Molecular Biogeochemistry - Tutorial

NMR mini course Outline • Basics of NMR • 1H NMR • 13C NMR • COESY & NOESY Basics • Nuclei are immersed in a static magnetic field and exposed to a second oscillating magnetic field • Some nuclei experience this phenomenon, and others do not, dependent on whether or not they have spin • Superconducting magnet produces the Bo field necessary • Immediately within the bore of the magnet are the shim coils for homogenizing the Bo field • Within the shim coils is the probe • Probe contains the RF coils for producing the B1 magnetic field necessary to rotate the spins by 90o or 180o • RF coil also detects the signal from the spins within the sample • Sample is positioned within the RF coil of the probe • Nuclear magnetic resonance (NMR) spectroscopy involves using energy in the form of electromagnetic radiation (radio waves) to orient nuclei into the less energetically favorable beta state • When the energy is removed, the energized nuclei relax back to the alpha state • The fluctuation of the magnetic field associated with this relaxation process is called resonance and this resonance can be detected and converted into peaks • Nuclei that are not in identical structural situations do not experience the external magnetic field to the same extent 1H • • • • • Nuclei are shielded or deshielded due to small local fields generated by circulating sigma, pi and lone pair electrons Shielded = lower frequency Hydrogens near an electronegative atom require a higher frequency to flip, thus they appear at a higher ppm in the spectrum Exact frequency depends on solvent If paramagnetic species are included, range become much wider Chemical Shifts Multiplicity N+1 rule -OH usually exchanges rapidly and is a broad singlet 13C NMR Chemical Shift (δ) Type of Carbon 13C has only about 1.1% natural abundance (of carbon atoms) C is about 400 times less sensitive than H nucleus to the NMR phenomena Chemical shifts measured with respect to tetramethylsilane, (CH3)4Si (i.e. TMS) Chemical shift range is normally 0 to 220 ppm Similar factors affect the chemical shifts in 13C as seen for H NMR Long relaxation times (excited state to ground state) mean no integrations (carbon signals don’t relax equally) Due to low abundance, do not usually see 13C13C coupling Carbons couple with the hydrogen atoms that are directly attached to them "Normal" 13C spectra are "broadband, proton decoupled" so the peaks show as single lines Number of peaks indicates the number of types of C From NOE enhancements we can usually say the peak intensity range CH3>CH2>CH>C 10-40 ppm Alkane C's 115-150 ppm Aromatic 40-70 ppm Adjacent to an electronegative atom 160-185 Carbonyl carbon of acid derivatives 65-90 ppm sp carbon of an alkyne 190-220 ppm carbonyl carbon of ketones and aldehydes 110-140 ppm sp2 carbon of an alkene or sp of a nitrile Multiple functional groups (Chemical shift effects are approximately additive ) 2D Techniques: COSY • COrrelation SpectroscopY • Determine the connectivity of a molecule by determining which protons are spin-spin coupled • The proton spectrum is plotted on each of the two axes • The diagonal within the box is the spectrum as seen from "above" • Off-diagonal peaks denote splitting between protons on adjacent carbons 2D Techniques: NOESY • Nuclear Overhauser Effect SpectroscopY is useful for determining which signals arise from protons that are close to each other in space even if they are not bonded