Pro EZGC Update: Comprehensive 209 compound Library of Brominated Diphenyl Ethers and a New Column Format

While attending Dioxin 2018 in Krakow, I noticed that several academic researchers were studying  the toxicity of specific PBDE congeners not on the standard target compound list for EPA method 1614 (or the EU equivalent). Though PBDE mixtures have been phased out of production and use, the concentrations in the environment have not been declining and are currently still widely monitored. Researchers at Environment Canada demonstrated that although decabromodiphenyl ether (BDE 209) is the primary PBDE in the flame retardant decaBDE, it can be metabolically debrominated by fish (and possibly other animals), forming a variety of penta- and hexa- brominated diphenyl ethers (Stapleton, Alaee et al. 2004).

The standard column used for the analysis of brominated diphenyl ethers by EPA Method 1614 (Figure 1) is the Rtx-1614, a 15 m x 0.25 mm ID x 0.10 µf column with a 5% diphenyl type phase modified for elevated thermal stability. The short column length is critical for EPA method 1614 because BDE 209 is thermally labile; its response relates directly with elution temperature, and because BDE 209 is the primary ingredient in decaBDE, accurate quantitation is critical for determining the scope of contamination.

Figure 1 – Wellington Laboratories BFR-PAR calibration standard collected on a 15 m Rtx-1614. Decabromodiphenyl ether is eluting just after 20 minutes, and the resolution of BDEs 49 and 71 is close, but meets method selectivity criteria.

It is difficult to identify individual PBDE congeners in large homologue groups because their mass spectra are virtually identical (isobaric) and their retention times are similar on the 15 meter column (Figure 2).

Figure 2 – (Top) The elution profile of the 42 hexabromodiphenyl ether homologues collected on a 15m Rtx-1614 column installed in an Agilent GC-MS equipped with an HES. (Bottom) The elution profile of the 42 hexabromodiphenyl ether homologues modeled in ProEZGC

In an attempt to help improve the speed and accuracy of congener identification, especially in the tri- through hepta- brominated homologue groups, we are offering a new high efficiency Rtx-1614 column format (60 m x 0.25 mm x 0.1 μm) as a custom column (PN CC1915) and expanded our ProEZGC PBDE library include all 209 congeners (plus 16 other significant BFRs). A quick literature search leads me to believe I’ve generated the first comprehensive PBDE retention index library. With better separation over a larger time scale, it will be easier to identify individual congeners by using the relative retention time calculated from the EZGC model and select carbon 13 labeled isomers as internal standards.

Figure 3 – (Top) Elution profile of the 42 hexabromodiphenyl ether homologues collected on a 60m Rtx-1614 installed in a Thermo TSQ 9000. The filter used to collect the native hexa-brominated species (black trace) was 641.5 to 483.7. The C13 labeled relative retention time compounds (blue trace) used a mass filter of 653.6 to 493.8. (Bottom) Elution profile of the 42 hexabromodiphenyl ether homologues modeled on a 60m Rtx-1614 using online ProEZGC. The red boxes show an area where there was some disagreement between the real world and modeled chromatogram – the purple box shows what appears to be a gap in the MRM data and possibly missing peaks

It should be noted that model accuracy is highly dependent on an accurate column length and flow. Due to the internal diameter variation inherent to fused silica capillary columns, it is essential that the effective column length be calculated using the column holdup time and head pressure. I translated the 15m run conditions to the 60m column, but my retention times did not match with the model as well as the 15m runs did. These changes in elution temperatures could explain the minor elution profile differences seen between the TSQ 9000 run and the online ProEZGC model (Figure 3) in the region enclosed by the red box. It is also possible that some of the isomers have a much reduced response because I’m only looking at one transition, and different substitution patterns can yield different fragmentation patterns (missing peaks in the purple box). The SIM chromatogram shown for the 15m column data is a sum of multiple ions, but primarily m/z = 483.6.

Finally, the 60m Rtx-1614 is not appropriate for quantitative analysis of the octa-, nona-, or decabrominated diphenyl ethers because they can experience extensive thermal degradation at elevated elution temperatures and extended time on column.

References

Stapleton, H. M., et al. (2004). “Debromination of the Flame Retardant Decabromodiphenyl Ether by Juvenile Carp (Cyprinus carpio) following Dietary Exposure.” Environmental Science & Technology 38(1): 112-119.

 

 

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