Correlation spectroscopy combination experiments

When combining isotope filtering/editing with coherence transfer steps to multidimensional experiments, then further size restrictions apply. For example, isotope edited / filtered H TOCSY or COSY experiments are generally limited to systems of <10 kDa, because of their sensitivity to T2 relaxation. In larger systems, heteronuclear correlation spectroscopy can be used for the correspondingly labeled component, while structural information about both the labeled and unlabeled moiety can be extracted from isotope edi-ted/filtered NOESY spectra, respectively. 

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. 

All of the protons in each of 12 thermo- and photochromic BIPS were assigned through a combination of homonuclear decoupling experiments and correlation spectroscopy. The relative stereochemistry of the gem-dimethyl groups could be assigned on the basis of Nuclear Overhauser Effect (NOE) experiments.131... 

Glass et al., were the first to mention the enormous sensitivity gain achievable by H-detected Se correlation spectroscopy " They showed for a number of representative organoselenium compound that the theoretical SIN ratio enhancement factor of 72, as compared to direct Se detection, can nearly be achieved so that this technique should be applied for recording biomolecules in low concentration, e.g., in selenoproteins. If inverse detection is combined with isotopic enrichment of Se, the factor may be increased up to 800. Selenomethionine, selenocysteine, and related compounds have been recorded using Se, H HETCOR experiments. ... 

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. 

Both nitrogen atoms of quinine are evident and, without hesitation, we can make assignments. If, instead, we had isolated quinine as a new unknown, natural compound we could envision a procedure in which we extend the concept of identifying compounds from a combination of spectra to include heteronuclear NMR 15N NMR would assume a natural place alongside mass, infrared, and other NMR spectrometries. Furthermore, as we made the transition from simple H and 13C spectra to correlation spectroscopy in Chapter 6, we asked Is there more to 15N NMR than proton-decoupled (and proton-coupled) spectra The question is rhetorical it is obvious that correlation experiments are possible and... 

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 a dual-color cross-correlation fluorescence spectroscopy (DCCFS) experiment [46], a sample containing two fluorophores with different emissions in each molecule was irradiated with two lasers (or with one laser) to perform simultaneous excitation of the fluorophores. The DCCFS in combination with the confocal laser microscopy allows the separation of microscopic volume with two different fluorophores from volume with only one of them and, therefore, the monitoring of dissociation of the dual-labeled molecules or association of two single-labeled molecules. Optical setup as realized in an inverted microscope to perform simultaneous excitation of the fluorophores (Figure 11.14). 

MQ NMR spectroscopy combined with heteronuclear NMR correlation experiments further increase spectral resolution and simplify the assignment of the detected NMR resonances. Moreover, information on interactions with neighboring atoms can be obtained along with the measurements of the interatomic distances. One of the first combined MQ experiments was conducted with the cross-polarization (CP) technique, which is normally used 1... 

ENDOR techniques work rather poorly if the hyperfine interaction and the nuclear Zeeman interaction are of the same order of magnitude. In this situation, electron and nuclear spin states are mixed and formally forbidden transitions, in which both the electron and nuclear spin flip, become partially allowed. Oscillations with the frequency of nuclear transitions then show up in simple electron spin echo experiments. Although such electron spin echo envelope modulation (ESEEM) experiments are not strictly double-resonance techniques, they are treated in this chapter (Section 5) because of their close relation and complementarity to ENDOR. The ESEEM experiments allow for extensive manipulations of the nuclear spins and thus for a more detailed separation of interactions. From the multitude of such experiments, we select here combination-peak ESEEM and hyperfine sublevel correlation spectroscopy (HYSCORE), which can separate the anisotropic dipole-dipole part of the hyperfine coupling from the isotropic Fermi contact interaction. 

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