Determination of OH-PCBs and OH.PBDEs in biological matrices : Method development, identification and quantification

Abstract

In the present study, an analytical method for determination of hydroxylated polychlorinated biphenyls (OH-PCBs) and polybrominated diphenyl ethers (OH-PBDEs) and a selection of PCBs, PBDEs and pesticides in biological matrices has been developed. The method is a modified version of the method used for determination of PCBs, pesticides and brominated flame retardants at the Laboratory of Environmental Toxicology (MT-lab) at the Norwegian School of Veterinary Science (NVH).

Blood samples from sheep and seals and liver samples from chicken, cattle and seals were used for the method development. After acidifying with 1 M H2SO4, extraction with cyclohexane and acetone was performed twice. The combined organic phases were evaporated and the lipid content determined gravimetrically. The residue was redissolved in cyclohexane and concentrated H2SO4 was added to remove lipids. Extraction with 1 M KOH in 50% ethanol was performed twice to separate the neutral and phenolic analytes. The organic phase containing neutral compounds such as PCBs and PBDEs, was analysed by GCECD (PCBs and pesticides) and GC-ECNI-MS (PBDEs). The alkaline phase containing the phenolic analytes was acidified and re-extracted with cyclohexane. Derivatization was performed with acetic anhydride:pyridine (1:1) to yield the acetylated analytes, which were subsequently determined using GC-ECNI-MS.

The method was validated for OH-PCBs and one OH-PBDE by spiking sheep blood (~5 g) at five different levels in the range 0.025 - 0.5 ng/g blood, and cattle liver (~4 g) at two different levels; 0.063 - 0.625 ng/g liver. The accuracy given as recovery relative to the internal standard 4 -OH-[13C12]CB159 was in the range 28-128% for blood, with the lowest result being assigned to 4-OH-CB187, which generally gave low recoveries. The other OH-PCBs were within the range 71-128%, and the variation coefficients were in the range 0.9-53%. The recoveries of OH-PCBs and 6-OH-BDE47 from liver samples were in the range 47-123%, with variation coefficients in the range 0.3-6.5%. Estimated instrumental limits of detection for OH-PCBs and the OH-PBDE were in the range 0.01-0.2 ng/ml (0.02-0.4 pg injected). The method limits of detection were in the range 0.001-0.036 ng/g blood and 0.001-0.031 ng/g liver, respectively. The method linearity was in the range 0.9962-0.9992 for the validated concentration range.

Validation results for PBDEs, PCBs, DDT and its metabolites, HCB and Mirex are also included in the thesis. All of these gave validation results within the requirements in the accredited quality system at the MT-lab.

The validated method has been used to investigate levels of OH-PCBs, 6-OH-BDE47 and PBDEs in plasma samples of female polar bears (n = 10) from Svalbard sampled in 1996/1997, and liver samples of male harbour seals (n = 5) from the Norwegian Coast sampled in 2002. Levels of the two most abundant congeners, 4-OH-CB187 and 4-OHCB146, in polar bear plasma were in the ranges 56.7-178 ng/g (recovery-corrected) and 28.1- 106 ng/g, respectively. The most abundant PBDE congener was BDE-47 which was present in the range 0.101-0.357 ng/g polar bear plasma. Levels of the OH-PCBs were considerably lower in the harbour seal liver samples. 4-OH-CB187 ranged from 0.088-0.159 ng/g liver, while the BDE-47 was present in the range 1.34-5.36 ng/g.