Organic Structure Analysis Professor Marcel Jaspars

Aim This course aims to extend student’s knowledge and experience with nuclear magnetic resonance (NMR) and mass spectroscopy (MS), by building on the material taught in the 3rd Year Organic Spectroscopy course, and also to develop problem solving skills in this area.

Learning Outcomes By the end of this course you should be able to: Assign 1H and 13C NMR spectra of organic molecules. Analyse complex first order multiplets. Elucidate the structure of organic molecules using NMR and MS data. Use data from coupling constants and NOE experiments to determine relative stereochemistry. Understand and use data from 2D NMR experiments.

Synopsis A general strategy for solving structural problems using spectroscopic methods, including dereplication methods. Determination of molecular formulae using NMR & MS Analysis of multiplet patterns to determine coupling constants. Single irradiation experiments. Spectral simulation. The Karplus equation and its use in the determination of relative stereochemistry in conformationally rigid molecules. Determination of relative stereochemistry using the nuclear Overhauser effect (nOe). Rules to determine whether a nucleus can be studied by NMR & What other factors must be taken into consideration. Multinuclear NMR-commonly studied heteronuclei. Basic 2D NMR experiments and their uses in structure determination.

Books Organic Structure Analysis, Crews, Rodriguez and Jaspars, OUP, 2009 Spectroscopic Methods in Organic Chemistry, Williams and Fleming, McGraw-Hill, 2007 Organic Structures from Spectra, Field, Sternhell and Kalman, Wiley, 2008 Spectrometric Identification of Organic Compounds, Silverstein, Webster and Kiemle, Wiley, 2007 Introduction to Spectroscopy, Pavia, Lampman, Kriz and Vyvyan, Brooks/Cole 2009

Four Types of Information from NMR

1H NMR Chemical Shifts in Organic Compounds

13C NMR Chemical Shifts in Organic Compounds

5 Minute Problem #1 MF = C6H12O Unsaturated acyclic ether

Six Simple Steps for Successful Structure Solution Get molecular formula. Use combustion analysis, mass spectrum and/or 13C NMR spectrum. Calculate double bond equivalents (DBE). Determine functional groups from IR, 1H and 13C NMR Compare 1H integrals to number of H’s in the MF. Determine coupling constants (J’s) for all multiplets. Use information from 3. and 4. to construct spin systems (substructures) Assemble substructures in all possible ways, taking account of DBE and functional groups. Make sure the integrals and coupling patterns agree with the proposed structure.

Double Bond Equivalents CaHbOcNdXe [(2a+2) – (b-d+e)] DBE = 2 C2H3O2Cl =

Tabulate Data Shift (ppm) Int. Mult (J/Hz) Inference 6.48 1H dd, 14, 7 4.17 d, 14 3.97 d, 7 3.69 2H t, 7 1.65 quint, 7 1.42 sext, 7 0.95 3H

Solution

Shift Prediction Prediction

THE PROCESS OF STRUCTURE ELUCIDATION Organic Structure Analysis, Crews, Rodriguez and Jaspars

Dereplication Dediscovery

Dereplication

Dereplication

Determining the Molecular Formula Using NMR and MS Data DEPT-135 Organic Structure Analysis, Crews, Rodriguez and Jaspars

Determining the Molecular Formula Using NMR and MS Data

Determining the Molecular Formula Using NMR and MS Data

MS Errors Experimental accurate mass measurement (from MS) was 136.1256 suggesting C10H16 is the correct formula. The error between calculated and experimental mass is: 136.1256 - 136.1248 = 0.0008 = 0.8 mmu Formula dbe Accurate mass C10H16 3 136.1248 C9H12O 4 136.0885 C8H8O2 5 136.0522 C7H4O3 6 136.0159 C9H14N 3.5 136.1123 C8H12N2 136.0998

Molecular Formula Calculators James Deline MFCalc

Isotope Ratio Patterns: C100H200 For 12Cm13Cn 1403.6 1404.6

ThermoFinnigan LTQ Orbitrap, Determining molecular formulae by HR-ESI/MS ThermoFinnigan LTQ Orbitrap, Xcalibur software examples from MChem group practicals 2009 (Rainer Ebel)

[M+H]+

[M+Na]+

Analysis of isotope patterns experimental calculated “monoisotopic peak” (mainly 12C713C1H935Cl14N16O2)

5 Minute Problem #2 Al Kaloid, an Honours Chemistry Student at Slug State University has synthesised either A or B below. He is uncertain which one it is but he’s tabulated the 13C NMR shifts and their multiplicities. Can you help Al by determining which one it is? A B

Answer Base Values:

ChemDraw

The NMR effect

Spin-spin coupling (splitting) No coupling Coupling

Spin-spin coupling (splitting)

Origin of spin-spin coupling

Coupling in ethanol

Coupling is mutual

Coupling in ethanol To see why the methyl peak is split into a triplet, let’s look at the methylene protons (CH2). There are two of them, and each can have one of two possible orientations- aligned with, or against the applied field This gives rise to a total of four possible states: Hence the methyl peak is split into three, with the ratio of areas 1 : 2 : 1

Coupling in ethanol Similarly, the effect of the methyl protons on the methylene protons is such that there are 8 possible spin combinations for the three methyl protons: The methylene peak is split into a quartet. The areas of the peaks have the ratio of 1:3:3:1.

Pascal’s triangle n relative intensity multiplet 0 1 singlet 1 1 1 doublet 2 1 2 1 triplet 3 1 3 3 1 quartet 4 1 4 6 4 1 quintet 5 1 5 10 10 5 1 sextet 6 1 6 15 20 15 6 1 septet

Coupling patterns

First Order In CH3CH2OH we could explain coupling by the n + 1 rule, this is called 1st order coupling

Second Order Like CH3CH2OH expect 7 lines but get many more. Dn/J < 6

Common Coupled Spin Systems

Common Coupled Spin Systems

Complex 1st Order Spin Systems

Iterative application of the n + 1 rule

5 Minute Problem #3. Given the 1H NMR chemical shifts and coupling constants for allyl alcohol, explain the observed spectrum (OH peak omitted)

A doubled quartet (dq)

What about this?

ddt

Spin Simulation Real Spectrum

Pro-R and Pro-S 1

Homotopic, Enantiotopic, Diastereotopic

Methyl groups

Chemical Equivalence/Magnetic Non Equivalence

What is going on?

Result Expect:

Using Coupling Constants

Glucose

Glucose

5 Minute Problem #4 Work out which of d 2.1 and d 2.5 is equatorial and which is axial. Also work out the 3 dihedral angles for d 2.1, d 2.5, d 2.8, d 6.8. There are also peaks at: 6.80, 1H, d, J = 0.5 Hz; 1.95, 3H, s; 0.93, 9H, s. 30 Hz

Solution

Removing Couplings Changing Solvents da ≠ db Each coupled to Hc Jab ≠ Jac CDCl3 Ha, dd Jab ≠ Jac C6D6 da = db Coupled to Hc Jab = Jac Ha, t Jab = Jac

Removing Couplings Spin decoupling Coupling due to Ha is removed See Hb, Hc at db, dc With mutual Jbc CDCl3 irradiated Signal due to Ha disappears CDCl3 Irradiate at 4.11 ppm

Spin Decoupling OFF A↓ X ↓ A↓ X↑ A↑ X↓ A↑ X↑

Spin Decoupling Two spins, A (nA), X (nX) with JAX Irradiate nX with RF power, A loses coupling due to X ON A↓ X ↑ ↓ A↑ X↑ ↓ Average of X ↑ and X ↓

Nuclear Overhauser Effect

Size of NOE

Effect of NOE on 13C NMR 10/5/9 3 8 2 6 4 1 7

13C – 1H NOE at equilibrium (small molecule) ● C↑H ↓ ●●●● C↓H ↑ ●●●●● C↑H ↑

13C – 1H NOE irradiation on H ●● C↓H↓ C ●●● C↑H ↓ Hsat Hsat ●● C↓H ↑ C ●●● C↑H ↑

13C – 1H NOE irradiation on H left ON ●● C↓H↓ C ●●● C↑H ↓ Hsat Hsat ●● C↓H ↑ C ●●● C↑H ↑

13C – 1H NOE result ● C↓H↓ C ●●●● C↑H ↓ Hsat Hsat ● C ●●●● C↑H ↑

1H-1H NOE example H1 H2O H4 H3

1H – 1H NOE at equilibrium (small molecule) S↓I↓ ●● ●● S↑I ↓ S↓I ↑ ●●●● S↑I ↑ nI nS

1H – 1H NOE irradiation on S ● S↓I↓ W1S (sat) W1I ●●● ● S↑I ↓ S↓I ↑ W1I W1S (sat) ●●● S↑I ↑ nI nS

1H – 1H NOE irradiation on S left ON ● S↓I↓ W1S (sat) W1I ●●● ● S↑I ↓ W1I W1S (sat) ●●● S↑I ↑

1H – 1H NOE result ½ S↓I↓ W1S (sat) W1I ●●●½ ½ S↑I ↓ W1I W1S (sat) nI nS

NOE 3D example

NOE 3D example

Events Accompanying Resonance Organic Structure Analysis, Crews, Rodriguez and Jaspars

ONE-PULSE SEQUENCE Organic Structure Analysis, Crews, Rodriguez and Jaspars

ONE-PULSE SEQUENCE (90o)x 1H Preparation Detection Organic Structure Analysis, Crews, Rodriguez and Jaspars

Fourier Transformation FT

Relaxation and Peak Shape

Rotational Correlation Time tc wo = 2pno

Nuclear spin Example Atomic mass Atomic number Spin, I 13C, 1H, 17O, 15N, 3H Odd Odd or Even 1/2, 3/2, 5/2 etc 12C, 16O Even 2H, 14N 1, 2, 3 etc 6 1 8 7 1 6 8 1 7

Receptivity 29Si 4.7% -5.32 13C 1.1% 6.73 1H 100% 26.75 Nucleus C Relative g Receptivity Relative receptivity 29Si 4.7% -5.32 13C 1.1% 6.73 1H 100% 26.75

Multinuclear NMR

15N NMR Shifts

31P NMR Shifts

Coupling

Effect of 31P on 1H NMR

Effect of 31P on 1H NMR

Effect of 31P on 13C NMR 5 4 3 1

The 2nd Dimension

BASIC LAYOUT OF A 2D NMR EXPERIMENT Organic Structure Analysis, Crews, Rodriguez and Jaspars

How a 2D NMR experiment works Contour plot n is the number of increments Organic Structure Analysis, Crews, Rodriguez and Jaspars

TYPES OF 2D NMR EXPERIMENTS AUTOCORRELATED Homonuclear J resolved 1H-1H COSY TOCSY NOESY ROESY INADEQUATE CROSS-CORRELATED Heteronuclear J resolved 1H-13C COSY HMQC HSQC HMBC HSQC-TOCSY Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY & & Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – DEPT DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA A B C D E F dC f e d’ d c b a Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA ATOM dC (ppm) DEPT dH (ppm) A 131 CH 5.5 B 124 5.2 C 68 4.0 D 42 CH2 3.0 2.5 E 23 CH3 1.5 F 17 1.2 Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HSQC DATA diastereotopic protons Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – COSY DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – COSY DATA ATOM dC (ppm) DEPT dH (ppm) COSY (HH) A 131 CH 5.5 b, c, d/d’, f B 124 5.2 a, d/d’, f C 68 4.0 a, d/d’, e D 42 CH2 3.0 2.5 a, b, c, d, e E 23 CH3 1.5 c, d/d’ F 17 1.2 a, b Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – COSY DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – COSY DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – COSY DATA HSQC suggests diastereotopic protons: 3.08/2.44 ppm 1.86/2.07 ppm Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HMBC DATA And many more… Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HMBC DATA ATOM dC (ppm) DEPT dH (ppm) COSY (HH) HMBC (CH) A 131 CH 5.5 b, c, d/d’, f b, c, d, f B 124 5.2 a, d/d’, f a, d, f C 68 4.0 a, d/d’, e a, d, e D 42 CH2 3.0 2.5 a, b, c, d, e a, b, c, e E 23 CH3 1.5 c, d/d’ c, d F 17 1.2 a, b Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HMBC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HMBC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY – HMBC DATA Organic Structure Analysis, Crews, Rodriguez and Jaspars

Combinatorial explosion STRATEGY BASED ON C-H CONNECTIVITY RETROSPECTIVE CHECKING Combinatorial explosion Pieces: Possibilities: Organic Structure Analysis, Crews, Rodriguez and Jaspars

STRATEGY BASED ON C-H CONNECTIVITY RETROSPECTIVE CHECKING And similarly for COSY data Organic Structure Analysis, Crews, Rodriguez and Jaspars

PROSPECTIVE CHECKING Pieces: Organic Structure Analysis, Crews, Rodriguez and Jaspars

2D EXERCISE 1. For a simple organic compound the mass spectrum shows a molecular ion at m/z 98. The following data has been obtained from various 1D and 2D NMR experiments. Using this information determine the structure of the molecule in question and rationalise the 2D NMR data given. Atom dC (ppm) dH (ppm) 1H - 1H COSY (3 bond only) 1H  13C Long range (2 - 3 bonds) A 218 s - A-b, A-c, A-d, A-e B 47 t 1.8 dd b-d B-c, B-d, B-e, B-f C 38 t 2.3 m c-e C-b, C-d, C-e D 32 d 1.5 m d-b, d-e, d-f D-b, D-c, D-e, D-f E 31 t 2.2 m e-c, e-d E-b, E-c, E-d, E-f F 20 q 1.1 d f-d F-b, F-d, F-e Organic Structure Analysis, Crews, Rodriguez and Jaspars

An additional peak is present in the 1H NMR at 11.6 ppm (bs). Atom 2D EXERCISE 2. For a simple organic compound the mass spectrum shows a molecular ion at m/z 114. The following data has been obtained from various 1D and 2D NMR experiments. Using this information determine the structure of the molecule in question and rationalise the 2D NMR data given. An additional peak is present in the 1H NMR at 11.6 ppm (bs). Atom dC (ppm) dH (ppm) 1H - 1H COSY (3 bond only) 1H  13C Long range (2 - 3 bonds) A 178 s - A-d, A-b B 136 d 5.7 m b-c, b-d B-d, B-c, B-e C 118 d 5.5 m c-b, c-e C-b, C-d, C-e, C-f D 38 t 3.0 d d-b D-b, D-c E 25 t 2.1 m e-c, e-f E-b, E-c, E-f F 13 q 1.0 t f-e F-c, F-e Organic Structure Analysis, Crews, Rodriguez and Jaspars