Heteronuclear experiments

HETCOR (Heteronuclear Correlation) refers to a 2D NMR experiment that correlates couplings between different nuclei (usually 1H and a heteroatom, such as 13C or 15N). Heteronuclear experiments can easily be extended into three or more dimensions, which can be thought of as experiments that correlate couplings between three or more different nuclei. Because there are two different frequency domains, there are no diagonal peaks like there are in COSY or TOCSY. Recently, inverse-detected HETCOR experiments have become extremely useful and commonplace, and it will be those experiments that will be covered here. Inverse-detection refers to detecting the nucleus with the higher gyromagnetic ratio, which offers higher sensitivity. It is ideal to determine which nucleus has the highest gyromagnetic ratio for detection and set the probe to be the most sensitive to this nucleus. In HETCOR, the nucleus that was detected first in a 1H -13C experiment was 13C, whereas now 1H is detected first in inverse-detection experiments, since protons are inherently more sensitive. Today, regular HETCOR experiments are not usually in common laboratory practice.

The HMQC (Heteronuclear Multiple-Quantum Coherence) experiment acquires a spectrum (see Figurea) by transferring the proton magnetization by way of 1JCH to a heteronucleus, for example, 13C. The 13C atom then experiences its chemical shift in the t1 time period of the pulse sequence. The magnetization then returns to the 1H for detection. HMQC detects 1JCH coupling and can also be used to differentiate between geminal and vicinal proton couplings just as in COSY-45. HMQC is very widely used and offers very good sensitivity at much shorter acquisition times than HETCOR (about 30 min as opposed to several hours with HETCOR).

However, because it shows the 1H -1H couplings in addition to 1H -13C couplings and because the cross peaks appear as multiplets, HMQC suffers when it comes to resolution in the 13C peaks. The HSQC (Heteronuclear Single-Quantum Coherence) experiment can assist, as it can suppress the 1H -1H couplings and collapse the multiplets seen in the cross peaks into singlets, which greatly enhances resolution (an example of an HSQC is shown in Figureb). Figure shows a side-by-side comparison of spectra from HMQC and HSQC experiments, in which some of the peaks in the HMQC spectrum are more resolved in the HSQC spectrum. However, HSQC administers more pulses than HMQC, which causes miss-settings and inhomogeneity between the RF pulses, which in turn leads to loss of sensitivity. In HMBC (Heteronuclear Multiple Bond Coherence) experiments, two and three bond couplings can be detected. This technique is particularly useful for putting smaller proposed fragments of a molecule together to elucidate the larger overall structure. HMBC, on the other hand, cannot distinguish between 2JCH and 3JCH coupling constants. An example spectrum is shown in Figured.

Spectra of heteronuclear experiments: 

(a) A spectrum of xylobiose (structure shown in Figureb) taken from a 1H-13C HMQC 2D NMR experiment. 

(b) A spectrum of codeine taken from an HSQC 1H-13C 2D NMR experiment. 

Spectrum from Acorn NMR, Inc. c) The chemical structure of codeine. 

d) Another spectrum of xylobiose taken from a 1H-13C HMBC 2D NMR experiment. 

Panels (a) and (d) from F. Sauriol, NMR Webcourse, Department of Chemistry, Queen’s University, Ontario, www.chem.queensu.ca/facilities/nmr/nmr/webcourse/.

Side-by-side comparison of an HMQC spectrum (a) and an HSQC spectrum (b). 

The HSQC experiment offers better resolution than the HMQC as well as sharper peaks. 

HSQC helps solve the problem of overlapping peaks, which is often seen in NMR experiments. 

The sample in both spectra is codeine. Spectra from Acorn NMR, Inc.