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Benzodiazepines and GHB Detection and Pharmacology Edited by Salvatore J. Salamone Humana Press Benzodiazepines and GHB FORENSIC SCIENCE- AND- MEDICINE Steven B. Karch, MD, SERIES EDITOR BUPRENORPHINE THERAPY OF OPIATE ADDICTION, edited by PASCAL KINTZ AND PIERRE MARQUET, 2002 ON-SITE DRUG TESTING, edited by AMANDA J. JENKINS AND BRUCE A. GOLDBERGER, 2001 BENZODIAZEPINES AND GHB: DETECTION AND PHARMACOLOGY, edited by Salvatore J. Salamone, 2001 TOXICOLOGY AND CLINICAL PHARMACOLOGY OF HERBAL PRODUCTS, edited by Melanie Johns Cupp, 2000 CRIMINAL POISONING: INVESTIGATIONAL GUIDE FOR LAW ENFORCEMENT, TOXICOLOGISTS, FORENSIC SCIENTISTS, AND ATTORNEYS, by John H. Trestrail, III, 2000 A PHYSICIAN'S GUIDE TO CLINICAL FORENSIC MEDICINE, edited by Margaret M. Stark, 2000 BENZODIAZEPINES AND GHB Detection and Pharmacology Edited by Salvatore J. Salamone, PhD Roche Diagnostics Corp. Indianapolis, IN Humana Press Totowa, New Jersey To my children Katherine, Norah, and Hannah © 2001 Humana Press Inc. 999 Riverview Drive, Suite 208 Totowa, New Jersey 07512 www.humanapress.com All rights reserved. No part of this book may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, microfilming, recording, or otherwise without written permission from the Publisher. The content and opinions expressed in this book are the sole work of the authors and editors, who have warranted due diligence in the creation and issuance of their work. The publisher, editors, and authors are not responsible for errors or omissions or for any consequences arising from the information or opinions presented in this book and make no warranty, express or implied, with respect to its contents. This publication is printed on acid-free paper. ∞ ANSI Z39.48-1984 (American Standards Institute) Permanence of Paper for Printed Library Materials. Cover design by Patricia F. Cleary. For additional copies, pricing for bulk purchases, and/or information about other Humana titles, contact Humana at the above address or at any of the following numbers: Tel: 973-256-1699; Fax: 973-256-8341; E-mail: [email protected], or visit our Website at www.humanapress.com Photocopy Authorization Policy: Authorization to photocopy items for internal or personal use, or the internal or personal use of specific clients, is granted by Humana Press Inc., provided that the base fee of US $10.00 per copy, plus US $00.25 per page, is paid directly to the Copyright Clearance Center at 222 Rosewood Drive, Danvers, MA 01923. For those organizations that have been granted a photocopy license from the CCC, a separate system of payment has been arranged and is acceptable to Humana Press Inc. The fee code for users of the Transactional Reporting Service is: [0-89603-981-1/01 $10.00 + $00.25]. Printed in the United States of America. 10 9 8 7 6 5 4 3 2 1 Library of Congress Cataloging in Publication Data Benzodiazepines and GHB: detection and pharmacology p. ; cm. -- (Forensic science and medicine) Includes bibliographical references and index. ISBN 0-89603-981-1 (alk. paper) 1. Benzodiazepines--Analysis. 2. Gamma-hydroxybutyrate--Analysis. 3. Forensic pharmacology. I. Salamone, Salvatore J. II. Series. [DNLM: 1. Body Fluids--chemistry. 2. Forensic Medicine--methods. 3. Benzodiazepines--isolation & purification. 4. Benzodiazepines--metabolism. 5. Benzodiazepines--pharmacology. 6. Hair--chemistry. 7. Hydroxybutyrates--isolation & purification. 8. Hydroxybutyrates--metabolism. 9. Hydroxybutyrates-- pharmacology. W 750 D4799 2001] RA1242.B43 D48 2001 614'.1--dc21 2001024305 Preface Since the first benzodiazepines were introduced to the market in 1960, there has been an evolution of these drugs toward lower dosage, shorter action, and faster clearance. As a consequence, this new generation of benzodiaz- epines eluded detection in many laboratories. Benzodiazepines can be abused in several different ways. These drugs are often given in conjunction with alcohol to enhance the desired effect, thus requiring lower doses. This situation has created added challenges to the toxicologist. Use of normal screening and confirmation methods may not detect the lower levels of these drugs, and in some cases the laboratory may not have capability in the methodologies to detect a particular drug. Another recent challenge for the toxicologist is the detection of γ-hydroxy- butyric acid (GHB). The use of this drug is widespread and it is easily obtained or prepared in clandestine laboratories. Similar to the low-dosed benzodiazepines (LDBs) the effect is also potentiated by alcohol. Its detection is difficult owing to the rapid clearance and the low concentrations that appear in urine or serum. Detection methods for GHB are not common, but labora- tories are now developing new methods as its popularity is highlighted. The purpose of Benzodiazepines and GHB: Detection and Pharmacology is to provide some background on the pharmacology and metabolism of LDB and GHB and to help the toxicologist develop methodologies that will enable better detection of these drugs in various body fluids, as well as in hair. The first chapter provides background on the LDBs by dealing with the pharmacol- ogy and metabolism of these drugs. Chapter 2 deals with immunoassay detection of LDBs, reviewing the current state of testing and providing methodologies that will increase the sensitivity of immunoassay reagents. Chapters 3 and 4 focus on methods for the detection of Rohypnol® and other LDBs by mass spectrometry. Chapter 5 addresses the detection of benzodiazepines in hair. Chapter 6 addresses the pharmacology and detection of GHB, and finally v vi Preface Chapter 7 presents a case study examining the prevalence of drugs used in cases of alleged sexual assault. Benzodiazepines and GHB: Detection and Pharmacology would not have been possible without the cooperation of all the authors. I wish to thank each of them for their efforts. I would like to thank Dr. Stephen Karch and Humana Press for the opportunity to be involved with such an endeavor. Salvatore J. Salamone, PhD Contents Preface .................................................................................................... v Contributors .......................................................................................... xi CHAPTER 1 Pharmacology of Flunitrazepam and Other Benzodiazepines Rudolf Brenneisen and Lionel Raymond ................................................. 1 1. Introduction .......................................................................................................... 1 2. Chemistry ............................................................................................................. 2 3. Pharmacokinetics ................................................................................................. 4 3.1. Absorption, Distribution, and Plasma Levels ............................................. 4 3.2. Half-Life vs Duration of Action .................................................................. 7 3.3. Metabolism and Excretion........................................................................... 7 4. Pharmacodynamics .............................................................................................. 8 4.1. Sites of Action.............................................................................................. 8 4.2. Acute Effect ............................................................................................... 10 4.3. Chronic Administration and Abuse Liability ........................................... 11 4.4. Pharmacokinetics vs Pharmacodynamics ................................................. 12 4.5. Rohypnol®: The Debate ............................................................................ 13 References .............................................................................................................. 14 vii viii Contents CHAPTER 2 Immunoassay Detection of Benzodiazepines Tamara N. St. Claire and Salvatore J. Salamone ...................................17 1. Introduction ........................................................................................................ 17 2. Optimal Enzyme Activity .................................................................................. 19 2.1. Conditions .................................................................................................. 19 2.2. Activity ....................................................................................................... 20 3. Cutoff ................................................................................................................. 22 4. Immunoassays .................................................................................................... 22 5. Methods .............................................................................................................. 28 5.1. Manual β-Glucuronidase Treatment ......................................................... 28 5.2. Automated β-Glucuronidase Treatment ................................................... 28 5.2.1. Online with the Hitachi 717............................................................ 28 5.2.2. Online with the COBAS INTEGRA 700 ....................................... 29 5.2.3. A General Scheme........................................................................... 29 6. Conclusion ......................................................................................................... 29 References .............................................................................................................
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  • Drug Screening: Actual Status, Pitfalls and Suggestions for Improvement Drogenscreening: Gegenwa¨ Rtiger Stand, Fehlermo¨ Glichkeiten Und Verbesserungsvorschla¨Ge

    Drug Screening: Actual Status, Pitfalls and Suggestions for Improvement Drogenscreening: Gegenwa¨ Rtiger Stand, Fehlermo¨ Glichkeiten Und Verbesserungsvorschla¨Ge

    J Lab Med 2004;28(4):317–325 ᮊ 2004 by Walter de Gruyter • Berlin • New York 2004/03304 Drug Monitoring und Toxikologie Redaktion: V. W. Armstrong Drug screening: Actual status, pitfalls and suggestions for improvement Drogenscreening: Gegenwa¨ rtiger Stand, Fehlermo¨ glichkeiten und Verbesserungsvorschla¨ge Wolf R. Ku¨ lpmann* pretierbar. In Wirklichkeit mu¨ ssen viele Aspekte beru¨ ck- sichtigt werden, um einen aussagekra¨ ftigen Befund zu Klinische Chemie, Medizinische Hochschule Hannover, erhalten, z.B. Pra¨ analytik, Spezifita¨ t und Empfindlichkeit Hannover, Germany des Verfahrens oder Qualita¨ tssicherung. Obwohl die Abstract mechanisierten Meßverfahren naturgema¨ ß quantitative Ergebnisse liefern, wird aufgezeigt, daß es sich in der Immunoassays for drug screening are often regarded as Regel empfiehlt, qualitative Befunde zu erstellen. Die procedures which are easily performed and interpreted. Verfahren erlauben aber im Gegensatz zu den auch fu¨r But in fact, many aspects have to be considered to diese Zwecke verwendeten Teststreifen die Entschei- obtain a meaningful result. Among these are sampling, dungsgrenze (cut-off) der jeweiligen Fragestellung anzu- specificity and sensitivity of assays, and quality assess- passen, z.B. bei akuter Intoxikation, chronischem Abusus ment. Although the mechanised procedures yield quan- oder U¨ berwachung des Drogenentzugs. titative results, there are good reasons to generally report Ein schwerwiegender Nachteil von Immunoassays zum qualitative findings. The mechanised procedures allow Nachweis von Amphetamin und a¨ hnlichen Substanzen, the adjustment of the cut-off concentration to the clinical Barbituraten, Benzodiazepinen und Opiaten besteht dar- setting, e.g. in the case of acute intoxication, chronic in, daß eine starke Kreuzreaktivita¨ t des Antiko¨ rpers abuse or drug withdrawal, an important advantage as gegenu¨ ber pharmakologisch wenig wirksamen Verbin- compared to test strips.