HSQC

The applicability of the HSQC experiment in natural product structure elucidation studies remains to be evaluated. To the best of our knowledge, no natural product structure elucidation studies have been reported using HSQC rather than HMQC to establish direct proton-carbon chemical shift correlations. This will undoubtedly change in time. A modification of the HSQC experiment, inverted direct response HSQC-TOCSY, has, however, been reported using strychnine as a model compound (Domke 1991). In the case of congested proton spectra, there may be advantages inherent to the use of HSQC instead of HMQC. 

IZSX---- IZSX cos fis(j + IzSy sin fisq - IxSxcosClst1 + IXSZ sin (lsq 

at time 9 we have a double quantum coherence (IXSX), which will not generate any signal, and antiphase / magnetization (IXSZ) multiplied by the sine 

FIGURE 12.8 Pulse sequence for the heteronuclear single quantum coherence experiment. See text for discussion of the state of the spin system at the times indicated. 

If broadband decoupling is applied to the 5 spins (e.g., 13C), a proton spectrum decoupled from 13C is obtained in dimension o 2. We have ignored the H—H couplings, which also evolve during the experiment, as they are usually a small 

FIGURE 12.9 Example of heteronuclear single quantum coherence (HSQC) applied to allylbutyl ether (300 MHz).The correlations of H and 1 C chemical shifts are clearly shown. Note the similarity to Fig. 10.10, which displays a HETCOR spectrum. For a sample of this sort, where signal/noise ratio is no problem, there is little to choose between the two techniques, but HSQC is inherently much more sensitive. 

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HC HMQC (heteronuclear multiple quantum coherence) and HC HSQC (heteronuclear single quantum coherence) are the acronyms of the pulse sequences used for inverse carbon-proton shift correlations. These sensitive inverse experiments detect one-bond carbon-proton connectivities within some minutes instead of some hours as required for CH COSY as demonstrated by an HC HSQC experiment with a-pinene in Fig. 2.15. 

Figure 2.15. HC HSQC experiment (contour plot) of a-pinene [ CDCI3, 5 % v/v, 25 °C, 125 MHz for C, 500 MHz for h, 4 scans, 256 experiments]. This experiment gives the same information as Fig. 2.14 within 8 minutes instead of two hours required for the CH-COSY in Fig. 2.14 due to higher sensitivity because of proton detection and stronger magnetic field. Deviations of proton shifts from those in Fig. 2.14 arise from the change of the solvent. The methylene protons collapsing in Fig. 2.14 at Sh = 2.19 (200 MHz) display in this experiment an AB system with = 2.17 and Sg = 2.21 (500 MHz)...

Two-dimensional C//correlations such as C//COSY or HC HMQC and HSQC provide the Jqh connectivities, and thereby apply only to those C atoms which are linked to H and not to non-protonated C atoms. Modifications of these techniques, also applicable to quaternary C atoms, are those which are adjusted to the smaller Jqh and Jqh couplings (2-25 Hz, Tables 2.8 and 2.9) Experiments that probe these couplings include the CH COLOC (correlation via long range couplings) with carbon-13 detection (Fig. 2.16) and HC HMBC (heteronuclear multiple bond coherence) with the much more sensitive proton detection (Fig. 2.17)... 

These two-dimensional CH shift correlations indicate CH relationships through two and more bonds (predominantly Jch and Jch connectivities) in addition to more or less suppressed Jch relationships which are in any case established from the CH COSY contour diagram. Format and analysis of the CH COLOC or HMBC plots correspond to those of a C//COSY or HSQC experiment, as is shown for a-pinene (1) in Figs. 2.14 - 2.17. 

Conditions CDCI3, 25 °C, 500 MHz (H), 125 MHz ( C). (a) //NMR spectrum and HH COSY plot of ethyl groups (b) HC HSQC plot with inserted zoomed section of ethyl groups ... 

Carbon atoms and protons are assigned by means of the proton-carbon connectivities as identified in the HC HSQC and HMBC experiment (b and c). The latter also permits the derivation of the connection of the ethyl groups to the porphyrin ring. The cross signals in the relevant part a of the HH COSY plot (a) are used to connect the methyl and methylene subunits to the ethyl groups. 

Additionally, all carbon-proton bonds can be assigned by means of the HC HSQC experiment b ... 

HSQC Heteronuclear single quantum coherence, e.g. inverse CH correlation via one-bond coupling providing the same result as HMQC but using an alternative pulse sequence... 

Proton resonances for aU residues were assigned using a combination of COSY, TOCSY, HSQC, and HMBC experiments. The large values of y(NH/H-C(/9)) and the small values of J(H-C(a)/H-C(y9) were indicative of antiperiplanar and synclinal arrangements respectively, around those bonds. In addition, medium-range NOE connectivities H-C(/ )i/NH +i, H-C(a)i/NH, + i, NHi/NH +i were consistent... 

D-TOCSY- H- N-HMQC/HSQC H spin systems, N shifts... 

D-NOESY- H- C-HMQC/HSQC Sequential assignment of the spin systems, in particular, identification of nOes between side chains... 

Haliclonacyclamine E (13) and arenosclerins A (14), B (15), and C (16) have been isolated from the marine sponge Arenosclera brasiliensis, endemic in Brazil. Crude extracts of this sponge displayed potent cytotoxic and antibiotic activities, and were subjected to fractionation by sihca-gel flash chromatography, medium pressure chromatography on a SiOH cyanopropyl-bonded column, and reversed-phase Cis column chromatography to give compounds 13-16 [18]. The structure elucidation was based on spectroscopic analysis, including HRFABMS, COSY, HSQC, HSQC-TOCSY, and HMBC NMR... 

COSY, HMQC, HMBC, and HSQC-TOCSY NMR spectra. The relative stereochemistry of each of the piperidine rings in 21 was estabhshed by analysis of NMR and NOESY spectra. However, the authors have not obtained experimental data to relate the relative stereochemistry of each piperidine moiety to the other. They suggested the relative stereochemistry indicated in 21, based on the Baldwin and Whitehead biogenetic proposal [24] (see below) and also by comparison with data for haliclonacyclamine E (5) and arenosclerin A (14). 

Nolls, P., Espinosa, J. F., Parella, T. Optimum spin-state selection for all multiplicities in the acquisition dimension of the HSQC experiment. 

Nolls, P., Parella, T. Spin-edited 2D HSQC-TOCSY experiments for the measurement of homonuclear and heteronuclear coupling constants application to carbohydrates and peptides. /. Magn. Reson. 2005, 176, 15-26. 

HSQC is based on proteins containing C-labelled methyl groups and is increasingly used to complement HSQC experiments [37]. 

The method is more sensitive due to the higher proton multiplicity and is suitable for screening proteins up to 40kDa. The key binding site residues of a protein can be identified by using a known inhibitor to identify cross-peaks in the HSQC spectrum [38]. 

The subset of peaks that can serve as markers for the binding of replacements were identified by comparing the HSQC spectra of N-AK... 

Fig. 2 Chemical shift perturbation and chemical shift mapping, (a) Portions of the [15N, 1H]-HSQC spectra of Bcf-xL recorded in absence (black) and in presence of each of the four molecules (in colors). Resonance assignments for amino acid residues that exhibit large shifts are reported, (b) Structure of Bc1-Xl in complex with the BH3 peptide from Bak (PDB code 1BXL) showing the chemical shift changes in Bcl-xL upon ligand binding (blue, large shits yellow, no shifts the Bak peptide is reported in cyan). Adapted from [48]...

Stopped-flow H C-HSQC NMR spectrum of a-tocopherol acetate. 

Fig. 2. (Left) 750-MHz 15N- H HSQC spectrum and (right) 750-MHz HNCO spectrum of apomyoglobin at pH 2.3,10 mM acetate-

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