Study On The Hydroxylated Metabolitic Pathway Of Polychlorinated Biphenyls In Orgnisms And Exploration In Toxicity Of Metabolites

Polychlorinated biphenyls (PCBs) are ubiquitous and typical persistent organic pollutants(POPs) in environment with accumulation,persistence and toxicity. Its hydroxylated metabolites have become another new focus in environment and food safety field for their accumulation and potential endocrine disruption effect. GC/MS and GC/MS/MS methods were established to detect hydroxylated-PCBs in aquatic product. And in order to reveal the metabolic pathway of PCB101 in mouse, the method to detect hydroxylated-PCBs in mouse tissues was established using gas chromatography with mass spectrometric detector. In the present study, we studied metabolitic pathway of PCB101 in mouse and eel. A human breast cancer cell line(MCF-7) was used to determine cell proliferation effects of PCB101 and its hydroxylated metabolites.This paper developed the GC/MS and GC/MS/MS methods to determine hydroxy-PCBs in aquatic products. The optimum condition of silylation derivatization was as follows:temperature 60℃, derivatizing time 40min. Before extracting targets compounds, HCl was added to the sample to modify the pH to 3. Sulfuric acid and silica column were used as clean-up procedure to eliminate proteins and lipids. And the recoveries of seven hydroxy-PCBs ranged from 70.6-101.1%, RSD ranged from 4.6-12.0%. The detection limit of seven hydroxy-PCBs determined by GC/MS ranged from 0.32 to 0.63μg/kg, and that of GC/MS/MS ranged from 0.02 to 0.14μg/kg. Mass spectrometric detector is much more selective and can increase the accuracy in identifying the target compounds from the interfering substances.The GC/MS method was developed to determine hydroxy-PCBs in mouse tissues. PCB and hydroxy-PCB were extracted at the same time using hexane/ethyl acetate (v/v,1/1) as extracting solvent. The present study adopted KOH to isolated hydroxy-PCBs from PCBs.The confirmed optimum concentration of KOH was 0.5mol/L. The recovery of PCB in spiked mouse muscle and blood were 82.8% and 85.6% respectively, RSD were 5.2% and 4.7% respectively. The recoveries of hydroxy-PCBs in spiked muscle ranged from 76.1 to 101.0%, RSD ranged from 1.8% to 8.9%; The recoveries of hydroxy-PCBs in spiked blood ranged from 74.3% to 113.2%, RSD ranged from 1.5% to 7.0%. The detection limit of PCB was 0.52μg/kg and that of hydroxy-PCBs were from 0.18 to 0.35μg/kg.This paper studied the accumulation and metabolism of PCB101 in mouse and eel. The concentration of PCB101 accumulated to the highest level in mouse tissues within 7days after intraperitoneal injection at the dose of 5μg/g, and during the second month's exposure, the concentration curve of PCB101 became gentle. This paper confirmed that the main metabolites of PCB101 in mouse were 3-OH-PCB101 and 4'-OH-PCB101, and 3-OH-PCB101 was the most abundant. The metabolic pathway of PCBs proceeded by cytochrome P450 enzyme mediated oxidation system. An OH-group replaced an H-in aromatic ring. And this replacement tended to happened in meta and para position. The other two possible metabolites were presumed to be 3'-OH-PCB and 4-OH-2,2',3,5,5'PCB.To eel, the exposure dose was 1μg/kg, and the PCB101 was exposured by mouth solved in oliver oil. The PCB101 concentration in eel tissues was accumulated to the highest level within 5days after exposure. 3-OH-PCB101 and 4'-OH-PCB101 were not detected in eel tissues during the exposure period.A human breast cancer cell line(MCF-7) was used to stduy the cell proliferation effects of PCB 101 and its two hydroxylated metabolites including 3-OH-PCB101 and 4'-OH-PCB101. Concentration gradients of three compouds were 5×10-6,5×10-7,5×10-8,5×10-9mol/L respectively.Alcohol and estrogen control were designed to ensure the accuracy of research. The results showed that the three compounds had no effect on the MCF-7 cell proliferation.This paper provides technical support for the potential risk of hydroxylated-PCBs to the safety of aquatic products. And it is meaningful to reflect the level of hydroxylated-PCBs and total bioaccumulated PCBs in orgnisms. Meanwhile, this paper provides reference materials for the further related research.