COSY type experiments exploring through-bond homonuclear correlations

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Peptides Faculty of Health and Medical Sciences University of Copenhagen Phone: +45 3533 6411; Fax:

1 COSY type experiments exploring through-bond homonuclear correlations Assistant Professor Kenneth Kongstad Bioanalytical Chemistry and Metabolomics Research Group Section for Natural Products and Peptides Faculty of Health and Medical Sciences University of Copenhagen Phone: ; Fax: Bioanalytical Chemistry and Metabolomics Research Group

2 Lecture outline ü Introduction to the second dimension ü Phase sensitive vs Absolute COSY ü DQF-COSY ü TOCSY Dias 2

3 Dias 3 2-Dimensional NMR (2D-NMR) 2D-NMR adds additional time domain to the experiment Preparation evolution mixing detection P, E and M is a single 90 pulse for a 1D 1 H NMR experiment

4 Dias 4 2-Dimensional NMR (2D-NMR) 2D-NMR adds additional time domain to the experiment ppm FT 0 F1 FT 8 8 F2 0 ppm

5 Dias 5 2-Dimensional NMR (2D-NMR) 2D-NMR adds additional time domain to the experiment Preparation evolution mixing detection Preparation is where equilibrium magnetization are pertubed Evolution is the time domain where the second dimension is created Mixing decide the type of 2Dexperiment Detection is the second time domain for direct detection of the FID

6 Dias 6 Simple 2-Dimensional NMR experiment Preparation evolution mixing detection Preparation is where equilibrium magnetization are pertubed Evolution is the time domain where the second dimension is created Mixing decide the type of 2Dexperiment Detection is the second time domain for direct detection of the FID

7 Dias 7 Vector-representation Longer t 1 than for (c)

8 Dias 8 Evolution Evolution time is varied systematically in order to obtain the second dimension

9 Dias 9 Variation of t 1 Systematical variation of t 1

10 Dias 10 Variation of t 1 generation of second dimension

11 Dias 11 2-Dimensional NMR (2D-NMR) 2D-NMR adds additional time domain to the experiment ppm FT 0 F1 FT 8 8 F2 0 ppm

12 Dias 12 F2 Result 2D-spectrum with frequency domain F2 along one axis and frequency domain F1 along the other. F1

13 Dias 13 Representation of 2D-spectra 2D-spectra usually displayed as stacked spectra or contour plots. Stacked spectrum Contour plot Cross peaks Diagonal peaks Diagonal peaks Cross peaks

14 Dias 14 Types of 2D-experiments A very large number of 2D-experiments have been developed over the years. The most commonly used are: HOMONUCLEAR EXPERIMENTS: ü COSY (COrrelation SpectroscopY) giving cross peaks due to scalar coupling. ü NOESY (Nuclear Overhauser Effect SpectroscopY) giving cross peaks for nuclei that are spatially close ü TOCSY (TOtal Correlation SpectroscopY) giving cross peaks between all hydrogens in a spin system ü J-resolved experiment ü Long-Range COSY, Relayed COSY, etc. (COSY variants) ü HOHAHA (TOCSY variant)

15 Dias 15 Types of 2D-experiments A very large number of 2D-experiments have been developed over the years. The most commonly used are: HETERONUCLEAR EXPERIMENTS: ü HSQC (Heteronuclear Single Quantum Coherence) giving cross peaks between 13 C and their direct bound hydrogens, i.e., 1 J C,H ü HMBC (Heteronuclear Multiple Bond Correlation) giving cross peaks due to 2 J C,H, 3 J C,H, 4 J C,H, etc. ü HETCOR, HSC, HMQC (HSQC analogs) ü COLOC (HMBC analogs)

16 Dias 16 The second dimension Homonuclear Heteronuclear F 1 ( 1 H) F 1 ( 13 C) F 2 ( 1 H) F 2 ( 1 H) Through bond coupling Through space coupling Chemical exchange

17 COSY Correlation SpectroscopY Cross peaks between protons that couple with each other 90 x 90 x t 1 t 2 Acquire several FIDs with increasing t 1 values. Dias 17 One spinsystem

18 COSY Correlation SpectroscopY Cross peaks between protons that couple with each other 90 x 90 x t 1 t 2 Acquire several FIDs with increasing t 1 values. Dias 18 Two spinsystems

19 Dias 19 Typical 2D homonuclear NMR experiments COSY: ü Absolute value COSY ü Phase sensitive COSY ü Double Quantum Filtered COSY TOCSY: ü TOCSY ü HOHAHA

20 Dias 20 Vector-representation

21 Dias 21 Absolute value COSY Advantages: Simple; well suited for automation Disadvantages: Poor resolution; no cross peak fine structure

22 Dias 22 Phase sensitive COSY DIAGONAL PEAKS In-phase doubledispersion lineshapes CROSS PEAKS Antiphase double absorption

23 Dias 23 DQF-COSY (Double-Quantum Filtered) Advantages ü Diagonal in antiphase absorption mode easier to see resonances close to the diagonal. ü Coupling constants from crosspeak fine structure ü Suppress peaks from singlet resonances

24 Dias 24 DQF-COSY (Double-Quantum Filtered) Gives coupling constants from cross peak fine structure Active couplings antiphase Passive couplings in phase Cross peaks due to small couplings or broad lines are more likely to be missing.

25 TOCSY Dias 25 Advantages: Correlations between all protons in spinsystem; high sensitivity; easy to get correlation to broad resonances Disadvantages: No cross peak fine structure

26 Dias 26 TOCSY COSY 90 x 90 x 90 x t 1 τ m Spinlock t 1

27 Dias 27 TOCSY COSY 90 x 90 x 90 x t 1 τ m Spinlock t 1

28 Dias 28 TOCSY 90 x t 1 τ m Spinlock Strong coupling Coherence transferred throughout spinsystem

High-Resolutio n NMR Techniques i n Organic Chemistry TIMOTHY D W CLARIDGE

High-Resolutio n NMR Techniques i n Organic Chemistry TIMOTHY D W CLARIDGE Foreword Preface Acknowledgements V VI I X Chapter 1. Introduction 1.1. The development of high-resolution NMR 1 1.2. Modern