The doctoral dissertations of the former Helsinki University of Technology (TKK) and Aalto University Schools of Technology (CHEM, ELEC, ENG, SCI) published in electronic format are available in the electronic publications archive of Aalto University - Aaltodoc.
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Library Search-Based Drug Analysis in Forensic Toxicology by Liquid Chromatography–Mass Spectrometry

Merja Gergov

Dissertation for the degree of Doctor of Science in Technology to be presented with due permission of the Department of Chemical Technology for public examination and debate in Auditorium KE 2 (Komppa Auditorium) at Helsinki University of Technology (Espoo, Finland) on the 29th of October, 2004, at 12 o'clock noon.

Overview in PDF format (ISBN 951-22-7279-2)   [455 KB]
Dissertation is also available in print (ISBN 951-22-7278-4)

Abstract

Liquid chromatography–mass spectrometry (LC–MS) with electrospray (ES) ionization in the positive ion mode was applied to drug analysis in forensic toxicology. Several types of screening and confirmation strategies were developed, based essentially on searching of comprehensive MS libraries and databases that were created in-house for the purpose.

Two spectral libraries, both containing spectra for approximately 400 drugs, were created. Spectra for the LC–MS/in-source collision-induced dissociation (CID) library were acquired by continuously switching between low (25 V) and high (90 V) orifice voltages during the run, and the summed mass spectra were added to the library. A library for liquid chromatography–triple quadrupole mass spectrometry (LC–MS/MS) was created by obtaining product ion spectra at a collision energy of 35 eV, and additionally at 20 eV or 50 eV, if the latter were more informative. The product ion spectra were reproducible in the long-term, based on a four-year experiment. Comparison of two libraries, independently created with similar instruments in different laboratories, showed that the libraries were fully compatible. Comparison of spectra obtained with different manufacturers' instruments suggested that after standardization of collision energy and gas pressure, LC–MS/MS spectral libraries are suitable for interlaboratory use.

The libraries were utilized in two fully automated methods for selected drug groups, comprising screening and confirmation. Analysis of 16 β-blocking drugs in urine involved two-step MS analysis, including a screening step based on monitoring of protonated molecules and a confirmation step based on LC–MS/MS product ion spectra. A method for simultaneous screening and quantitation of 18 antihistamine drugs in blood involved multiple-reaction monitoring (MRM) for screening and quantitation and product ion spectra for confirmation of the drugs. The antihistamines were identified and quantified at concentrations ranging from subtherapeutic to toxic in blood. A data-dependent experiment (DDE) was utilized for automation of both procedures, from starting the sample batch to printing of the library search results.

Two different approaches for comprehensive screening of drugs were developed. A method for simultaneous screening of 238 drugs in blood was developed using LC–MS/MS with multiple-reaction monitoring. Identification was based on RT and the presence of protonated molecule and one representative fragment ion. The method was shown to be sufficiently selective and sensitive to allow the majority of the drugs to be detected at therapeutic concentrations. Another approach was based on accurate mass determination with liquid chromatography–time-of-flight mass spectrometry (LC–MS/TOF), using a target database of monoisotopic masses for 433 drugs. The method was tentatively suitable for rapid screening of drugs in urine even without referral to primary reference standards.

This thesis consists of an overview and of the following 5 publications:

  1. Gergov M., Robson J. N., Duchoslav E. and Ojanperä I., 2000. Automated liquid chromatographic/tandem mass spectrometric method for screening β-blocking drugs in urine. Journal of Mass Spectrometry 35, number 7, pages 912-918.
  2. Gergov M., Robson J. N., Ojanperä I., Heinonen O. P. and Vuori E., 2001. Simultaneous screening and quantitation of 18 antihistamine drugs in blood by liquid chromatography ionspray tandem mass spectrometry. Forensic Science International 121, numbers 1-2, pages 108-115.
  3. Gergov M., Weinmann W., Meriluoto J., Uusitalo J. and Ojanperä I., 2004. Comparison of product ion spectra obtained by liquid chromatography/triple-quadrupole mass spectrometry for library search. Rapid Communications in Mass Spectrometry 18, number 10, pages 1039-1046.
  4. Gergov M., Ojanperä I. and Vuori E., 2003. Simultaneous screening for 238 drugs in blood by liquid chromatography–ionspray tandem mass spectrometry with multiple-reaction monitoring. Journal of Chromatography B 795, number 1, pages 41-53.
  5. Gergov M., Boucher B., Ojanperä I. and Vuori E., 2001. Toxicological screening of urine for drugs by liquid chromatography/time-of-flight mass spectrometry with automated target library search based on elemental formulas. Rapid Communications in Mass Spectrometry 15, number 8, pages 521-526.

Keywords: liquid chromatography–mass spectrometry, screening, library search, drugs, forensic toxicology

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© 2004 Helsinki University of Technology

Last update 2011-05-26