Method for the detection of desmethylbromethalin in animal tissue samples for the determination of bromethalin exposure

Bromethalin, a potent neurotoxin, is widely available for use as a rodenticide. As access to other rodenticides is reduced due to regulatory pressure, the use of bromethalin is likely to increase with a concomitant increase in poisonings in nontarget animals. Analytical methods for the detection of bromethalin residues in animals suspected to have been exposed to this rodenticide are needed to support post-mortem diagnosis of toxicosis. This paper describes a novel method for the analysis of desmethylbromethalin (DMB), bromethalin's toxic metabolite, in tissue samples such as liver, brain, and adipose. Samples were extracted with 5% ethanol in ethyl acetate, and an aliquot of the extract was evaporated dry, reconstituted, and analyzed by reverse phase ultrahigh-performance liquid chromatography-mass spectrometry. The mass spectrometer utilized electrospray ionization in negative ion mode with multiple reaction monitoring. This method was qualitatively validated at a level of 1.0 ng/g in liver tissue. The quantitative potential of the method was also evaluated, and a method detection limit of 0.35 ng/g wet weight was determined in fat tissue. DMB was detected in tissue samples from animals suspected to have been poisoned by this compound. To the authors' knowledge, there have been no other methods reported for analysis of DMB in tissue samples using LC-MS/MS.