A two-dimensional HPLC separation for the enantioselective determination of hexabromocyclododecane (HBCD) isomers in biota samples.

A new method for enantioselective analysis of isomers of hexabromocyclododecane (HBCD) is described, using a two-dimensional high-performance liquid chromatography (HPLC) approach to avoid coelution, in particular between (+) α-HBCD, (+) β-HBCD, or (+) γ-HBCD. After isomer separation on a conventional column, the single isomers are transferred to an enantioselective HPLC column using heart cuts. Two enantioseparations are conducted in two separate partial chromatograms: one for α-HBCD and one for β- and γ-HBCD. The result is a completely undisturbed enantioselective separation for α-HBCD at a resolution of 4.11. A peak capacity of 107 was achieved. This peak capacity is utilized by the six peaks of the three isomers with two enantiomers each by 6%. This method was applied to samples of sand eel oil, glaucous gull, and ringed seal. The calibration was performed by treating each enantiomer as a single analyte using a multilevel internal standard calibration. Enantiomeric fractions of 0.495-0.501 with standard deviations (SDs) of 0.056-0.071 were determined for racemic standards of α-HBCD, while the values for fish oil were 0.548-0.562 with SD of 0.018-0.041, depending on the respective mass spectrometric transition.