Total correlation spectroscopy combination experiments

All the spectroscopic approaches applied for structural characterization of mixtures derive from methods originally developed for screening libraries for their biological activities. They include diffusion-ordered spectroscopy [15-18], relaxation-edited spectroscopy [19], isotope-filtered affinity NMR [20] and SAR-by-NMR [21]. These applications will be discussed in the last part of this chapter. As usually most of the components show very similar molecular weight, their spectroscopic parameters, such as relaxation rates or selfdiffusion coefficients, are not very different and application of these methodologies for chemical characterization is not straightforward. An exception is diffusion-edited spectroscopy, which can be a feasible way to analyze the structure of compounds within a mixture without the need of prior separation. This was the case for the analysis of a mixture of five esters (propyl acetate, butyl acetate, ethyl butyrate, isopropyl butyrate and butyl levulinate) [18]. By the combined use of diffusion-edited NMR and 2-D NMR methods such as Total Correlation Spectroscopy (TOCSY), it was possible to elucidate the structure of the components of this mixture. This strategy was called diffusion encoded spectroscopy DECODES. Another example of combination between diffusion-edited spectroscopy and traditional 2-D NMR experiment is the DOSY-NOESY experiment [22]. The use of these experiments have proven to be useful in the identification of compounds from small split and mix synthetic pools. 

This structure was clearly supported by the data obtained by irradiation of the protons of HDA-j8-CyD and observing the response for the protons of CL [60]. Thus, upon irradiation of the H-3 protons of HDA-j8-CyD an intermolecular NOE was observed only for the protons of the fert-butyl moiety of CL. In combination with the NOE response observed for the aromatic protons of CL upon irradiation of the acetyl group of HDA-j8-CyD these data indicate that the fert-butyl moiety is included in the cavity and the phenyl moiety is located outside the cavity close to the secondary rim of HDA-j8-CyD. Thus, the ROESY experiment shows a significant difference between the structures of the CL complexes with fi-CyD and HDA-j8-CyD. ID and 2D transversal ROESY (T-ROESY) experiments confirmed that the effect observed in the ID ROESY spectra were solely of intermolecular origin and that there was no significant contribution due to intramolecular TOCSY (total correlation spectroscopy) magnetization transfer. Thus, all ROESY experiments clearly indicated that CL forms intermolecular inclusion complexes with -CyD and HDA-jS-CyD. The CL molecule is included in the cavity of both CyDs from the secondary wider rim. The most distinct difference between the two complexes is that the phenyl moiety of CL is most likely included in the cavity of j8-CyD whereas the fert-butyl moiety is included in the cavity of HDA- 8-CyD. 

In principle, 2D NMR structural determination of proteins in solution involves the linking of information derived from a combination of three complementary experiments - TOCSY (Total Correlated Spectroscopy), DQF-COSY (Double-Quantum Filtered Correlated Spectroscopy) and NOESY (Nuclear Overhauser Effect Spectroscopy). The TOCSY (58) spectra are used to identify spin systems of the amino acids in the protein. COSY (59,60) spectra yield complementary information to the TOCSY, which can be used to obtain the direct scalar connectives. The NOESY (61) spectra yield information on the through space relationship of protons in the protein. It should be noted that because of the vast abundance of hydrogen atoms in a protein molecule proton NMR is the preferred technique to determine the solution structure of a protein. 

Mixing sequences for total through-bond correlation spectroscopy in solids (TOBSY) have been developed for fast MAS experiments. Possible sequences with the desired Hamiltonian (the homonuclear isotropic J interaction) have been identified using lowest order average Hamiltonian theory combined with numerical simulations as a function of the MAS frequency. An experimental TOBSY spectrum of a uniformly C-labelled decapeptide at 20 kHz MAS has been obtained using one of the new sequences. The spectrum allows to assign the resonances to the respective spin systems. 

---