NOESY spectra pulse sequence

Figure 3.24 The pulse sequence for two-dimensional nuclear Overhauser effect spectroscopy (NOESY), The pulse sequence is divided into the preparation (P), evolution or tx (E), mixing (M), and detection or t2 (D) periods. The data are recorded in the detection period for many equally spaced values of tx and double Fourier-transformed to give the two-dimensional frequency spectrum.

The pulse sequences for HMQC-COSY and HMQC-NOESY experiments are presented in Fig. 6.10. The 3D HMQC-COSY spectrum of a N labeled tripeptide is shown in Fig. 6.11. Since the coherence transfer involved in this experiment is N([Pg.362]

Fig. 17. ID RELAY-NOESY spectra of the oligosaccharide 5 acquired using the pulse sequence of fig. 13(d) with the initial polarization transfer from overlapping anomeric protons of two terminal glucoses in 5. (a) Partial H spectrum of 7 at 600 MHz after 8 scans. The following parameters were used to acquire the two original ID RELAY-NOESY spectra (not shown) rsei = 50 ms, tq = 39 ms, A = 9.09 ms, ti = 120 ms, Tr = 64 ms, r, = 400 ms.

Fig. 6. Selected spectral regions of a NOESY spectrum of BPTI recorded with the pulse sequence of fig, 5(A), except that the first spin-lock pulse was omitted and a Bo gradient was applied during the NOESY mixing time. Protein concentration 20 mM in 90% H2O / 10% D2O, pH 6.9, 36°C. The relaxation reagent GdDTPA-BMA was added at a concentration of 750 pM to enhance the relaxation of the water protons. Spin-lock pulse 2 ms, rm(NOE) = 50 ms, r = 190 ps. Positive and negative levels were plotted without distinction. The arrow identifies the cross section containing the intermolecular water-protein cross peaks. (Reproduced by permission of the American Chemical...

In the case of isoorientin 6"-0-caffeate isolated from Gentiana arisanensis, the C NMR spectrum was assigned by H-decoupled spectra, DEPT pulse sequence, H- C COSY spectrum, long-range C- H COSY, and NOESY experiments the H NMR spectrum was analyzed with the aid of H- H COSY and H- C COSY. 

Results similar to those shown in the slice of Fig. 8.22 can be obtained with the so-called NOE-NOESY sequence [36]. Here a hyperfine shifted signal, e.g. I2-CH3 of the above compound, is selectively saturated, and then the NOESY pulse sequence is applied. The NOESY difference spectrum obtained by subtracting a NOESY spectrum without presaturation of the I2-CH3 signal is shown in Fig. 8.23. Here, some more cross peaks are evident with respect to the 3D NOESY-NOESY experiment because secondary NOEs develop much more when the primary NOEs from the I2-CH3 signal evolve in a steady state experiment like the NOE-NOESY rather than in a transient-type experiment like the NOESY-NOESY. In Fig. 8.23, dipolar connectivity patterns are apparent among protons... 

In principle, all the combinations of homonuclear 2D spectroscopies can be performed to originate a 3D spectrum (COSY-COSY, NOESY-COSY, NOESY-TOCSY, etc.). The considerations made in this chapter for the most basic experiments can be easily extended to their combinations. The general guideline should always be that the more complex the pulse sequence is, the more the experimental sensitivity will suffer from fast nuclear relaxation. 

Figure 12.12a gives a good illustration of the need for going to a third dimension to facilitate the interpretation of a crowded 2D spectrum. The NOESY spectrum of a uniformly 15N-enriched protein, staphylococcal nuclease, has so many cross peaks that interpretation is virtually impossible. However, it is possible to use, 5N chemical shifts to edit this spectrum, as indicated in Fig. 12.121) and c in a three-dimensional experiment. With the 15N enrichment, NOESY can be combined with a heteronuclear correlation experiment, in this case HMQC, but HSQC could also be used. A 3D pulse sequence can be obtained from two separate 2D experiments by deleting the detection period of one experiment and the preparation period of the other to obtain two evolution periods (q and t2) and one detection period (f3). In principle, the two 2D components can be placed in either order. For the NOESY-HMQC experiment, either order works well, but in some instances coherence transfer proceeds more efficiendy with a particular arrangement of the component experiments. We look first at the NOESY-HMQC sequence, for which a pulse sequence is given in Fig. 12.13. The three types of spins are designated I and S (as usual), both of which are H in the current example, and T, which is 15N in this case.

NMR samples contained 0.6 ml receptor (0.5-2.0 mM) dissolved in refolding buffer (vide supra) with 10% DjO. One-dimensional F NMR spectra were obtained at 470 mHz on a General Electric GN 500 spectrometer fitted with a 5 mm F probe. Parameters included 16K data points, 3.0 second relaxation delay and 25 Hz linebroadening for processing spectra. T, relaxation times were measured by the inversion recovery method. The two-dimensional F NOESY NMR spectrum was obtained on a Varian Unity Plus 500 using the standard Varian pulse sequence. A total of 128 experiments with a mixing time of 0.3 seconds were performed with collection of 1024 data points. Quadrature detection in the second dimension was obtained through the method of States and Haberkom. C ( H NMR spectra were obtained on a Varian 500 Unity Plus fitted with a 10 mm broadband probe. 

Solvent suppression is a particular problem in LC/NMR and has been a theme throughout its development. Early methods for suppression of the pro-tonated solvent signals which otherwise dominate the NMR spectrum made use of binomial pulse sequences [124-126]. Methods in use today either use fully deuterated solvents, or make use of solvent suppression schemes such as the NOESY presaturation technique [127], WATERGATE [28,128], WET [29,129], or excitation sculpting [30,130,131]. These methods have for some time made it possible to study relatively low-level (several %) impurities [132,133]. The need... 

Fig. 8.33 Pulse sequence schematic for the gradient ID NOESY experiment. The double pulsed field gradient spin echo (DPFGSE) refocuses only for that resonance subject to the selective pulse. All other magnetization is left defocused in the xy-plane. Ultimately, a NOESY spectrum is recorded only for the selected re-...

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