Making sense of the toxin screen

Felipe Reggeti, Nick Schrier, Tracy Van Raaij

Animal Health Laboratory, University of Guelph, Guelph, ON

AHL Newsletter 2021;25(1):22.

What happens when toxicity is suspected, but there is limited or no history of exposure to a poison and the clinical presentation, laboratory data and postmortem findings do not provide any directions?

The AHL toxicology section offers several highly sensitive tests for screening of biological specimens, feed and suspect materials for the presence of toxins.  These techniques include specific tests (e.g. bromethalin, ethylene glycol), as well as general methods for identification of unknown substances.  The most comprehensive of these is the “toxin screen” (toxiscr).  This is a qualitative multi-residue method that combines gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the identification of targeted (known) and non-targeted (unknown) substances.  The readers are referred to the AHL LabNote 59 for a list of compounds that can (and a few that cannot) be identified by this screen (1).  However, the GC-MS component also includes an “open screen” for unknown volatile substances.  These are identified by matching an unknown compound’s spectrum with a mass spectral library that contains approximately 200,000 chemicals (NIST/EPA/NIH Mass Spectral Library - NIST08); additional substances can be subsequently added.  This method is an excellent first option to investigate exposure to unknown substances; however, it cannot identify all possible toxins and it has limitations that are important to understand:

  • The extraction method determines the presence or absence of the substance of interest in the sample. There are many ways to obtain a liquid extract from the sample in order to run it through the instrument.  Depending on physical-chemical structure, the solvents used in a particular method may or may not extract or retain the substance of interest.  Because toxicology labs have their preferred extraction protocols, they may be able to “see” different substances in the same sample.
  • Results are qualitative (adequate to confirm exposure); however, quantification is possible provided a standard (the purified substance) is available.  This would require additional validation and would increase costs.
  • Since extracts from biological samples consist of complex mixtures of molecules, interpretation of chromatograms requires great expertise to separate substances that are normal components of the sample (e.g. fatty acids), or background environmental contaminants from those that are potentially significant.

As a matter of example, here are a few substances that have been identified in our laboratory with the toxin screen and the interpretation provided in the context of the (often times limited) available information:

GC-MS open screen

GC-MS volatile targeted compounds

LC-MS non-volatile targeted compounds