extract was transferred to a 2-ml graduated vial with isooctane.

Lipid Determination
The method of lipid analysis defined by Sloan et al. was used. 11 Prior to clean up by column chromatography, a volume of sample extract equivalent to 1 g of tissue was pipetted into a pre-weighed (after acetone rinsing and drying) aluminum drying pan. Preweighing was performed to the nearest 0.1 mg. The extract was allowed to evaporate under static conditions in a fume hood for 2 hours. The pan was weighed again to the nearest 0.1 mg and the percent extractable lipid was computed as 100 x (1-weight of residual lipid).

Sample Cleanup
A 600-mm x 19 mm cleanup column was prepared by blocking the hole with glass wool and adding 3 g of activated silica gel (60 to 100 mesh, calcined at 650ºC for 24 hours in a muffle furnace, and then stored at 130ºC until use). Before use, the silica gel was deactivated with 1 ml distilled water. The column was topped with 1 cm of preheated sodium sulfate previously heated at 650ºC for 8 hours in a furnace, and stored in a clean bottle in a desiccator. The column was analysis in the present study. After filleting, a homogenous tissue sample was created using a commercial-grade food blender washed with phosphatefree soap and water and rinsed with hexane between samples to avoid cross contamination. Each sample was ground three times, with the tissue mixed between grinding to ensure a homogenous sample. Two packets of tissue were prepared and wrapped in aluminum foil, numbered and frozen. One packet was used for PCBs analysis and the other for pesticide analysis.

Sample Preparation
Samples were extracted in batches of 6-8 samples including all the necessary quality assurance/quality control (QA/QC) measures specified in the method used for this study. All of the solvents were pesticide quality grade. Fish tissue previously homogenized using a commercial grade food blender (washed with phosphatefree soap and water and rinsed with hexane between samples to avoid cross-contamination) was thawed and Research were expressed in wet weight bases and were not corrected for recoveries.

Analytical Methods
Twenty seven (27)  The model and type of GC used in the confirmation study was a Shimadzu GCMS QP2010 and capillary column type: HP1MS (30m x 0.25um x 0.25mm). The GC was checked to ensure that it was in good condition. Thereafter, it was flushed with the carrier gas. Calibration was done using reference standards; 0.063; 0.125; 0.25; 0.5; 1.0 ppm. These standards were run six times to calculate the mean, range, and standard deviation along with peak column performance, peak height and resolutions. Helium gas was used because of its inertness. The statistical data analysis of the results included t-tests and correlations using the Statistical Package for the Social Sciences (SPSS) software package.

Results
The biometric data are shown in Table 1  The concentrations of marker PCBs are presented in Figure 1. PCB-28, 52, 138, 180, 101, 118 and 153 were detected in all of the analyzed brackish water fish samples.

Marker PCBs
Marker PCBs, sometimes called "Dutch seven" or ICES7, are seven PCBs that have been measured for many years as an indication of the total PCB contamination. They include PCB-28, PCB-52, PCB-101, PCB-118, PCB-138, PCB-153, and PCB-180. One of these seven, PCB118, is classified as a dioxin-like PCB  Table 6 shows the coplanar PCBs that have been assigned toxicity equivalent factors (TEFs) by the WHO.
Abbreviations: ADI, acceptable daily intake; TEQ, toxic equivalent *TEQ in ng (TEQ) kg -1 , ADI in ng (TEQ) kg -1 / body weight (bw)/day. An acceptable daily intake of 10 pg TEQ/kg bw/day can be used to assess the health risks of the intake of mixtures of PCDDs/Fs and PCB congeners. This ADI, which has been adopted by several countries, including Canada and the Netherlands, has been revised downwards by the WHO to a range of 1-4 pg TEQ kg bw/day. The Health Council of the Netherlands has specified a healthbased exposure limit of 1 pg TEQ/ kg bw/day. In addition to these ADIs, some countries have regulations for dioxin-like PCB concentrations or international TEQ in specific food types (summarized in Buckland et al.). 13 In the United States, a limit value has been set for dioxins in fish. This guideline recommends not consuming fish with dioxin levels greater than 25 ng TEQ kg-1 on a wet weight basis (Food and Drug Administration, cited in the US Environmental Protection Agency (USEPA), 1987). In Ontario, there is a guideline value for sport fish of 15 ng TEQ kg-1. The regulation of dioxins and furans for the protection of wildlife is complicated by the bioaccumulative nature of these compounds. To address this issue, the Canadian Council of Ministers of the Environment has proposed both a tissue residue guideline (50 ng I-TEQ kg-1 fat) and a dietary intake guideline (1.1 ng I-TEQ kg-1 fresh weight) for the protection of aquatic wildlife. 26 The TEQ values in brackish water fish from this study exceeded the regulatory limits in all of the fish samples and the ADI values also exceeded the regulatory limits in all of the samples, with the exception of 0.61 ng I-TEQ kg-1 and 0.71 ng I-TEQ kg-1detected in Pristipoma jubelini and Pseudotolitus senegalensis, respectively.

Conclusions
The total PCB levels in the brackish fish samples were relatively high at >1000 µg/kg, above the WHO/FAO guideline of 1000 µg/kg fresh weight. The EU guideline value for fish of 220 µg/kg fresh weight was exceeded in about 80% of the brackish water fish samples. Consumption of fish from the River Niger may expose humans to polychlorinated biphenyls. In addition, since contamination of the fish samples was an indication of river contamination, water quality is of great concern and there is a need for additional PCB data on water quality to be distributed to the community, followed by mitigation measures.

Copyright Policy
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