Poisoning response is quick, as LC-MS does the trick

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  • Published: Mar 1, 2018
  • Author: Ryan De Vooght-Johnson
  • Channels: HPLC
thumbnail image: Poisoning response is quick, as LC-MS does the trick

Speedy analytical results needed with poisoning cases

Analytical methods can be extremely useful in cases of suspected poisoning by pharmaceuticals, but only if they give results prior to clinical decisions being taken. Such acute poisoning incidents may occur due to attempted suicide, children getting hold of adult medication or the wrong medication being taken by accident. Often the patient is unable to describe exactly what has occurred; indeed, they may have lost consciousness prior to arrival at a medical facility. Rapid analysis of blood samples is needed so that appropriate treatment can be given.

The University of Saarland scientists aimed to produce techniques for detecting a wide range of pharmaceuticals in blood samples. Previous work (by Meyer et al.) described a rapid method of detecting 40 drugs by GC-MS, but additional methods are needed as many pharmaceutical compounds are unsuitable for GC. UHPLC coupled with MS/MS was used as a rapid method for detecting 45 drugs and metabolites.

New UHPLC-MS/MS method applied to rapid drug analysis

Blood samples were centrifuged and trimipramine-d3 was added to the supernatant plasma as an internal standard. The plasma was extracted using liquid–liquid extraction (LLE) with a solvent mixture of diethyl ether and ethyl acetate. Evaporation of the combined extracts gave a residue that was taken up into methanol prior to chromatographic analysis. UHPLC employed an Accela system fitted with a Hypersil GOLD Phenyl column (both from Thermo Fisher). Gradient elution was carried out using aqueous 10 mM ammonium formate containing 0.1% formic acid at pH 3 and an organic phase of 0.1% formic acid in acetonitrile. The amount of organic phase was increased from 1% to 80% over 5 minutes, and then up to 99% over the next 0.5 mins, giving a total run time of 5.5 minutes. A cleaning flush protocol was then carried out using various solvent ratios of methanol and water in order to minimise any carry-over.

Mass spectrometry employed a Thermo Fisher TSQ Quantum Access instrument, which is a triple quadrupole MS/MS device. An APCI (atmospheric-pressure chemical ionisation) source was used, which previous work had shown to give more accurate results than the usual electrospray ionisation (ESI). The mass spectrometry was carried out in selected reaction monitoring (SRM) mode, quantifying and qualifying transitions being identified for the 45 compounds examined.

A standard mixture of the 45 compounds was prepared and stored at 8 °C. When needed, the mixture was dissolved in methanol, added to blank plasma and extracted as above. Quantification was carried out using one-point calibration in order that fast analyses could be run. Despite only using a one-point calibration, it was shown that most of the compounds gave reasonable linearity (defined as a slope of 1.0 ± 30%). With regard to accuracy, 32 of the 45 compounds gave sufficient accuracy (defined as ±30%) at lower levels (‘therapeutic’ levels), while 41 out of the 45 gave sufficient accuracy at higher levels (‘overdose’ or ‘toxic’ levels).

The amount of drugs present in the blood samples was classified into one of four groups: ‘toxic’, ‘overdose’, ‘therapeutic dose’ and ‘sub-therapeutic dose’. Comparison of the ‘emergency’ methods (the current UHPLC method and the GC method of Meyer et al.) with lengthy, validated therapeutic drug monitoring (TDM) methods for eight drugs from three patients gave the same classifications regardless of the method used, although the differences between the ‘emergency’ methods and the TDM methods ranged from 13% up to 55%.

New emergency method gives rapid results

The emergency UHPLC-MS/MS method, in combination with the complementary GC method of Meyer et al., gave fast, reasonably accurate results for a wide range of compounds. The use of short run times and one-point calibration allows results to be obtained quickly. Traditional methods do give greater accuracy, but this is of little use if the patient has died in the meantime.

Related Links

Drug Testing and Analysis, 2018, 10, 164-176. Michely et al.. A multi-analyte approach to help in assessing the severity of acute poisonings – Development and validation of a fast LC–MS/MS quantification approach for 45 drugs and their relevant metabolites with one-point calibration.

Drug Testing and Analysis, 2014, 6, 472-481. Meyer et al.. Development and validation of a fast and simple multi-analyte procedure for quantification of 40 drugs relevant to emergency toxicology using GC-MS and one-point calibration.

Wikipedia, Atmospheric-Pressure Chemical Ionization

Article by Ryan De Vooght-Johnson

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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