C13 NUCLEAR MAGNETIC RESONANCE

C13 NUCLEAR MAGNETIC RESONANCE (NMR) , DEPT, COSY & NOESY

1. C 13 NUCLEAR MAGNETIC RESONANCE (NMR) , DEPT, COSY & NOESY Sujitlal Bhakta Department of Chemistry Ravenshaw University Cuttack, Odisha, 753 003

2. Introduction to C-13 NMR 13 C occurs naturally as 1.11% of total C and the NMR signal is weaker than 1 H. Fourier Transform NMR is used to collect a spectrum C 13 resonances occur from 0 to 220 ppm ( δ ). 13 C peaks are split by the attached hydrogens .

3. Nuclei with an odd mass or odd atomic number have "nuclear spin" (in a similar fashion to the spin of electrons). This includes   1 H and   13 C( butnot   12 C). The spins of nuclei are sufficiently different that NMR experiments can be sensitive for only one particular isotope of one particular element.   The NMR behaviour of   1 H and   13 C nuclei has been exploited by organic chemist since they provide valuable information that can be used to deduce the structure of organic compounds. These will be the focus of our attention.Since a nucleus is a Charged particle in motion, it will develop a magnetic field.   1 H and   13 C   have nuclear spins of 1/2 and so they behave in a similar fashion to a simple, tiny bar magnet.   In the absence of a magnetic field, these are randomly oriented but when a field is applied they line up parallel to the applied field, either spin aligned or spin opposed.   The   more highly populated state is the lower energy spin state spin aligned situation.   Two schematic   representations of these arrangements are shown below:   BASIC PRINCIPLES OF NMR

4. Nuclear Magnetic Resonance Nuclear spin m = g I h m - magnetic moment g - gyromagnetic ratio I - spin quantum number h - Planck’s constant m I is a property of the nucleus Mass # Atomic # I Odd Even or odd 1/2, 3/2, 5/2,… Even Even Even Odd 1, 2, 3 As an exercise determine I for each of the following 12 C, 13 C, 1 H, 2 H, 15 N .

5. Nucleus Spi n Quantum Number (I) Natural Abundance (%) Gyromagnetic Ratio (10-7 rad / T sec) Sensitivit y † ( % vs . 1H) Electric Quadrupule Momen t (Q) (e·1 24 c m 2) 1 H 2 H 13 C 15 N 19 F 31 P 1/2 1 1/2 1/2 1/2 1/2 99.9844 0.0156 1.108 0.365 100 100 26.7520 4.1067 6.7265 -2.7108 25.167 10.829 100.0 0.965 1.59 0.104 83.3 6.63 ————— 0.00277 ————— ————— ————— ————— Nuclear Magnetic Resonance

6. B o w w = g B o = n /2 p w - resonance frequency in radians per second, also called Larmor frequency n - resonance frequency in cycles per second, Hz g - gyromagnetic ratio B o - external magnetic field (the magnet) Apply an external magnetic field (i.e., put your sample in the magnet) z m m w Spin 1/2 nuclei will have two orientations in a magnetic field +1/2 and -1/2.

7. B o w z m m w +1/2 -1/2 Net magnetic moment

8. B o = 0 B o > 0 Randomly oriented Highly oriented B o Ensemble of Nuclear Spins N S Each nucleus behaves like a bar magnet.

9. The net magnetization vector z x y w w z x y M o - net magnetization vector allows us to look at system as a whole z x w one nucleus many nuclei

10. B o = 0 B o > 0 E D E Allowed Energy States for a Spin 1/2 System antiparallel parallel D E = g h B o = h n -1/2 +1/2 Therefore, the nuclei will absorb light with energy D E resulting in a change of the spin states.

11. Energy of Interaction D E = g h B o = h n The frequency, n , corresponds to light in the radiofrequency range when B o is in the Teslas. This means that the nuclei should be able to absorb light with frequencies in the range of 10’s to 100’s of megaherz. Note: FM radio frequency range is from ~88MHz to 108MHz. 77 Se, g = 5.12x10 7 rad sec -1 T -1 n = g B o /2 p

12. Nuclear Spin Dynamics z x y M o z x y M o z x y M o RF off RF on RF off Effect of a 90 o x pulse

13. Nuclear Spin Evolution z x y M o z x y M o w z x y Time x y RF receivers pick up the signals I

14. Free Induction Decay The signals decay away due to interactions with the surroundings. A free induction decay, FID, is the result. Fourier transformation, FT, of this time domain signal produces a frequency domain signal. FT Time Frequency

15. Spin Relaxation There are two primary causes of spin relaxation: Spin - lattice relaxation, T 1 , longitudinal relaxation . Spin - spin relaxation, T 2 , transverse relaxation. lattice

16. Nuclear Overhauser Effect Caused by dipolar coupling between nuclei. The local field at one nucleus is affected by the presence of another nucleus. The result is a mutual modulation of resonance frequencies. N S N S

17. Nuclear Overhauser Effect The intensity of the interaction is a function of the distance between the nuclei according to the following equation. I = A (1/r 6 ) I - intensity A - scaling constant r - internuclear distance 1 H 1 H r 1,2 1 2 1 H 3 r 1,3 r 2,3 Arrows denote cross relaxation pathways r 1,2 - distance between protons 1 and 2 r 2,3 - distance between protons 2 and 3 The NOE provides a link between an experimentally measurable quantity, I, and internuclear distance. NOE is only observed up to ~5Å.

18. Scalar J Coupling Electrons have a magnetic moment and are spin 1/2 particles. J coupling is facilitated by the electrons in the bonds separating the two nuclei. This through-bond interaction results in splitting of the nuclei into 2I + 1states. Thus, for a spin 1/2 nucleus the NMR lines are split into 2(1/2) + 1 = 2 states. 1 H 12 C 12 C 1 H Multiplet = 2nI + 1 n - number of identical adjacent nuclei I - spin quantum number

19. Scalar J Coupling The magnitude of the J coupling is dictated by the torsion angle between the two coupling nuclei according to the Karplus equation. C C H H H H q J = A + Bcos ( q ) + C cos 2 ( q) A = 1.9, B = -1.4, C = 6.4 q 3 J Karplus Relation A, B and C on the substituent electronegativity .

20. Torsion Angles Coupling constants can be measured from NMR data. Therefore, from this experimental data we can use the Karplus relation to determine the torsion angles, q. Coupling constants can be measured between most spin 1/2 nuclei of biological importance, 1 H, 13 C, 15 N, 31 P The most significant limitation is usually sensitivity, S/N .

21. Chemical Shift, δ The chemical is the most basic of measurements in NMR . The Larmor frequency of a nucleus is a direct result of the nucleus , applied magnetic field and the local environment . If a nucleus is shielded from the applied field there is a netreduction if the magnetic field experienced by the nucleus which results in a lower Larmor frequency. d is defined in parts per million, ppm. 13 C Chemical shifts are most affected by:  hybridization state of carbon  electronegativity of groups attached to carbon

22. TYPICAL CHEMICAL SHIFTS 190-220 d aldehydes, ketones 160-190 d esters, amides, carboxylic acids, acyl halides 110-160 d arenes , alkenes 50-110 d alkynes, sp 3 C attached to functional groups 0-50 d sp 3 C-Csp 3 , where 4 o >3 o >2 o >1 o

23. The zero point is defined as the position of absorption of a standard, tetramethylsilane (TMS): This standard has only one type of C and only one type of H. 13 C chemical shift

24. Chemical Shifts Electronegative groups attached to the  C-H  system decrease the electron density around the protons, and there is less shielding ( i.e.   deshielding ) so the chemical shift increases. 

25. 13 C NMR

26. Magnetic Anisotropy The word " anisotropic " means "non-uniform".  So magnetic anisotropy means that there is a "non-uniform magnetic field". Electrons in π systems ( e.g.  aromatics, alkenes, alkynes, carbonyls  etc .) interact with the applied field which induces a magnetic field that causes the anisotropy.  As a result, the nearby protons will experience 3 fields: the applied field, the shielding field of the valence electrons and the field due to the π system. Depending on the position of the proton in this third field, it can be either shielded (smaller d) or deshielded (larger d), which implies that the energy required for, and the frequency of the absorption will change.

27. Solvent 1H NMR Chemical Shift 13C NMR Chemical Shift Acetic Acid 11.65 ( 1 ) , 2.04 (5) 179.0 ( 1 ) , 20.0 (7) Acetone 2.05 (5) 206.7 (13) , 29.9 (7) Acetonitrile 1.94 (5) 118.7 ( 1 ) , 1.39 (7) Benzene 7.16 ( 1 ) 128.4 ( 3 ) Chloroform 7.26 ( 1 ) 77.2 ( 3 ) Dimethyl Sulfoxide 2.50 (5) 39.5 (7) Methanol 4.87 ( 1 ) , 3.31 (5) 49.1 (7) Methylene Chloride 5.32 ( 3 ) 54.00 (5) Pyridine 8.74 ( 1 ) , 7.58 ( 1 ) , 7.22 ( 1 ) 150.3 ( 1 ) , 135.9 ( 3 ) , 123.9 (5) Water (D 2 O) 4.8   NMR Solvent Signals   The chemical shifts (d)of solvent signals observed for 1 H NMR and   13 C NMR spectra Are listed in the following table. The multiplicity is shown in parentheses as 1 for singlet , 2 for doublet, 3 for triplet, etc. 

28. Solvent Chemical Shift of H 2 O (or HOD) Acetone 2.8 Acetonitrile 2.1 Benzene 0.4 Chloroform 1.6 Dimethyl Sulfoxide 3.3 Methanol 4.8 Methylene Chloride 1.5 Pyridine 4.9 Water (D2O) 4.8 Signals for water occur at different frequencies in  1 H NMR spectra depending on the solvent used. Listed below are the chemical shift positions of the water signal in several common solvents. Note that H 2 O is seen in aprotic solvents, while HOD is seen in protic solvents due to exchange with the solvent deuteriums .   NMR Water Signals

29. Predicting Chemical Shifts

30. Predicted Chemical Shifts of C a and C b

31. Chemical Shift Prediction with Functional Groups

32. Predicted 13 C Chemical Shifts m- Xylene

33. Use Base Value From Table 4.5

34. Carbon-13 Proton-Coupled Patterns

35. 13 C Off-resonance & Broadband decoupled spectra Broadband Off-resonance Off-resonance decoupling eliminates interactions of hydrogens on adjacent carbons. Broadband decoupling eliminates splitting of C by Hs attached to that C. However, proton decoupled (broadband) spectra are not split by H.

36. Spectrum at 75 MHz and 150 MHz

37. 13 C NMR (100 MHz, CDCl 3 ) δ 158.5, 157.9, 152.7, 149.6, 147.6, 140.8, 137.8, 131.0, 129.1, 128.8, 126.8, 124.6, 122.4, 121.0, 119.6, 118.0, 117.2, 115.0, 114.6, 112.5, 108.0, 75.0, 65.0, 15.0. C-13 NMR OF 8'-ethoxy-4-hydroxy-3'-nitro-2'-phenyl-2H,2'H-3,4'-bichromen-2-one

38. Summary There are three primary NMR tools used to obtain structural information Nuclear Overhauser effect - internuclear distances J Coupling - torsion angles Chemical shift - local nuclear environment ( Chemical exchange can also be monitored by NMR .)

39. Distortionless enhancement by polarization transfer ( DEPT ) spectra permit identification of CH 3 , CH 2 , and CH carbon atoms. DEPT 45 shows 1 o , 2 o ,and 3 o carbons. So any broadband peak not in DEPT 45 is 4 o . DEPT 90 shows only 3 o carbons. DEPT 135 shows 1 o and 3 o carbons as positive peaks and 2 o carbons as negative peaks . DEPT In DEPT, a second transmitter irradiates 1H during the sequence, which affects the appearanceof the 13Cspectrum. some 13C signals stay the same some 13C signals disappear some 13C signals are inverted

40. DEPT : D istortionless E nhancement by P olarization T ransfer Heteronuclear expt. Detection: 13 C Distinguish CH, CH2, CH 3 By suitable combination of  =45, 90 & 135 spectra All CH’s Only CH CH & CH 3 up CH 2 down

41. DEPT Spectra Quaternary carbons (C) do not show up in DEPT. CH and CH3 unaffected C and C=O nulled CH2 inverted

42. Simulated DEPT Spectra of Ethyl Phenylacetate Normal C-13 spectrum DEPT-45 DEPT-90 DEPT-135

43. COSY & NOESY COSY - Correlation spectroscopy Gives experimental details of interaction between hydrogens connected via a covalent bond NOESY - Nuclear Overhauser effect spectroscopy Gives peaks between pairs of hydrogen atoms near in space (1.5-5 Å ) (and not necessarily sequence)