Advances in Forensic Applications of Mass Spectrometry

Jehuda Yinon, Jehuda Yinon

• Publisher: CRC Press
• Year: 2003
• ISBN:
  9781135505066
• Edition: 1
• Topic: Law
• Subtopic: Forensic Science
• Index: Law

Recent developments in analytical instrumentation have had an enormous influence on forensic analysis. The mass spectrometer is now an integral part of every forensic laboratory, resulting in greater analytical accuracy, more reliable identification, and lower detection limits. As the instrumental method of choice among forensic analysts, the mass

1 Screening for Drugs in Body Fluids by GC/MS

HANS H. MAURER

Contents

1.1 Introduction!

1.1.1 Matrices for Drug Screening

1.1.2 Sample Preparation

1.2 Screening for Drugs in Blood, Serum, or Plasma by GC/MS

1.3 Screening for Drugs in Alternative Matrices by GC/MS

1.4 Screening for Drugs in Urine by GC/MS.

1.4.1 Screening Procedures for Detection of Particular Drug Classes in Urine by GC/MS

1.4.1.1 Screening Procedures for Detection of Particular Drug Classes of Basic or Neutral Drugs and/or Their Metabolites in Urine

1.4.1.2 Screening Procedures for Detection of Particular Drug Classes of Acidic Drugs and/or Their Metabolites in Urine

1.4.2 General Screening Procedures for Simultaneous Detection of Several Drug Classes in Urine by GC/MS

1.4.2.1 General Screening Procedure for Drug Classes Relevant in Doping after Enzymatic Hydrolysis, SPE, and Combined TMS and TFA Derivatization

1.4.2.2 General Screening for Most of the Basic and Neutral Drug Classes in Urine after Acid Hydrolysis, LLE, and AC.

1.4.2.3 General Screening Procedures for Simultaneous Detection of Several Classes of Acidic Drugs in Urine after Extractive Methylation

1.5 Conclusions and Perspectives.

Acknowledgments

References

1.1 Introduction

In forensic toxicology, proof of abuse of illegal drugs or of murder by poisoning are important tasks. Furthermore, drugs, which may reduce the penal responsibility of a defendant, or which may reduce the fitness to drive a car, must be monitored in body fluids or tissues.

In clinical toxicology, the diagnosis or the definite exclusion of an acute or chronic poisoning is of great importance. Furthermore, patients addicted to alcohol, medicaments, or illegal drugs have to be monitored. For determination of clinical death as a prerequisite for explantation of organs, the presence of drugs which may depress the central nervous system must be analytically excluded. The compliance of patients can be monitored by determination of the prescribed drugs. Finally, monitoring of drugs with a narrow therapeutic range can be performed by the clinical toxicologist. Similiar problems arise in forensic toxicology.

In doping control, the use or abuse of drugs that may stimulate the buildup of muscles, enhance endurance during competition, lead to reduction of body weight, or that may reduce pain caused by overexertion must be monitored, typically in urine.

An efficient toxicological analysis is the basis of competent toxicological judgement, consultation, and expertise. The choice of methods in analytical toxicology depends on the problems to be solved. Usually, the compounds to be analyzed are unknown. Therefore, the first step is the identification of the compounds of interest which can then be quantified, e.g., in plasma. The screening strategy of systematic toxicological analyses (STA) must be very extensive because several thousands of drugs or pesticides have to be considered. It often includes screening and confirmatory tests. If only a single drug or category has to be monitored, immunoassays can be used for screening in order to differentiate between negative and presumptively positive samples. Positive results must be confirmed by a second independent method that is at least as sensitive as the screening test and that provides the highest level of confidence in the result. Without doubt, GC/MS, especially in the full-scan electron ionization (EI) mode, is still the reference method for confirmation of positive screening tests.^1–21 Nevertheless, LC/MS has also been applied for screening and confirmation of particular drugs or drug classes, especially in blood.^22–31

The two-step strategy, immunoassay screening and MS confirmation, is employed only if those drugs or poisons have to be determined that are scheduled, e.g., by law or by international organizations, and for which immunoassays are commercially available. If this is not the case, the screening strategy must be more extensive, because several thousands of drugs or pesticides are on the market worldwide.³² For these reasons, STA procedures are necessary that allow the simultaneous detection of as many toxicants in biosamples as possible. As already mentioned, GC/MS procedures are most often used today. HPLC coupled to diode array detectors (DAD)^33–39 have also been described for general screening purposes, but the specificity is lower than that of full-scan EI MS. Valli et al.⁴⁰ combined GC/MS blood screening with urine REMEDi testing and Saint-Marcoux et al.²⁶ combined GC/MS, HLPC-DAD, and LC/MS. LC/MS procedures in this field were several times reviewed in the last years, e.g., by Maurer,⁴¹ Van Bocxlaer et al.,⁴² Marquet,²⁸ and Bogusz.^31,43

Most of the STA procedures cover basic (and neutral) drugs, which include the majority of toxicants. For example, most of the psychotropic drugs have basic properties like neurotransmitters. Nevertheless, some classes of acidic drugs or drugs producing acidic metabolites, like cardiovascular drugs such as angiotensin converting enzyme (ACE) inhibitors and angio-tensin II AT₁ receptor blockers, dihydropyridine-type calcium channel blockers (metabolites), diuretics, coumarin anticoagulants, antidiabetics of the sulfonylurea type, barbiturates, or nonsteroidal antiinflammatory drugs (NSAIDs), are relevant to clinical and forensic toxicology or doping. Therefore, GC/MS screening procedures are described here not only for detection of basic and neutral, but also acidic drugs in biosamples. After the unequivocal identification, reliable quantification of the drugs can also be performed by GC/MS, especially if stable isotopes are used as internal standards (for example, see References 44, 45), or in case of rather polar or unstable compounds, by LC/MS, even using universal internal standards like deuterated trimipramine.^22,24,25,46 However, whatever technique is used, quantification procedures must be validated according to international guidelines, which have recently been reviewed by Peters and Maurer.⁴⁷

1.1.1 Matrices for Drug Screening

Blood (plasma, serum) is the sample of choice for quantification. However, if the blood concentration is high enough, screening can also be performed herein. This is especially advantageous if only blood samples are available and/or the procedures allow simultaneous screening and quantification. ^6,9,32,48,49 In driving under the influence of drugs (DUID) cases, blood analysis is even mandatory. A GC/MS screening procedure has been described for about 100 acid, neutral, and basic drugs in horse plasma.⁵⁰ Methods for postmortem drug analysis have been reviewed recently.⁵¹

GC/MS analysis of drugs in alternative matrices like hair, ^8,19,52 sweat and saliva,^7,19 meconium,⁵³ or nails⁵⁴ have also been described, but a comprehensive screening for a series of various drugs has not yet been described in alternative matrices, probably because the concentrations are too low for full-scan GC/MS detection. Negative-ion chemical ionization (NICI) allows markedly lowering the detection limits,^20,24,44 but this technique is not suitable for comprehensive screening because the analytes must contain an electronegative moiety and the NICI mass spectra are less informative and reproducible than EI spectra.

In conclusion, urine is still the sample of choice for comprehensive screening for, and identification of, unknown drugs or poisons, mainly because concentrations of drugs are relatively high in urine and the samples can be taken noninvasively.^{1,2,4,16,18,32} However, the metabolites of these unknowns must be identified, additionally or even exclusively. In (horse) doping control, urine is also the common sample for screening.⁵⁵

1.1.2 Sample Preparation

Suitable sample preparation is an important prerequisite for GC/MS analysis in biosamples. It may involve cleavage of conjugates, isolation, and derivati-zation, preceded or followed by cleanup steps. Cleavage of conjugates can be performed by fast acid hydrolysis or by gentle but time-consuming enzymatic hydrolysis.² However, the enzymatic hydrolysis of acyl glucuronides (ester glucuronides of carboxy derivatives like NSAIDs) may be hindered due to acyl migration,⁵⁶ an intramolecular transesterification at the hydroxy groups of the glucuronic acid, which leads to β-glucuronidase-resistant derivatives. If the analysis must be finished within a rather short time as in emergency toxicology, it is preferable to cleave the conjugates by rapid acid hydrolysis. ^57–62 Alkaline h...

Table of contents

• Cover Page
• Title Page
• Copyright Page
• Introduction
• The Editor
• Contributors
• List of Abbreviations
• 1: Screening for Drugs in Body Fluids by GC/MS
• 2: Liquid Chromatography/Mass Spectrometry in Forensic Toxicology
• 3: Substance Abuse in Sports: Detection of Doping Agents in Hair by Mass Spectrometry
• 4: Forensic Applications of Isotope Ratio Mass Spectrometry
• 5: Identification of Ignitable Liquid Residues in Fire Debris by GC/MS/MS
• 6: Analysis of Explosives by LC/MS