Pulse sequence ROESY

Figure 5.45 Pulse sequence used in the ROESY experiment. The data obtained from odd and even scans are stored separately.

The pulse sequence for the ID ROESY experiment using purged half-Gaussian pulses is shown in Fig. 7.7. The purging is required to remove the dispersive components, since these are not completely eliminated by the weak spin-lock field employed in the ID ROESY experiment. 

Figure 7.7 A ID ROESY pulse sequence with purged half-Gaussian excitation. (Reprinted from Mag. Reson. Chem. 29, H. Kessler et al., 527, copyright (1991), with permission from John Wiley and Sons Limited, Baffins Lane, Chichester, Sussex P019 lUD, England.)...

In the subsequent ID ROESY-TOCSY experiment (pulse sequence of fig. 7(c)), a selective TOCSY transfer was applied from H-4c. During the... 

Fig. 8. ID ROESY-TOCSY. (a) H spectrum of the oligosaccharide 3 (5 mg/0.5 ml D2O). (b) ID ROESY spectrum of 3 acquired using the pulse sequence of fig. 7(a) with selective excitation of the H-lb proton. Duration of the 270° Gaussian pulse and the spin-lock pulse ( yBi/ K = 2.8 kHz) was 49.2 ms and 0.5 s, respectively. The spin-lock pulse was applied 333.3 Hz downfield from the H-lb resonance. The time used for the frequency change was 3 ms. (c) ID ROESY-TOCSY spectrum acquired using the pulse sequence of fig. 7(c) and the selective ROESY transfer from H-lb followed by a selective TOCSY transfer from H-4c. Parameters for the ROESY part were the same as in (b). A 49.2 ms Gaussian pulse was used at the beginning of the 29.07 ms TOCSY spin lock. 256 scans were accumulated. A partial structure of 3 is given in the inset. Solid and dotted lines represent TOCSY and ROESY...

The ID TOCSY-ROESY experiment is illustrated on the same molecule using the pulse sequence of fig. 7(d). This time the magnetization of H-4c was generated during the initial selective TOCSY transfer from H-lc (fig. 9(b), pulse sequence of fig. 7(b)). In the subsequent ID TOCSY-ROESY experiment, the ROE transfer from H-4c confirmed the expected... 

Fig. 1. Basic pulse sequence and CP diagram for gradient-based spin-locked ID exf>eriments. A 1 (— 1) 2 gradient ratio selects N-type data (solid lines) while 1 (— 1) (—2) selects P-type data (dashed lines). When SL stands for a -filtered DIPSI-2 pulse train, a ge-lD TOeSY is performed. On the other hand, when SL stands for a T-ROESY pulse train, a GROESY experiment is performed. S stands for the gradient length.

The use of spin-lock pulses for water suppression is illustrated with the NOESY and ROESY pulse sequences (fig. 5). Using the Cartesian product operator description [9], the effect of the NOESY pulse sequence of fig. 5(A) is readily illustrated ... 

Fig. 5. Pulse sequences of NOESY and ROESY with spin-lock purge pulses for water suppression. (A) NOESY pulse sequence. The spin-lock pulses are typically of length 0.5 ms and 2 ms, and r = 1/SW, where SW is the spectral width in the acquisition dimension. Phase cycle (pi = x,—x) 4>2 = 4 x,x,—x,—x) ...

Compared to other multidimensional experiments the exchange experiments are fairly simple and, thus, easy to optimize. Experiments are robust with regard to the pulse imperfections and miscalibration. All artifacts except coherence transfer can be removed with standard phase cycling of RF pulses and receiver. The coherence transfer can be removed by appropriate pulse sequences, preferably with T-ROESY. 

Fig. 8.2. Some of the most common 2D pulse sequences that can be employed using a proper choice of parameters to record 2D spectra of paramagnetic molecules (A) NOESY, (B) ROESY, (C) COSY, (D) ISECR COSY, (E) zero-quantum (double quantum) COSY, (F) TOCSY, (G) HMQC, (H) HSQC. Sequences (A), (B) and (F) are also used to obtain EXSY spectra. SL indicates a soft spin-lock sequence, while MLEV17 indicates a train of spin-locking hard pulses that optimizes the development of J/j coupling. In the reverse heteronuclear experiment (G) the upper and lower levels refer to H and heteronucleus, respectively. The phase cycles are not indicated. For clarity of discussion, all initial pulses can be thought to be applied along the y axis, in such a way that the coherence after the first 90° pulse is always along x. ...

It should be noted that NOESY and ROESY pulse sequences also provide EXSY spectra, and therefore EXSY cross peaks may appear simultaneously in the 2D NOESY and ROESY spectra. EXSY cross peaks are always positive in both types of experiment, whereas dipolar cross peaks are negative in EXSY spectra independently of molecular weight and in NOESY spectra of small molecules. Therefore, in macromolecules the sign for NOESY and EXSY cross peaks is the same, and the two phenomena cannot be distinguished in NOESY experiments. In contrast, ROESY cross peaks have different sign from EXSY cross peaks and can be distinguished and even plotted selectively in ROESY experiments. These considerations are summarized in Table 8.3 for the reader s convenience. 

Chapter 5 still covers 2-D correlation but has been reorganized, expanded, and updated, which reflects the ever increasing importance of 2-D NMR. The reorganization places all of the spectra together for a given compound and treats each example separately ipsenol, caryophyllene oxide, lactose, and a tetrapeptide. Pulse sequences for most of the experiments are given. The expanded treatment also includes many new 2-D experiments such as ROESY and hybrid experiments such as HMQC-TOCSY. There are many new Student Exercises. 

The pulse sequence for ICP experiments appears simple a 90° proton pulse is followed immediately by a spin lock radio-frequency (rf) field of strength B that is phase shifted by 90° relative to the first pulse. By a spin-lock field is meant a strong rf field B that is on resonance with the given nucleus it keeps magnetization in a spin-locked orientation parallel to the B direction where the decay of magnetization is governed by T p. At present the strong continuous B field is replaced by multipulse sequences that are well known from other spin-lock experiments such as TOCSY, ROESY etc. Simultaneously,... 

A 2002 review by Reynolds and Enriquez describes the most effective pulse sequences for natural product structure elucidation.86 For natural product chemists, the review recommends HSQC over HMQC, T-ROESY (transverse rotating-frame Overhauser enhancement) in place of NOESY (nuclear Over-hauser enhancement spectroscopy) and CIGAR (constant time inverse-detected gradient accordion rescaled) or constant time HMBC over HMBC. HSQC spectra provide better line shapes than HMQC spectra, but are more demanding on spectrometer hardware. The T-ROESY or transverse ROESY provides better signal to noise for most small molecules compared with a NOESY and limits scalar coupling artefacts. In small-molecule NMR at natural abundance, the 2D HMBC or variants experiment stands out as one of the key NMR experiments for structure elucidation. HMBC spectra provide correlations over multiple bonds and, while this is desirable, it poses the problem of distinguishing between two- and three-bond correlations. 

The experimental method for obtaining ROESY is essentially the same as that for HOHAHA, application of a spin-lock pulse sequence for mixing at the end of the evolution period. HOHAHA effects can interfere with ROESY measurements but are minimized by using lower rf power and offsetting the pulse frequency to interfere with the Hartmann-Hahn condition. 

Many other pulse-sequence techniques besides COSY can be used to produce multidimensional NMR spectra. It will suffice here to simply list the acronyms of some of the better known methods EXSY (exchange spectroscopy), NOESY (nuclear Overhauser effect spectroscopy), TOCS Y (total correlation spectroscopy), ROESY (rotational nuclear Overhauser effect spectroscopy). The nuclear Overhauser effect (NOE) refers to a change in intensity of one NMR peak when another peak is irradiated. 

The rotating-frame NOESY experiment (ROESY) provides some advantages for small and medium-sized, as well as large, molecules. The pulse sequence for ROESY (previously... 

Since the pulse sequence is the same for EXSY and NOESY, NOESY (or ROESY) cross peaks might be mistaken for EXSY cross peaks. They can be distinguished in the phase-sensitive experiment, since EXSY and ROESY peaks have opposite phases, as do EXSY and NOESY peaks in the fast motion regime. For example, two resolved OH or NH resonances may exhibit EXSY cross peaks from slow proton exchange. These peaks could be mistakenly taken to be NOESY peaks and interpreted incorrectly in terms of stereochemistry. 

The first of these arises when the long spin-lock pulse acts in an analogous fashion to the last 90" pulse of the COSY experiment so causing coherence transfer between J-coupled spins. The resulting peaks display the usual antiphase COSY peak stmcture and tend to be weak so are of least concern. A far greater problem arises from TOCSY transfers which arise because the spin-lock period in ROESY is similar to that used in the TOCSY experiment (Section 5.7). This may, therefore, also induce coherent transfers between J-coupled spins when these experience similar rf fields, that is, when the Hartmann-Hahn matching condition is satisfied. Since the ROESY spin-lock is not modulated (i.e. not a composite pulse sequence), this match is restricted to mutually coupled spins with similar chemical shift offsets or to those with equal but opposite... 

In Figures 7 and 8 the 2D proton exchange spectrum of the hydrothermally dealuminated zeolite H-Y loaded with ca. 40 H2O per unit cell (identical to the sample in Figure 6C) is shown as contour and stacked plot, respectively. A homonuclear 2D ROESY pulse sequence [5] and a spin lock pulse of 2 ms were used. The absence of cross peaks between the signals at 1.8+0.2 ppm and... 

FIGURE 19-34 The 500-MHz one-dimenstonal H NMR spectrum (a) and a portion of the two-dimensional ROESY spectrum (b) of a nineteen-amino acid protein. The pulse sequence used collapses multiplels due to H- H spin-spin coupling inlo singlets. The cross peaks of the dipolar interactions make it possible lo completely assign the proton NMR spectrum. (Adapted from A, Kaerner and D. L Rabensiein, Magn. Reson. Chem., 1998.36.601. Copyright t998 Interscience/Wiley.)... 

The information regarding the structure of the (R,S)-CL complexes with HDA-)9-CyD was obtained using ROESY experiments, which were performed with three different pulse sequence for the complex between CyDs and a 3 1 (w/w) mixture of (S)- and (R)-CL. 

Fig. 8.16 ROESY pulse sequence [35, 36, 52, 53]. Protons are labeled with their respective chemical shifts during the evolution time, ti, as with the COSY and NOESY experiments.

Fig. 17A,B. Pulse sequences for HMQC-NOESY (A) (Sohn and Opella 1989) and HMQC-ROESY (B) (Kawabata et al. 1992a)...

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