Analytical strategies for successful enantioselective separation of atropisomeric polybrominated biphenyls 132 and 149 in environmental samples.

Some of the polybrominated biphenyls (PBBs) found in the environment are axially chiral, due to hindered rotation about the interannular phenyl-phenyl bond. This applies for PBB congeners having two or more bromine substituents in ortho-position to this bond. In this study analytical methods were developed that allow determining the enantiomer fraction (EF) of axially chiral (atropisomeric) PBBs in environmental samples. A white-tailed sea eagle egg was used as test sample. The egg extract was purified and further fractionated by normal phase (NP) high performance liquid chromatography (HPLC), yielding enriched fractions of axially chiral PBB 132 and PBB 149. Gas chromatographic (GC) enantioseparation of the atropisomers of PBB 149 was achieved on one of nine tested modified cyclodextrin phases. Due to coelution with an unknown brominated compound, conventional GC/ECNI-MS, which is based on the detection of the bromide ion, did not allow for the establishment of the EF. However, by means of GC/EI-MS-MS it was possible to verify an EF of 0.42-0.43, i.e. a significant enantiomeric enrichment of the second eluting atropisomer of PBB 149 in the white-tailed sea eagle egg. This is the first proof of non-racemic proportions of a chiral PBB in environmental samples. Despite the testing of nine different chiral stationary phases, GC enantioseparation of PBB 132 or other atropisomeric PBB congeners failed. For this reason, an enantioselective reversed-phase HPLC method was developed. This method proved to be a powerful tool for the separation of PBB atropisomers. It was found that even a standard of the di-ortho substituted PBB 153 could be partially separated into atropisomers at 0 degrees C but already enantiomerized at 5 degrees C. For establishing the EF of PBB 132 in the bird egg sample a combination of enantioselective HPLC followed by non-chiral gas chromatography was employed. Using enantioselective HPLC, the atropisomers of PBB 132 were quantitatively targeted into two separate fractions at room temperature (20 degrees C). After addition of internal standards for volume adjustment the relative amounts of the atropisomers in the isolated fractions were quantified by using non-chiral GC/EI-MS analysis. A deviation from the racemic mixture of the atropisomers of PBB 132 in the egg extract could not be statistically proven.