A. S. Edison University of Georgia 2016 Fast Course in NMR Lecture 9 Jan/Feb, 2016 Big picture 1D Review of how we do it Why 2D? What information does 2D give?

A. S. Edison University of Georgia 2016 Human Urine Bouatra et al: Estimate at least 3079 compounds

A. S. Edison University of Georgia 2016 NMR Implementation Very Strong Magnet Probe ~400,000x Earth’s magnetic field Up to 14x greater than average MRI magnet The NMR console is a radio transmitter and receiver

A. S. Edison University of Georgia 2016 Common Coils for NMR Probes HTS coils Side view of single pair Top view of HTS probe layout for 4 nuclei Jason Kitchen (NHMFL) kindly provided the wire drawings Solenoid Flow probe: Protasis Saddle Coil Standard coil for 1 H Alderman-Grant coil Standard coil for X nuclei The basic requirements for a probe are that it has an electrical conductor oriented to deliver a magnetic field B 1 that is perpendicular to the static field B 0.

A. S. Edison University of Georgia 2016 Fourier transform The nuclear spins induce an oscillating voltage in the coil

A. S. Edison University of Georgia 2016 The anatomy of 1D 1 H NMR This urine spectrum arises from 10s to several 100s of simpler spectra

A. S. Edison University of Georgia 2016

A. S. Edison University of Georgia 2016 J-coupling For spin ½, splitting at one nucleus is i+1 where i is the number of 1 Hs at coupled nucleus quartet doublet Coupling occurs up to about 3 chemical bonds

A. S. Edison University of Georgia 2016

A. S. Edison University of Georgia 2016

A. S. Edison University of Georgia 2016 NMR is quantitative (when done with full relaxation) Urine is much more complicated!

A. S. Edison University of Georgia D NMR Reduces overlap in complicated spectra Provides a lot of additional information through correlations Can ID molecules de novo using 2D NMR Can be used as a basis for database matching: COLMAR

A. S. Edison University of Georgia D TOCSY spectrum of human urine

A. S. Edison University of Georgia 2016 TOCSY and 1D

A. S. Edison University of Georgia 2016 Information Content of Common 2D NMR Experiments COSY (Correlation Spectroscopy): J coupling (generally up to 3 covalent bonds) TOCSY (Total Correlation Spectroscopy): J coupling along coupled networks Blue is one TOCSY network, green is another HSQC (Heteronuclear Single Quantum Correlation): Directly bonded 13 C- 1 H or 15 N- 1 H HMBC (Heteronuclear Multiple Bond Correlation): 2 or 3 bond 13 C--- 1 H or 15 N H NOESY (Nuclear Overhauser Effect Spectroscopy) Or ROESY (Rotating Frame Overhauser Effect Spectroscopy): 1 H to 1 H distances up to 5-6 Å

A. S. Edison University of Georgia 2016 Using 2D NMR to ID Metabolites COSY spectrum

A. S. Edison University of Georgia 2016 This chain is almost symmetric. How do we know which end is which?

A. S. Edison University of Georgia 2016 HSQCHMBC COSY Working with multiple 2D datasets.