ROESY mixing

If we used a continuous-wave spin lock (ROESY mixing scheme) for TOCSY mixing, the only way efficient transfer could occur via 7 couplings within a spin system would be if the two protons have opposite resonance offsets (v0 - vr), where vr is the carrier or reference frequency. This is a special case of the Hartmann-Hahn match ... 

In both ROESY mixing and TOCSY mixing, the transfer is from in-phase net magnetization on the spin-lock axis to in-phase net magnetization on the spin-lock axis for example, ly ->1 (TOCSY) or ly - - Iy (ROESY). This is analogous to the NOESY transfer, It -> — for small molecules and in contrast to the INEPT transfer 21 -> 21 1 , which is antiphase to antiphase. 

Exercise Estimate the Varian power level settings for TOCSY mixing (8333 Hz), for ROESY mixing (3333 Hz), and for H decoupling (2778 Hz) if the 90° pulse is 21.3 jjis at a power setting of 59 dB. 

The ROESY used to be a bit difficult to set up because the low power spin-lock RF had to come from a different source than the hard pulse RF. Now rapid solid-state power switching is so routine that all H RF comes from the same source, with no variation of phase or frequency. ROESY tends to replace the NOESY experiment for NOE measurements, especially for small molecules where T2 is relatively long. Because the NOE builds up about twice as fast in the x -y plane as it does on the z axis, ROESY mixing times are set to about half of what would be the NOESY mixing time. There is one additional parameter to set up the power level ( Z i field strength ) of the spin lock pulse. This is typically... 

Figure 8.49. The tic-toc motion of spin-locked vectors during the Tr-ROESY mixing sequence.

The pioneering work in this field, a two-dimensional relayed-NOE experiment proposed by Wagner [7], was quickly followed by the appearance of several related NMR techniques [8-17]. Application of isotropic mixing during the J-transfer period yielded the 2D TOCSY-NOESY [11, 15] and NOESY-TOCSY [12, 14] experiments. When spin-lock sequences were applied to both J and NOE-transfers, the 2D TOCSY-ROESY and ROESY-TOCSY experiments [10, 16, 17] emerged. 

The ID TOCSY module has been used in many pseudo-3D experiments (or alternatively referred to as ID analogues of 3D experiments in the literature) such as ID TOCSY-NOESY or ID TOCSY-ROESY experiments. The TOCSY part of these experiments are similar to that of a regular ID TOCSY where a selective excitation of a desired signal is followed by a MLEV17-type isotropic mixing. The second polarization transfer (NOESY or ROESY) step can either be non-selective [29, 59-61] or selective [62-65]. 

The easiest way to reduce the amplitude of TOCSY cross peaks in the ROESY spectra is to record a spectrum with minimal spin-lock power [23]. The other possibility is to modulate the frequency of the spin-lock field [25]. However, the most convenient way is to apply a series of 180° pulses instead of a single continuous-wave pulse during the mixing time, as is done in the T-ROESY experiment. Figure 4(D) shows the T-ROESY spectrum of cyclo(Pro-Gly) recorded with = 300 ms. Although the... 

In contrast to the 1D experiment, where steady-state NOEs may be obtained, only the less intense transient NOE.s are measured in the NOESY experiment. ROEs can only be obtained as transient effects in both the ID and the 2D experiment. Furthermore the intensities of the NOESY and ROESY cross peaks depend upon the molecular size as well as the length of the mixing period. In the case of large molecules, e.g. polypeptides, rather short mixing times are usually chosen to avoid spin diffusion. 

The 2D NOESY and the 2D ROESY experiments may also be used to measure NOE-or ROE- build-up rates. This is accomplished using a series of experiments, where the mixing period D9 or the length of the spin-lock period respectively is incremented from experiment to experiment. From build-up rates relative internuclear distances may be estimated and calculated. 

MLEV17 sequence being one of the most used sequences [23]) in such a way as to continuously refocus the chemical shift evolution of the various signals in the xy plane. Analogously to ROESY experiments, the magnetization during the spin-lock (mixing) time disappears with T p (i.e. essentially Tj, see Section 3.4). It follows that coherence transfer in the xy plane, which is built up with a sin(7T J/jt) function, also decreases with time constant p p — p[p + p p)/2 ... 

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