Related Pergamon Titles of Interest BOOKS Tetrahedron Organic Chemistry Series: CARRUTHERS: Cycloaddition Reactions in Organic Synthesis GAWLEY & AUBE: Asymmetric Synthesis HASSNER & STUMER: Organic Syntheses Based on Name Reactions and Unnamed Reactions PAULMIER: Selenium Reagents & Intermediates in Organic Synthesis PERLMUTTER: Conjugate Addition Reactions in Organic Synthesis SIMPKINS: Sulphones in Organic Synthesis TANG & LEVY: Chemistry of C-Glycosides WILLIAMS: Synthesis of Optically Active Alpha-Amino Acids WONG: Enzymes in Synthetic Organic Chemistry JOURNALS BIOORGANIC & MEDICINAL CHEMISTRY BIOORGANIC & MEDICINAL CHEMISTRY LETTERS JOURNAL OF PHARMACEUTICAL AND BIOMEDICAL ANALYSIS TETRAHEDRON TETRAHEDRON: ASYMMETRY TETRAHEDRON LETTERS Full details of all Elsevier Science publications/free specimen copy of any Elsevier Science journal are available on request from your nearest Elsevier Science office. Modern NMR Techniques for Chemistry Research ANDREW E. DEROME* The Dyson Perrins Laboratory, University of Oxford, UK PERGAMON U.K. Elsevier Science Ltd, The Boulevard, Langford Lane, Kidhngton, Oxford 0X5 1GB, U.K. U.S.A. Elsevier Science Inc., 660 White Plains Road, Tarrytown, New York 10591-5153, U.S.A. JAPAN Elsevier Science Japan, Tsunashima Building Annex, 3-20-12 Yushima, Bunkyo-ku, Tokyo 113, Japan Copyright © 1987 Pergamon Press Ltd All Rights Reserved. No part of this publication may be reproduced, stored in a retrieval system or transmitted in any form or by any means: electronic, electrostatic, magnetic tape, mechanical, photocopying, recording or otherwise, without permission in writing from the publishers. First edition 1987 Reprinted 1987, 1988 (twice), 1990, 1991, 1993, 1995, 1997 Library of Congress Cataloguing in Publication Data Derome, Andrew E. Modem NMR techniques for chemistry research. (Organic chemistry series, v. 6) Includes index. I. Nuclear magnetic resonance spectroscopy. I. Title II. Series: Organic chemistry series (Pergamon Press); v. 6. QD96.N8D47 1987 543'.0877 86-20510 British Library Cataloguing in Publication Data Derome, Andrew E. Modem NMR techniques for chemistry research. — (Organic chemistry series, v. 6) I. Nuclear magnetic resonance spectroscopy I. Title II. Series 541.2'8 QD96.N8 ISBN 0-08-032514-9 (Hardcover) ISBN 0-08-032513-0 (Flexicover) Printed in Great Britain by BPC Wheatons Ltd, Exeter Foreword The development of commercially available high resolution NMR spectrometers in the 1950's provided the chemist with a new tool of enormous power. The direct relationship between spectral symmetry and the molecular symmetry of the sample enabled the routine solution of structural problems which previously required a level of intellectual power and chemical insight given to few. All this was achieved in the 100 milligram range, with instruments based on powerful electro- or permanent magnets. A second revolution has now occurred in the NMR field. This has come about through the development of rehable superconducting magnets coupled to the application of the pulse technique and its associated Fourier transform. Now dispersion and sensitivity have increased in leaps and bounds, so that sample size can be in the microgram range. Perhaps more important have been developments consequent on the pulse technique which permit enormously greater control and manipulation of the sample's magnetisation. As a result, the structural information now available to the chemist, through pulse NMR, is probably greater and more readily obtained than by any other single technique. In this book Andrew Derome takes the practising chemist through the practical aspects of this new method. Through this work a chemist, not trained as a physical NMR spectroscopist, can understand and use the enormously powerful tools for struc­ tural investigation available through this new generation of superconducting FT ma­ chines. It is strongly recommended to chemists and biologists both in academe and industry, who wish to reahse the full possibihties of the new wave NMR. Jack E. Baldwin, FRS The Dyson Perrins Laboratory Oxford University Andrew Ε, Derome. 1957-1991 The tragic death of Andy Derome earlier this year has robbed the field of chemical research of an outstanding teacher in the application of NMR spectroscopy to chemical problems. Throughout his brief but highly productive career he was able to bring together his knowledge of chemistry and his expertise in NMR to create the most powerful analytical techniques used by organic chemists. This book has served as a catalyst for the conversion of a multitude of chemists throughout the world into capable practitioners of modern NMR methods. As such it has significantly advanced the field of chemical research. Although he is no longer with us his magnum opus will, through its use by chemists, be a living memorial to his outstanding teaching ability. Jack E. Baldwin, FRS Dyson Perrins Laboratory University of Oxford August, 1991 Preface If you have just picked up this book with the thought of buying it, you may be asking yourself 'why go to all this trouble'. Here is a text which, although set in a qualitative framework, does spend a good deal of time discussing the physical principles behind NMR experiments; if you are a chemist or biologist you may shy away from such material. There is also a lot of experimental detail, which may seem superfluous if you are used to placing your spectroscopic problems in the hands of specialists. Presumably you are curious about the new developments in NMR, but maybe you would feel more comfortable with a survey which concentrated on spectral interpretation, leaving the technical details for somebody else. I believe there are several reasons why the applic­ ation of NMR to chemical and biological problems cannot be made properly using such a 'black box' approach, and I hope the following paragraphs will convince you of this. You can find a detailed description of what this book contains in the first chapter. Why is NMR such a useful technique? There are many reasons, but a central theme is that it can identify connections between entities. The gross features of chemical shift and signal intensity have, essentially, the same character as other spectroscopic methods such as infra-red absorption, and if that was all there was to NMR it would not outpace other methods to such a degree. But there is more: the fine structure in spectra, which arises from the coupling between nuclei, and various other interactions such as the nuclear Overhauser effect, depend on the relationships between nuclei; this is what gives NMR its special usefulness. Whether the aim is to probe the structure of an isolated, pure compound, or to measure proton-proton distances in a protein, or to extract the signal of a labelled metabolite from some biological soup, it is to those properties relating one nucleus with another that we turn. Over the last ten years or so the peculiar advantages of pulse NMR have come to be fully appreciated, leading to the development of many ingenious new ways to exploit these connections. Simply glancing at a list of experiments can be daunting; there are so many that it seems impossible to even begin to understand them. However, it is not so difficult. A few basic principles have been applied in various permutations; once these principles have been grasped any new experiment can be fitted easily into the overall picture. This is an important reason for making the effort to understand what is happening during pulse experiments; you will be better able to judge whether some new method is applicable to your particular problem. A nice feature of pulse NMR is that it is, on the whole, easier to understand than continuous wave NMR, even though the latter may be more familiar. In a pulse experiment the system spends most of its time evolving independently of outside stimuli, so there are less things to think of at once. Modern pulse NMR is performed exclusively in the Fourier transform mode. The reasons for this are set out in detail later in the text, but the use of the FT method is another important motivation for thinking about the mechanics of NMR experiments. Of course, it is useful to appreciate the advantages of the transform, and particularly the spectacular results which can be achieved by applying it in more than one dimension, but it is also essential to understand the limitations imposed by digital signal analysis. The sampling of signals, and their manipulation by computer, often limit the accuracy of various measurements of frequency and amplitude, and may even prevent the detection of signals altogether in certain cases. These are not diflftcult matters to understand, but they often seem rather abstract to newcomers to FT NMR. Even if you do not intend to operate a spectrometer, it is irresponsible not to acquire some familiarity with the interaction between parameters such as acquisition time and resolution, or repetition vii viii Preface rate, relaxation times and signal intensity. Many errors in the use of modern NMR arise because of a lack of understanding of its hmitations. Another reason for studying NMR, and especially its practical side, is that it is fun. Many chemical and, nowadays, biological projects culminate in an NMR experiment. After a year-long slog to synthesise some labelled substrate, it may only take a few days to perform the real experiment by NMR, whether it be following the mechanism of an organic reaction or watching a metabolic pathway inside whole cells. It is not fair that the excitement associated with such experiments should be reserved for specialist NMR operators: learn to do it yourself. The investment is the initial effort required to become familiar with a spectrometer (which may require a few late nights, unless you are very lucky), and the struggle to understand the experiments. The return is a broad under­ standing of the most important spectroscopic technique, and a refreshing and stimulating change from everyday lab. routine. Many people have helped during the preparation of this book, and I will now try to thank them; to anyone who has been inadvertently omitted I extend my apologies. First, I am indebted to Jane Maclntyre, for originally persuading me that the project was possible. Several of my colleagues in the Dyson Perrins and Inorganic Chemistry Laboratories have read the text, from the diverse perspectives of graduate student, NMR operator, electronics engineer, university lecturer and postdoctoral researcher, and pro­ vided many helpful suggestions. My sincere thanks to: Barbara Domayne-Hayman, Elizabeth McGuinness, Dermot O'Hare, Mike Robertson, Mike Robinson, Chris Schofield and Nick Turner. I also especially have to thank Elizabeth McGuinness and Tina Jackson for bearing the brunt of the NMR requirements of hundreds of research workers in the Oxford chemistry departments, thus leaving me sufficiently undisturbed to work on the text. Mike Robertson carried out extensive modifications to a commercial instrument to permit some of the more unusual experiments to be performed, while Tina Jackson provided technical assistance. A number of people have been kind enough to contribute figures, or to permit me to perform devihsh experiments on their compounds. I thank, from the Dyson Perrins Laboratory, Oxford: John Brown (for Figures 9.6, 9.7), George Fleet (for compound 1, Chapters 8-10), Nick Turner (for compound 6, Chapter 5) and Rob Young (for compound 6, Chapter 8); from the Inorganic Chemistry Laboratory, Oxford: Dermot O'Hare (for Figures 8.45, 9.10, 10.11); from the Physical Chemistry Laboratory, Oxford: Chris Bauer (for Figure 7.19), Ray Freeman (for Figure 8.21) and Peter Hore (for Figure 2.23); from the Biochemistry Department, Oxford: Jonathan Boyd (for Figure 8.36); from the University Chemical Laboratory, Cambridge: James Keeler (for Figures 8.24, 10.13); from the Ruder Boskovic Institute, Zagreb: Dina Keglevic (for compound 2, Chapter 8). Oxford Instruments Ltd. and Bruker Spectrospin also provided a number of figures, which are acknowledged in the relevant captions. Figure 5.10 was reproduced with permission from: J. H. Noggle and R. E. Schirmer, The Nuclear Overhauser Effect - Chemical Applications, Academic Press. 1972. In common with the other books in the Organic Chemistry series, this text was reproduced photographically from camera-ready copy. However, in order to achieve an acceptable appearance, I typeset the material myself on a Monotype Lasercomp phototy- pesetter at Oxford University Computing Service. I thank the Computing Service for access to this device, and Catherine Griffin for advice on its use. Christine Palmer, of the Inorganic Chemistry Laboratory, Oxford, miraculously produced the hne drawings from my semi-legible sketches, often at high speed; I thank her for this vital contribution. Final paste-up of the text was performed at Pergamon, under the expert supervision of Colin Drayton, who also managed to avoid making me feel bad even when the whole project was months behind schedule. Andrew Derome Introduction Nuclear resonances are affected by a variety of weak interactions between the nuclei and the electrons of molecules, between nuclei within the molecules and between nuclei in neighbouring molecules. If these interactions can be disentangled and interpreted, they are found to contain an extraordinarily rich mine of information about the structures and conformations of the molecules of the sample, about interactions between molecules and about molecular motion. All this is made possible by the very long relaxation times that characterise NMR spectra of spin ^2 nuclei in mobile liquids; line widths of 0.1 Hz or less at 500 MHz are by no means uncommon. This means that even resonances which lie very close together can be resolved. High resolution spectra, even of molecules of modest molecular weight, are therefore often quite complicated, but because the interactions are weak, cross terms are not very important if the measurements are made in strong magnetic fields, and the spectra are relatively easy to inteφret, It is not surprising, therefore, that high resolution NMR has become such an indispensable tool for the organic chemist. As the size of the sample molecules becomes greater, the complexity of the NMR spectra increases rapidly, lines overlap, and their interpretation becomes more difficult. At this stage much more sophisticated sequences of experiments must be made, the analysis of which can be technically demanding. Here the organic chemist is faced with a dilemma. How much effort should he or she invest in grappling with the not inconsiderable technicalities of the very powerful NMR techniques now available? It is certain that without some understanding of these methods he may not even realise how NMR can help him solve his particular problems, and even if he thinks help is available, he must beware of using NMR as an imperfectly understood black box. Most organic chemists to-day need to understand what NMR can do for them, and in this book Dr Derome sets out the principles and provides a practical guide to the use of NMR spectroscopy in terms accessible to organic chemists; it is a long journey but one which will be well worth while. The reader who perseveres with the book will be considerably enlightened. I hope the reader will also be sufficiently fired by the enthusiasm running through the text to read at some later stage the further references provided and achieve still greater depths of understanding of these elegant experiments. Sir Rex Richards, FRS University of Oxford xvn 1 What This Book Is About LI INTRODUCTION Figure 1.1 is a proton NMR spectrum of cholesteryl acetate, run at 60 MHz on a continuous wave spectrometer. During the sixties and early seventies, a period when much research into compounds of this type was carried out, such a spectrometer would have been regarded as a top rank research instrument. It is a remarkable reflection on the power of NMR to illuminate chemical problems (and on the prowess of those who pioneered its use) that the availability of spectra even as crude as this revolutionised organic chemistry. 4.5 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 Nowadays we can expect much more from our spectrometers (Figure Figure 1.1 A low-field, continuous wave 1.2), with instruments operating at almost ten times the frequency (and NMR spectrum. costing at least thirty times as much). The improvements in this spectrum are obvious, but the point I wish to make by showing it to you is perhaps less so. It is not good enough. The intrinsic limitations of NMR are such that, however strenuous our eñOrts to improve spectrometer technology, we will not be able to interpret the proton spectra of relatively simple compounds at sight. This will remain so into the foreseeable future, barring some entirely unexpected development in magnet construction. It is worth bearing in mind that, while the switch from electromagnets to super conducting magnets allowed an immediate jump in field strength of a factor of 3-4, further eñOrts in superconducting magnet design over almost twenty years have resulted in only another doubling of routinely available field. What This Book Is About ' ' I ' ' ' ' I ' ' ' I ' ' I ' ' 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 .5 0.0 PPM Figure 1.2 The same sample as for Fi­ gure I.I, but run at 500 MHz in Fourier We cannot rely on progress in instrument design to solve our spec­ transform mode. troscopic problems for us, but fortunately we do not need to do so. In parallel with technological development of NMR spectrometers, there has been an astonishing growth in our understanding of the properties of nuclear spin systems. Since about 1980, this has reached some kind of critical point, with the sudden development of a wide range of new experimental techniques. Everyone involved in chemistry research will certainly be aware by now that something unusual is happening to NMR, but still the impact of these developments has been fairly restricted. The technical nature of most NMR papers, and their spread through an unusual range of chemical, physical and biological journals, means that the non-specialist has little hope of keeping abreast of developments. In the teaching of NMR to students of chemistry, it sometimes seems that pulsed FT NMR has yet to supplant CW NMR in many courses pitched at organic chemists; modern NMR methods are almost never treated. This cannot remain so for much longer, because understanding how to use NMR to solve problems is fundamental to success in chemical research. In this book I hope to provide an introduction to modern NMR accessible to the non-specialist. It is intended to be self-contained, aside from some basic background knowledge requirements described in the next section. The reader I have in mind is the advanced student or beginning research worker, or those with more experience who wish to learn about