Detection of Drug Misuse: Biomarkers, Analytical Advances and Interpretation [1 ed.] 1782621571, 9781782621577

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Detection of Drug Misuse Biomarkers, Analytical Advances and Interpretation

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Detection of Drug Misuse Biomarkers, Analytical Advances and Interpretation Edited by

Kim Wolff King’s College London, UK Email: [email protected]

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Print ISBN: 978-1-78262-157-7 A catalogue record for this book is available from the British Library © The Royal Society of Chemistry 2017 All rights reserved Apart from fair dealing for the purposes of research for non-commercial purposes or for private study, criticism or review, as permitted under the Copyright, Designs and Patents Act 1988 and the Copyright and Related Rights Regulations 2003, this publication may not be reproduced, stored or transmitted, in any form or by any means, without the prior permission in writing of The Royal Society of Chemistry or the copyright owner, or in the case of reproduction in accordance with the terms of licences issued by the Copyright Licensing Agency in the UK, or in accordance with the terms of the licences issued by the appropriate Reproduction Rights Organization outside the UK. Enquiries concerning reproduction outside the terms stated here should be sent to The Royal Society of Chemistry at the address printed on this page. Whilst this material has been produced with all due care, The Royal Society of Chemistry cannot be held responsible or liable for its accuracy and completeness, nor for any consequences arising from any errors or the use of the information contained in this publication. The publication of advertisements does not constitute any endorsement by The Royal Society of Chemistry or Authors of any products advertised. The views and opinions advanced by contributors do not necessarily reflect those of The Royal Society of Chemistry which shall not be liable for any resulting loss or damage arising as a result of reliance upon this material. The Royal Society of Chemistry is a charity, registered in England and Wales, Number 207890, and a company incorporated in England by Royal Charter (Registered No. RC000524), registered office: Burlington House, Piccadilly, London W1J 0BA, UK, Telephone: +44 (0) 207 4378 6556. Visit our website at www.rsc.org/books Printed in the United Kingdom by CPI Group (UK) Ltd, Croydon, CR0 4YY, UK

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Preface The general aim in producing this book has been to illustrate the manner in which drug detection is carried out in different environments and with different matrices. In the last two decades, the interest in drug and alcohol use has markedly increased across the globe. This is due to both an increase in the availability and use of psychoactive substances and an increased focus from those involved in national and international public health and law enforcement agencies. The publication of the Detection of Drug Misuse comes at an important and intriguing time, with drug use in sport and in drug driving being focuses of international discourse. The manner in which drugs are detected in biological material and the way in which the results are interpreted and reported have received significant attention. It is hoped that this book will supplement knowledge and add to the wider process of review, consultation and evaluation that is taking place throughout the field and enable the reader to develop a view in terms of what, how and why we seek to detect misused drugs. It has been the aim to reflect upon the state-of-the-art analytical advances in the detection of drug and alcohol use, including innovative developments in sample collection. The intention is to provide academics, postgraduate and undergraduate students, drug and alcohol professionals and forensic and toxicological scientists with a high-quality book that covers key contemporary issues in the field. To this end, the authors have been chosen because of their particular expertise in different fields. Each chapter offers critical and up-to-date content on the contemporary issues in the detection of drug misuse. The sporting environment, civil aviation and drug driving are given particular attention. The authors are geographically dispersed, bringing the advantage of an international flavour to the sections on the ‘detection of new psychoactive substances’ and the section on ‘innovative approaches to sample collection’. In summary, this book provides an overview of the latest ideas, research and experience concerning drug detection of misused substances. Hopefully, Detection of Drug Misuse: Biomarkers, Analytical Advances and

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Interpretation will become a resource for the growing academy, especially at a time when these issues are gaining prominence in the public domain. Kim Wolff

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Contents Section I: Analytical Advances in Drug Detection 1 Urinalysis: The Detection of Common Drugs in Urine Michael David Osselton 1.1 Introduction and Historical Background 1.2 Urinary Drug Excretion 1.3 Urine Collection and Storage 1.4 Interpretation of Analytical Results 1.5 Approaches to Screening 1.6 Confirmatory Analysis References 2 Point-of-Care/Collection Testing: Application to Drugs of Misuse Testing Claire George and Alan Pang 2.1 Introduction 2.2 Principle of POCT Drug Tests 2.3 Parameters of Test Performance 2.3.1 Analytical Specificity 2.3.2 Cross-Reactivity 2.3.3 Analytical Sensitivity 2.4 Selection of a Drug Testing Device 2.5 Interpretation of POCT Results 2.5.1 True-Positive Results 2.5.2 True-Negative Results 2.5.3 False-Positive Results 2.5.4 False-Negative Results

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2.6 Quality and Accreditation 2.7 Innovation in POCT for Drugs of Misuse References 3 Analytical Advances in Drug Detection: Human Sports Drug Testing D. A. Cowan 3.1 Introduction 3.2 Sample Collection 3.3 Sample Screening Methods 3.3.1 Foreign Substances 3.3.2 Pseudo-Endogenous Substances 3.3.3 The ABP for Blood Samples 3.3.4 The Use of Biomarkers to Evidence Drug Misuse 3.3.5 Detection of Gene Doping 3.4 Conclusion References 4 Analytical Overview of Drug Detection: Civil Aviation Nigel P. Dowdall 4.1 Introduction 4.1.1 Aircraft Accidents: Drug and Alcohol Misuse 4.2 Regulations and Guidance 4.2.1 ICAO 4.2.2 USA 4.2.3 Europe 4.2.4 UK 4.2.5 Other Countries 4.3 Identifying Drug and Alcohol Misuse in Aviation 4.3.1 Prevalence 4.3.2 Drug and Alcohol Testing 4.3.3 Questionnaire Screening 4.3.4 Reporting by Enforcement Agencies, Health Professionals and ‘Whistle-Blowers’ 4.3.5 Reporting by Peers and Employers 4.4 Conclusion References 5 Detection of Misused Drugs: Natural and Synthetic Cathinones Anders Helander

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5.1 Cathinone 5.1.1 Origin and Use 5.1.2 Clinical Effects 5.2 Synthetic Substituted Cathinones 5.2.1 Origin and Use of Substituted Cathinones 5.2.2 Structures of Substituted Cathinones 5.2.3 Clinical Effects 5.3 Analysis of Synthetic Substituted Cathinones 5.3.1 Analytical Problems Related to NPSs 5.3.2 Immunoassay Screening 5.3.3 MS Analysis 5.3.4 Analysis of Parent Compounds or Metabolites 5.4 Conclusion References 6 Detection of Misused Drugs: Psychoactive Piperazines L. J. Schep, H. A. Poulsen and P. Gee 6.1 Introduction 6.2 Pharmacology and Toxicology 6.3 Methods of Detection of Piperazines in Biological Samples 6.3.1 Immunoassays 6.3.2 Gas Chromatography and Liquid Chromatography Mass Spectrometry 6.4 Identification of Psychoactive Piperazines in Powder and Tablet Formulations 6.5 Psychoactive Piperazines in Biological Samples 6.6 Conclusion References

Section II: Innovation in Sample Collection & Assay Methodology 7 Dried Blood Spots for Testing Drugs of Misuse R. Quraishi, R. Jain and A. Ambekar 7.1 Introduction 7.2 DBS Sampling: Technique and Relevant Factors 7.2.1 DBS Sampling Paper 7.2.2 Preparation of DBSs 7.2.3 Drying of DBS Samples

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7.2.4 Storage and Transportation of DBS Samples 7.3 DBSs to Test DOAs 7.3.1 Preparation of Calibrators, Quality Controls and Blanks for DBS Assays 7.3.2 Haematocrit and Blood Spot Volume 7.3.3 Extraction and Derivatisation of Drugs from DBSs 7.3.4 Assay Technique 7.3.5 Assay Validation: Precision, Accuracy and Sensitivity 7.3.6 Matrix Effect and Recovery 7.3.7 Clinical Validation and Stability 7.4 Advantages and Disadvantages of Using DBSs to Test DOAs 7.5 Conclusion and Future Perspective Acknowlegements References 8 Drug Testing in Exhaled Breath Markus R. Meyer and Olof Beck 8.1 Historical Background of Breath Drug Testing 8.2 Non-Volatiles in Exhaled Breath 8.3 Exhaled Breath Condensate (EBC) 8.4 Aerosol Particles in Exhaled Breath 8.5 Sampling Procedures 8.6 Analytical Methods and Detected Substances 8.7 Studies on Methadone and Clinical Trials 8.8 Possible Applications of Drug Testing in Exhaled Breath 8.9 Portable Detectors 8.10 Practical Aspects References 9 DNA/RNA Aptamers for Illicit Drug Molecules M. C. Parkin and N. Frascione 9.1 Introduction 9.1.1 Molecular Recognition and Binding for Drug Misuse Detection 9.1.2 Current Molecular Recognition for Drug Misuse Detection: Antibodies 9.1.3 Future Molecular Recognition for Drug Misuse Detection: Aptamers 9.2 Aptamer Selection

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9.2.1 SELEX and Its Variations 9.2.2 Cloning and Sequencing 9.2.3 KD Calculations 9.3 Current Applications of Aptamers for Drug Misuse Detection 9.3.1 Large-Molecule Targets 9.3.2 Small-Molecule Targets 9.3.3 Complexities of Aptamer Selection for Small-Molecule Targets 9.4 Aptamer-Based Detection Systems 9.4.1 Aptamer-Based Sensing 9.4.2 Portable Detection Systems 9.5 Conclusion References 10 Latent Fingerprints for Drug Screening Susan van der Heide and David A. Russell 10.1 Latent Fingerprints 10.1.1 Fingerprints for Drug Detection 10.2 Immunological and Aptamer-Based Methods for Detecting Drugs in Fingerprints 10.2.1 Immunoassays 10.2.2 Antibody-Functionalised Gold Nanoparticles 10.2.3 Antibody-Functionalised Magnetic Particles 10.2.4 Nanoplasmonic Imaging of Gold Nanoparticles 10.2.5 Aptamer-Functionalised Upconverting Nanoparticles 10.3 Vibrational Spectroscopy for Detecting Drugs in Fingerprints 10.3.1 Infrared Spectroscopy 10.3.2 Raman Spectroscopy 10.4 MS for the Detection of Drugs in Fingerprints 10.4.1 Chromatography with MS Detection 10.4.2 Imaging of Fingerprints with Surface-Assisted Laser Desorption/Ionisation MS 10.4.3 Imaging of Fingerprints with Time-of-Flight Secondary Ion MS 10.4.4 Direct Analyte-Probed Nanoextraction Coupled with Nanospray Ionisation MS 10.4.5 Imaging of Fingerprints with Matrix-Assisted Laser Desorption Ionisation MS 10.4.6 Imaging of Fingerprints with Desorption Electrospray

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Ionisation 10.5 Summary and Outlook Acknowledgements References 11 Microneedle Patches for Caffeine Detection and Quantification Ester Caffarel-Salvador, Aaron John Brady and Ryan F. Donnelly 11.1 Sources of Caffeine 11.2 Caffeine Pharmacokinetics 11.3 Pharmacological Effects of Caffeine 11.4 Misuse of Caffeine 11.5 Treatment 11.6 Monitoring Caffeine 11.7 Conclusion References 12 Detection of a Single Drug Exposure in Hair Pascal Kintz 12.1 Introduction 12.2 Review Method 12.3 Controlled Dose Studies 12.4 Models for Drug Incorporation into Hair 12.4.1 Physiochemical Properties of Drugs 12.4.2 The Specific Problem of Segmental Hair Analysis 12.4.3 Conflicting Results After Single Exposures to Zolpidem and Relevance in the Courts 12.5 Conclusion References

Section III: Interpretation and Reporting Results 13 Ethanol Analysis in Blood, Breath and Urine: Interpreting the Results Alan Wayne Jones 13.1 Introduction 13.2 Amounts of Alcohol Consumed 13.3 Alcohol and Traffic Safety 13.4 Statutory Alcohol Limits for Driving 13.5 Concentration Units

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13.6 Sampling of Blood, Breath and Urine 13.6.1 Blood Samples 13.6.2 Proficiency Testing 13.6.3 Urine Samples 13.6.4 Urine: Blood Concentration Ratios 13.6.5 Breath Samples 13.6.6 Blood: Breath Concentration Ratios 13.7 Alcohol in the Body 13.7.1 Absorption 13.7.2 Distribution 13.7.3 Metabolism 13.7.4 Excretion 13.8 Applications of the Widmark Equation 13.9 Effects of Ethanol on the Brain 13.10 Relationship Between BAC and Signs of Intoxication 13.11 Concluding Remarks 13.12 Future Considerations References 14 ‘Ecstasy’ Tablets: Batch Matching for Forensic Drug Intelligence Purposes in Malta Alan Wayne Jones 14.1 Introduction 14.1.1 Characterisation of Ecstasy Tablets 14.2 Drug Seizures 14.3 Visual Characteristics of MDMA/‘Ecstasy’ Tablets Seized in Malta 14.3.1 Measurable Features: Mass, Diameter and Thickness 14.3.2 Comparable Characteristics and Trends in Tablet Mass, Diameter and Thickness 14.3.3 Physical Characteristics: Hardness, Friability and Disintegration Rate 14.3.4 Novel Approaches when Assessing the Measurable Characteristics (Mass, Diameter, Thickness, Hardness, Friability and Disintegration Rate) of ‘Ecstasy’ Tablets 14.4 Conclusions References 15 The Usefulness of Metabolites in the Interpretation of Drug Test

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Results Michael Scott-Ham 15.1 Introduction 15.2 Pre-Analytical Considerations 15.3 Choice of Biological Matrix 15.4 Detection 15.5 Drugs/Drug Groups 15.5.1 Amphetamine, Methamphetamine and Methylenedioxymethamphetamine 15.5.2 Cannabis 15.5.3 Cocaine 15.5.4 Opiates/Opioids 15.5.5 Ketamine 15.5.6 Benzodiazepines 15.5.7 Medicinal Drugs 15.5.8 New Psychoactive Substances References 16 The Usefulness of Metabolic Ratios in the Interpretation of Steroid Misuse A. D. Brailsford 16.1 Introduction to Testosterone 16.2 Testosterone Misuse 16.2.1 Steroid Misuse in Society 16.2.2 Historical Aspects of Performance-Enhancing Drugs 16.2.3 Testosterone Misuse in Sports 16.3 The Testosterone/Epitestosterone (T/E) Ratio 16.3.1 Genetic Factors Influencing the Testosterone/Epitestosterone (T/E) Ratio 16.4 Longitudinal Athlete (Individual) Profiling 16.4.1 The Athlete Biological Passport 16.4.2 The ABP Steroid Module 16.4.3 Confounding Factors of Steroid Ratios 16.5 Glycoprotein Hormones 16.5.1 Luteinising Hormone 16.5.2 Human Chorionic Gonadotropin 16.5.3 Confounding Factors 16.6 Analytical Techniques 16.6.1 Isotope Ratios

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16.6.2 GC-C-IRMS Instrumentation 16.6.3 δ Values 16.6.4 IRMS and Steroids 16.7 Looking Forward References 17 Neurohypophyseal Hormones and Drugs of Misuse Jacinta L. Johnson, Michaela E. Johnson and Femke BuismanPijlman 17.1 Introduction to the Neurohypophyseal Hormones 17.1.1 Synthesis of the Neurohypophyseal Hormones 17.1.2 Neurohypophyseal Hormone Receptors 17.1.3 Physiological Roles and Regulation of the Neurohypophyseal Hormones 17.2 Laboratory Tests for Detecting Oxytocin 17.2.1 Detection of Oxytocin via Radioimmunoassay 17.2.2 Detection of Oxytocin via Enzyme Immunoassay 17.2.3 Detection of Oxytocin via Liquid ChromatographyMass Spectrometry (LC-MS) 17.3 Laboratory Tests for Detecting Vasopressin 17.3.1 Indirect Methods of Measuring Vasopressin 17.3.2 Direct Methods of Measuring Vasopressin 17.4 The Relationship Between Peripheral and Central Concentrations of Oxytocin and Vasopressin 17.5 The Relationship Between the Neurohypophyseal Hormones and Drugs of Misuse 17.5.1 Neurohypophyseal Hormones and Opioid Addiction 17.5.2 Neurohypophyseal Hormones and Cocaine Misuse 17.5.3 Neurohypophyseal Hormones and Use of Amphetamine-Type Stimulants 17.5.4 Neurohypophyseal Hormones and Use of 3,4Methylenedioxymethamphetamine 17.5.5 Neurohypophyseal Hormones and Alcohol Misuse 17.5.6 Oxytocin and Susceptibility to Addiction 17.6 Conclusion References Subject Index

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Section I Analytical Advances in Drug Detection

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1 Urinalysis: The Detection of Common Drugs in Urine Michael David Osseltona a

Department of Archaeology, Anthropology and Forensic Science, Faculty of Science and Technology, Bournemouth University, Christchurch House, Fern Barrow, Poole, BH12 5BB, UK *E-mail: [email protected]

1.1 Introduction and Historical Background Mathieu Orfila,2 Robert Christison3 and Alfred Swaine Taylor1 were amongst the first early toxicologists in the 18th and 19th centuries to record the use of urine as an aid to poison detection. “One of the strongest proofs of poisoning in the living subject is the detection of poison by chemical analysis in the matters vomited or in the urine, if the poison be one of those which are eliminated from the kidneys. The evidence is, of course, more satisfactory when the substance is discovered in the matter vomited or in the urine because it will show that the poison has really been taken, and will at once account for the symptoms.”1 At the time of Orfila, Christison and Swaine Taylor, the mechanisms by which poisons exerted their action in the body were unknown, and tests for the presence of poisons in urine were largely associated with administering the urine of suspected poisoning victims to animals, yet urine in the diagnosis and detection of drugs and poisons was later recognised as a valuable matrix for toxicological investigation. Only after the introduction of gas chromatography (GC) into forensic laboratories in

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the 1960’s were toxicologists able to move away from urine in order to explore the use of blood, where better interpretation can be made from analyses. Today, urine still occupies an important place in forensic, sports, clinical and workplace drug testing, thanks largely to developments in high-performance liquid chromatography linked to mass spectrometry (HPLC-MS).

1.2 Urinary Drug Excretion Urine is produced in the kidneys and comprises water (>95%) together with the waste products of metabolism. The kidney plays a key function in maintaining the health of an individual by regulating the salt and water balance in the body and providing a route for the elimination of metabolic excretion products and toxic substances. Urine comprises a concentrated solution of filtered waste products that pass out of the body via the bladder. In the healthy individual, the urine is a clear, pale yellow fluid containing water (>95%), urea (∼2%), creatinine (∼0.1%; >2 mmol L-1 or >226 mg L-1) and small quantities of salts, together with water-soluble drugs and their metabolites. The normal glomerular filtration rate is approximately 120 mL minute-1. After reabsorption of salts, glucose and water in the proximal tubules of the kidney, the filtrate reaches the loop of Henle with the same osmotic pressure as plasma at a rate of approximately 20 mL minute-1. After reabsorption of water in the distil tubules, the final urine flow is less than 0.5 mL minute-1. The rate of urine production varies with age, such that a 1-week-old baby produces 50–800 mL 24 hours-1, a 3-year-old child produces 500–700 mL 24 hours-1, a 10-year-old produces 700–1400 mL 24 hours-1 and a healthy adult produces 800–2000 mL 24 hours-1.4 Normal urine has a specific gravity of greater than 1.025. Specimens with a specific gravity of less than 1.001 may indicate dilution caused by excessive fluid consumption, diabetes insipidus, impaired renal function resulting from a low number of functioning glomeruli or, in workplace drug testing scenarios, an attempt by the donor to dilute their urine specimen to try to confound the testing process. Although specific gravity is infrequently measured in post-mortem and criminal toxicology, it is regularly measured in workplace drug testing laboratories as a test for deliberate specimen dilution. The pH of urine is naturally variable and may range between 4.5 and 8.0.

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Depending on the pH of the urine and the state of ionisation and/or water solubility of any drugs or their metabolites present in the glomerular filtrate, drugs and their metabolites become trapped in the urine and are subsequently excreted from the body. Although drug concentrations in the glomerulus are similar to those in blood, the reabsorption of water back into the blood as the urine passes through the convoluted tubules and loop of Henle results in increased drug concentrations in urine, such that drug concentrations may be between 10 and 100 times more concentrated in urine than those in the circulating blood. As a matrix that is largely aqueous and free from proteins and cells in normal healthy individuals (cf. blood), where drug concentrations are usually significantly higher than in blood and where drugs may accumulate prior to elimination from the body, urine provides an ideal sample for drug screening. As a consequence of this concentration effect, drugs and their metabolites may be detected in urine long after they become undetectable in blood; hence, urine analysis provides an extended window of detection for the forensic or clinical analyst. Approximate time intervals for the detection of drugs in urine following single dose consumption are shown in Table 1.1. Table 1.1

Approximate detection times for determining the presence of drugs and their metabolites in urine following the consumption of a single dose or therapeutic dose.a

Substance

Detection time (hours)b

Alcohol Amphetamine Diazepam (including metabolites) Temazepam Flunitrazepam/7-amino flunitrazepam THC Cocaine Benzoylecgonine GHB (γ-Hydroxybutyrate) Morphine 6-MAM (6-monoacetyl morphine) Codeine Methadone/EDDP MDMA Ketamine/norketamine LSD 2-Oxo-3-hydroxy-LSD