Analytical and Chromatographic Techniques in Radiopharmaceutical Chemistry

Analytical and Chromatographic Techniques in Radiopharmaceutical Chemistry

Edited by Donald M. Wieland Michael C. Tobes Thomas J. Mangner

With 115 Figures

Spri nger -Veri ag New York Berlin Heidelberg Tokyo

Donald M. Wieland Division of Nuclear Medicine

Department of Internal Medicine

The University of Michigan

Ann Arbor, MI48109

USA

Thomas J. Mangner Division of Nuclear Medicine

Department of Internal Medicine The University of Michigan Ann Arbor, MI48109 USA

Michael C. Tobes Division of Nuclear Medicine

Department of Internal Medicine

The University of Michigan

Ann Arbor, MI48109

USA

Library of Congress Cataloging in Publication Data Main entry under title: Analytical and chromatographic techniques in radio­pharmaceutical chemistry.

Updated and expanded versions of presentations at a symposium held June 4, 1984, in Los Angeles, Calif., under the sponsorship of the Radiopharmaceutical Science Council of the Society of Nuclear Medicine.

Includes bibliographies and index. 1. Radiopharmaceuticals--Analysis-Congresses.

2. Thin layer chromatography-Congresses. 3. Liquid chromatography-Congresses. 4. Chemistry, Analytic­Congresses. I. Wieland, Donald M. II. Tobes, Michael C. III. Mangner, Thomas J. IV. Society of Nuclear Medicine (1953- ). Radiopharmaceutical Science Council. [DNLM: 1. Chemistry, Analytical-methods­congresses. 2. Chromatography, High Pressure Liquid­congresses. 3. Chromatography, Thin Layer--congresses. 4. Radiochemistry-methods--congresses. QD 605 A532] RS190.R34A53 1985 615.84 85-14707

© 1986 by Springer-Verlag New York Inc.

Softcover reprint of the hardcover 1 st edition 1986

All rights reserved. No part of this book may be translated or reproduced in any form without written permission from Springer-Verlag, 175 Fifth Avenue, New York, New York 10010, U.S.A. The use of general descriptive names, trade names, trademarks, etc., in this publication, even if the former are not especially identified, is not to be taken as a sign that such names, as understood by the Trade Marks and Merchandise Marks Act, may accordingly be used

freely by anyone.

Typeset by BiComp, Inc., York, Pennsylvania.

987 6 5 4 3 2

ISBN-13: 978-1-4612-9331-6 e-ISBN-13: 978-1-4612-4854-5 DOl: 1 0.1 007/978-1-4612-4854-5

Foreword

In 1906, Michael T. Sweet first developed the chromatographic method by using an adsorbant to separate pigments. Since that time, the technological advances in TLC and HPLC have brought about new definitions of purity in parallel with the advances. Radiopharmaceutical chemistry is especially dependent on the chromat­ographic technique because of the relatively small amount of material in most radiopharmaceuticals-often so small that the usual physical methods of analytical chemistry cannot be used. As a result, this collection of papers represents the key to successful radiopharmaceutical development by setting the standard for the pres­ent-day definition of radiochemical purity.

William C. Eckelman, Ph.D. Diagnostics Associate Director The Squibb Institute for Medical Research New Brunswick, New Jersey

Preface

The chapters herein are updated and expanded versions of presentations that the authors made at a symposium held on June 4, 1984 in Los Angeles, California under the sponsorship of the Radiopharmaceutical Science Council of the Society of Nuclear Medicine. All manuscripts were refereed.

The intent of the symposium organizers was to enlist participants who work on a day-to-day basis with the analytical and chromatographic techniques to be discussed at the symposium. We feel confident that this distillation of hands-on experience will be of value to graduate students as well as experienced researchers in radio­pharmaceutical chemistry and related fields which use radiotracer methodology.

The short history of radiopharmaceutical chemistry has been marked by vivid examples of the value of conscientious use of analytical and chromatographic tech­niques. Nearly a decade ago, radio-TLC revealed the presence of a radioactive "impurity" in preparations of the adrenal cortex imaging agent I-131-19-iodocho­lesterol. Identification of this "impurity" showed in fact that it was the active agent I-131-6J3-iodomethyl-19-norcholesterol. The recent renaissance in Tc-99m radio­pharmaceuticals, especially the cationic heart agents, has been accompanied by successful application of reverse phase radio-HPLC to purity analyses of these agents. Application of similar radio-HPLC techniques to clinical mainstays such as Tc-99m bone agents, many of which were first synthesized in the "HPLC-Iess" days of the early 1970s, has revealed that certain of these agents are complex mixtures. Just as radio-HPLC was being embraced as the definitive technique for purity confirmation, a 1984 report using fluorine nuclear magnetic resonance re­vealed that F-18-2-fluorodeoxyglucose, a radiotracer of major importance in de­termining regional glucose metabolism by positron emission tomography, was in certain cases contaminated with varying amounts of its isomer, F-18-2-fluorodeoxy-

vi i i Preface

mannose. To date no radio-HPLC system has been reported that distinguishes these fluoro isomers.

The historical lesson is clear-no single analytical technique should be relied on. Working at nanogram levels, radiopharmaceutical chemists have always had cause to be light sleepers. Hopefully, reading this book will keep them up all night.

Donald M. Wieland, Ph.D. Michael C. Tobes, Ph.D. Thomas J. Mangner, Ph.D.

Acknowledgments

The editors and the Radiophannaceutical Science Council would like to thank the speakers, panel participants, poster presentors, and commercial exhibitors who participated in the symposium. A special thanks to Joanna Fowler of Brookhaven National Laboratories for moderating the panel discussion and to Richard Cham­berlain of the Society of Nuclear Medicine Central Office for organizational assis­tance. The editors are grateful to Linder Markham for secretarial assistance.

Research at the University of Michigan Medical Center in Radiophannaceutical Chemistry and Nuclear Medicine is supported by the Department of Internal Med­icine and the following grants from the National Institutes of Health: Contract No. HL-27555, NCI Training Grant, Contract No. 5-T32-CA09015-09 and the U.S. Department of Energy, Contract No. DE-AC02-76EV02031.

Contents

THIN-LAYER CHROMATOGRAPHY

Instrumental Evaluation of Thin-Layer Chromatograms

Colin F. Poole, Hal T. Butler, Myra E. Coddens, and Sheila A. Schuette 3

Introduction 3 Sample Application 4 Mode Selection for Scanning Densitometry 10 Instrumentation for Scanning Densitometry 13 Instrument Parameters that Affect the Performance of Slit-Scanning Densitometers 16 Protocol for Measuring the Sensitivity of a Slit-Scanning Densitometer 18 Qualitative Sample Identification by HPTLC and Scanning Densitometry 23 Quantitation of Separated Components by Scanning Densitometry 28 Reagents Used to Enhance Fluorescence of Organic Compounds on Thin-Layer Plates 32 Radiochromatography on Thin-Layer Plates 33 Conclusions 35 References 35

2 Radioanalytical Techniques: ITLC, TLC, Mini-Columns, and Electrophoresis

Alan P. Carpenter, Jf.

Introduction 39 ITLC 40 Radio-TLC 45 Mini-Columns 56

39

xii Contents

Electrophoresis 60 Conclusions 68 Future Considerations 68 References 68

3 Radio-Thin-Layer Chromatogram Imaging Systems-Performance and Design

Sheryl J. Hays

Introduction 71 Instrumentation 72 Resolution and Efficiency 73 Performance and Operation of the TLC Imaging Systems 74 Conclusion 75 References 77

4 Detection of Radiochromatograms and Electropherograms with Position­Sensitive Wire Chambers

Heinz Filthuth

Introduction 79 The TLC Linear Analyzer 81 Data Acquisition System 83 Multiplate Detector 84 Performance of the Linear Analyzer 85 Quantitative Measurements 86 Experience with the Linear Analyzer 90 Determination of the Radiochemical Purity of Radiopharmaceuticals 98 Summary 98 References 99

HIGH PRESSURE LIQUID CHROMATOGRAPHY

5 Components for the Design of a Radio-HPLC System

Adrian D. Nunn and Alan R. Fritzberg

Introduction 103 Columns and Packings 104 Components of a Radio-HPLC System 108 Conclusion 122 References 123

71

79

103

Contents xiii

6 Overall Radio-HPLC Design

Chester A. Mathis, Reese M. Jones, and Joseph H. Chasko

Introduction 125 Components of the Basic System 128 A More Complex Radio-HPLC System 130 Additional Considerations 137 A Specialized HPLC System 140 Summary 147 References 147

7 Quantitation of Radiolabeled Molecules Separated by High Pressure Liquid Chromatography

Michael J. Kessler

Introduction 149 Detection of HPLC-Separated Radioactive Compounds 150 Detector Design 154 Considerations in Use of Detector 158 Applications 163 Conclusion 167 References 167

8 Flow Detector Designs: Build Your Own or Buy?

Richard D. Hichwa

Introduction 171 Requirements 171 Types of Flow Cells 175 Data Acquisition and Analysis Considerations 177 Conclusions 178 References 179

APPLICA liONS

9 Radio-HPLC: Application to Organics and Metal Chelate Chemistry

Alan R. Fritzberg and Adrian D. Nunn

Introduction 183 Stability Under HPLC Conditions 188 Tc-N2S2 Chelates 188

125

149

171

183

Use of HPLC in Stereochemical Studies of Tc and Re Penicillamine Complexes 200

xiv Contents

HPLC in the Development and Analysis of HIDAs 201 Phosphines 204 Phosphonates 205 Conclusions 206 References 207

10 Concepts and Techniques Used in Metabolic Tracer Studies

Jorge R. Barrio, Randy E. Keen, Diane C. Chugani, Gerald Bida, Nagichettiar Satyamurthy, and Michael E. Phelps 213

Introduction 213 Requirements of Analytical Techniques for Metabolic Studies 214 Applications in Tracer Kinetic Models 221 Conclusions 227 References 227

11. Development of No-Carrier-Added Radiopharmaceuticals with the Aid of Radio-HPLC

D. Scott Wilbur

Introduction 233 Radio-HPLC 234 Scaling Radiolabeling Reactions to nca Levels 236 Identification of the nca Radiolabeled Compound 237 Examples of nca Radiolabeling Experiments 238 Conclusions 245 References 248

12 From Cyclotron to Patient via HPLC

Michael R. Kilbourn, Michael J. Welch, Carmen S. Dence, and Keith R. Lechner

Introduction 251 HPLC Systems 251 Columns 253 Solvent Systems 255 Dedicated vs General Instruments 256 Detectors 257 Pre-Columns and Filters 258 Is HPLC Truly Necessary? 258 Conclusions 259 References 259

233

251

Contents xv

13 Potential Artifacts in the Chromatography of Radiophannaceuticals

Thomas J. Mangner

Introduction 261 Choice of Analytical Methods 262 Radiochromatography 266 Potential Artifacts/Inconsistencies in Radiochromatography 270 Illustrative Examples 274 Summary 276 References 276

14 HPLC of Radiolabeled Antibodies

261

Donald J. Hnatowich 279

Introduction 279 Theory 280 An HPLC System for the Analysis of Radiolabeled Antibodies 283 Applications 285 Discussion 290 Conclusions 291 References 292

Index 295

Contributors

Jorge R. Barrio, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Gerald Bida, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Hal T. Butler, Department of Chemistry, Wayne State University, Detroit, MI 48202, USA

Alan P. Carpenter, Jr., Division of Analytical Chemistry, Radiopharmaceutical Research, Dupont/New England Nuclear Corporation, North Billerica, MA 01845, USA

Joseph H. Chasko, Donner Laboratory, Lawrence Berkeley Laboratory, University of California, Berkeley, CA 94720, USA

Diane C. Chugani, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Myra E. Coddens, Department of Chemistry, Wayne State University, Detroit, MI 48202, USA

Carmen S. Dence, Division of Radiation Sciences, The Edward Mallinckrodt In-stitute of Radiology, Washington University, St. Louis, MO 63110, USA

Heinz Filthuth, Laboratorium Prof. Dr. Berthold, D-7547 Wildbad 1, West Germany

Alan R. Fritzberg, NeoRx Corporation, Seattle, WA 98119, USA

Sheryl J. Hays, Warner-Lambert Company, Pharmaceutical Research Division, Ann Arbor, MI 48105, USA

xviii Contributors

Richard D. Hichwa, Division of Nuclear Medicine, University of Michigan Medical Center, Ann Arbor, MI 48109, USA

Donald J. Hnatowich, Department of Nuclear Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA

Reese M. Jones, Donner Laboratory, Lawrence Berkeley Laboratory, University of California, Berkeley, CA 94720, USA

Randy E. Keen, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Michael J. Kessler, Radiomatic Instruments & Chemical Co., Tampa, FL 33611, USA

Michael R. Kilbourn, Division of Radiation Sciences, The Edward Mallinckrodt Institute of Radiology, Washington University, St. Louis, MO 63110, USA

Keith R. Lechner, Division of Radiation Sciences, The Edward Mallinckrodt In­stitute of Radiology, Washington University, St. Louis, MO 63110, USA

Chester A. Mathis, Donner Laboratory, Lawrence Berkeley Laboratory, University of California, Berkeley, CA 94720, USA

Thomas J. Mangner, Division of Nuclear Medicine, University of Michigan Medical Center, Ann Arbor, MI 48109, USA

Adrian D. Nunn, Division of Radiopharmaceutical Research and Development, The Squibb Institute for Medical Research, New Brunswick, NJ 08903, USA

Michael E. Phelps, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Colin F. Poole, Department of Chemistry, Wayne State University, Detroit, MI 48202, USA

Nagichettiar Satyamurthy, Division of Nuclear Medicine and Biophysics, UCLA School of Medicine, Los Angeles, CA 90024, USA

Sheila A. Schuette, Department of Chemistry, Wayne State University, Detroit, MI 48202, USA

Michael J. Welch, Division of Radiation Sciences, The Edward Mallinckrodt In­stitute of Radiology, Washington University, St. Louis, MO 63110, USA

D. Scott Wilbur, NeoRx Corporation, Seattle, WA 98119, USA