Isotropic mixing experiments

Isotropic effective /-coupling tensors (/) with a scaling factor s 1 are characteristic for ideal homonuclear Hartmann-Hahn experiments and, in particular, for homonuclear isotropic mixing experiments (see Section X). Isotropic effective /-coupling tensors can also be created between heteronuclear spins i and m (see Section XI) however, this results in a reduced effective coupling constant with a scaling factor i 1/3 [see Eq. (115)]. 

In this section, we restrict the discussion to constants of motion during Hartmann-Hahn experiments that are induced by the symmetry of the mixing Hamiltonian and to selection rules for cross-peak signals in two-dimensional isotropic-mixing experiments. 

The conservation of coherence order has important consequences for the practical implementation of isotropic-mixing experiments (see Section XILA). Only coherences with coherence order p = — 1 during the evolu-... 

In principle, this type of band-selective Hartmann-Hahn transfer can be used to restrict coherence transfer to spins with resonances in certain frequency ranges. However, in these experiments the effective coupling constant is reduced by a factor of 1/2, which requires longer mbdng times. However, in broadband homonuclear isotropic-mixing experiments as discussed in Section IV.C.2 coupling constants are scaled by a factor... 

Fig. 10.18. IDR (Inverted Direct Response)—HSQC-TOCSY pulse sequence. The experiment first uses an HSQC sequence to label protons with the chemical shift of their directly bound carbons, followed by an isotropic mixing period that propagates magnetization to vicinal neighbor and more distant protons. The extent to which magnetization is propagated in the experiment is a function of both the size of the intervening vicinal coupling constants and the duration of the mixing period. After isotropic mixing, direct responses are inverted by the experiment and proton detection begins.

For all the reasons mentioned above, it is more desirable to obtain PASADENA spectra when applying the PHIP method, at least initially. Nevertheless, in certain cases ALTADENA spectra can be very helpful, too, especially for a quick screening, because these experiments can be performed without any specialized NMR probes. Rather, in these cases, standard NMR equipment will suffice totally. Furthermore, in other cases, the (normally unwanted) isotropic mixing can be used advantageously to obtain information about other additional aspects of the system. A typical such case results when it is desirable to transfer the original proton polar-... 

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. 

Fig. 18. The pulse sequence of a ID ge-NOESY-TOCSY experiment, tnoe is the NOE mixing time, 5 are optional delays which can be used for z-filtration [81] or for suppression of ROE effects in macromolecules (2 x (5 + Tgrad) = 0.5 x mixing time). DIPSI-2 [78] sequence was used for isotropic mixing. Phases were cycled as follows 0i = 2x, 2(—x) (j)2 = X, —x Ip = X, 2 —x), X. Rectangular PFGs, G = 6 Gauss/cm and Gi = 7 Gauss/cm, were applied along the axis for Xpad = 1 ms.

The pulse sequence for ID TOCSY is a ID modification of the original TOCSY experiment [2] introduced by Braunschweiler and Ernst. The TOCSY experiment was also referred to as HOHAHA (which stands for HOmonuclear HArtman-HAhn) by Bax and Davis [3]. The ID TOCSY experiment was proposed by Bax and co-workers [4, 5], and by Kessler et al. [6]. The essential features of the pulse sequence involve the use of selective excitation of a desired resonance, followed by a homonu-clear Hartman-Hahn (or isotropic) mixing period [2, 7]. That is, the unit -Pnonsei - in the 2D TOCSY pulse sequence is replaced by Fsei -where P stands for a pulse (or pulses), ti is the evolution period in the 2D experiment and r is a fixed delay. 

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]. 

Figure 4 Pulse sequences 2D LASSY (or CLASSY) experiments. (A) CLASSY IM pulse sequence with isotropic mixing (IM). The narrow, wider and wide rectangles represent the 45°, 90° and 180° pulses, r — 1/2/cc, A —3 fis. The following phase cycling was used — y>3 = 4x, 4y, 4( x), 4( y) ...

Many other implementations of the PEP principle are possible for TOCSY experiments. For example, rather than performing one experiment with and one experiment without an additional 180° pulse, the same result is achieved if both experiments contain a 90° pulse before and after the isotropic-mixing period. In the first experiment, both 90° pulses have opposite phases, whereas in the second experiment the phases of the two pulses are identical. Detailed phase-cycling protocols and an alternative processing scheme in which the two experimental data sets are combined in the time domain, rather than in the frequency domain, can be found in the original literature (Cavanagh and Ranee, 1990b Ranee, 1994). 

Zero-field NMR of liquid samples (Zax et al., 1984) is also an experiment, with an incremented isotropic mixing evolution period. In this case, the energy match condition is satisfied during the evolution period by physically shuttling the sample to zero field, rather than by applying rf irradiation schemes. 

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