Toxtyper selection speeds overdose detection

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  • Published: Sep 29, 2018
  • Author: Ryan De Vooght-Johnson
  • Copyright: Image: photka/Shutterstock
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Speedy identification needed with pharmaceutical overdoses

Credit: photka/Shutterstock.

Overdoses of pharmaceuticals, whether deliberate or accidental, need rapid analysis of the substances involved so that appropriate treatment can be given. The patient may be unconscious, confused or uncooperative, so is often of little help to medical staff. Qualitative and quantitative determination of the drugs present in blood is vital so that the correct treatment can be given.

There is a need for standard, proven methods to deal with emergency drug determination in plasma. One standardised protocol, the Bruker Daltonik Toxtyper system, uses LC-MS with fixed equipment and parameters, along with associated software and an MS library, to give qualitative data on a range of pharmaceutical compounds in blood plasma. The Saarland University researchers sought to extend the protocol to give rapid quantification of 22 drugs and two drug metabolites in a single run. The drugs included various painkillers, such as paracetamol (acetaminophen), and psychiatric pharmaceuticals, such as antidepressants.

Toxtyper protocol applied to 22 drugs and two metabolites

Five calibration solutions at different concentrations were prepared, each containing all 24 of the substances under investigation. Four QC solutions were also prepared: low, high, therapeutic and diluted (the drug concentration in the latter was typically half the ‘QC high’ concentration). The solutions were used to spike blank plasma solutions.

Plasma solutions (either spiked blanks or samples from patients) were extracted with a 1:1 mixture of ethyl acetate and diethyl ether. After evaporation the residue was taken up in methanol and mobile phase (aqueous acetonitrile containing ammonium formate and formic acid). HPLC employed a Dionex UltiMate 3000 system, fitted with a Thermo Scientific Acclaim 120 C18 column. Solvent A was 2 mM aqueous ammonium formate, containing 1% acetonitrile and 0.1% formic acid, while solvent B was acetonitrile, 2 mM with respect to ammonium formate, containing 1% water and 0.1% formic acid. The eluting solvent was kept at 1% B between 0 and 1 min, increased to 95% B between 1 and 8 min, kept at 95% B between 8 and 9 min, and held at 1% B between 9 and 11 min for re-equilibration.

Mass spectrometry used a Bruker amaZon speed ion trap instrument, run in autoMSn mode, recording MS2 (in some cases also MS3) spectra in positive or negative ion mode as required for each compound. The Toxtyper MS library was used to identify the compounds present.

The method was shown to give sufficient accuracy for use in emergency toxicology, where deviations of up to 30% in accuracy and precision are considered acceptable, since speed is the priority. The method was particularly inaccurate at low levels, but this is less important in the context of overdoses. Two compounds, paracetamol and carbamazepine, gave significant carry-over between injections when high levels were present, but this could be eliminated with a blank injection. Two pairs of compounds were not separated by the short gradient used, but adequate quantification could be obtained from the mass spectrometry. The method was tested on ten plasma samples from patients, containing drugs that were typically at toxic or therapeutic concentrations, where the results were in largely in line with those obtained by current methods.

Rapid analysis requires the storage of calibration parameters for extended periods of time so that no calibration injections are required in an emergency situation. Testing over 60 days showed that the saved calibration parameters (five-point calibration) were applicable over this time period for nearly all the 24 compounds, with only paracetamol showing deviations of more than 30% on more than one day. In practice, most testing facilities would check the calibration more often than 60 days.

Toxtyper methods shown to be applicable to quantification

The results show that the Toxtyper protocol and library enable rapid quantitative results for the 24 compounds examined. Such methods are extremely useful in emergency situations, where rapid analysis is needed, since multiple methods for different compounds can be replaced by a single fast run.

Related Links

Drug Testing and Analysis 2018, 1-10. Caspar et al.: "Blood plasma level determination using an automated LC–MSn screening system and electronically stored calibrations exemplified for 22 drugs and two active metabolites often requested in emergency toxicology."

Clinical Mass Spectrometry 2017, 4-5, 11-18. Ott et al.: "Detection of drugs of abuse in urine using the Bruker Toxtyper™: Experiences in a routine clinical laboratory setting."

Therapeutic Drug Monitoring 2018, 40, 642-648. Plecko et al.: "Evaluation of an ion trap LC-IT/MS instrument (Toxtyper) for drug of abuse screening in oral fluid."

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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