Phase Sensitive COSY

The COSY experiment can be either phase sensitive or nonphase sensitive (a.k.a. absolute-value COSY). The total integrated volume of a phase-sensitive COSY cross peak will always be zero because the cross peak will be composed of an equal distribution of positive and negative components. Phase-sensitive COSY spectra are useful for the measurement of 1-couplings between spins. 

If a cross peak arises from the coupling of two spins whose multiplicities are more than just doublets, the extraction of the couplings is slightly more involved. Only the I-coupling responsible for the generation of the cross peak will modulate the alternation in sign of the components of the cross peak. 

Whenever we attempt to measure 1-couplings accurately using a 2-D COSY data set, we must ensure that we have sufficient digital resolution (see Chapter 3) to give the desired precision, usually along the f2 axis. This matrix dimensional asymmetry may entail reprocessing the data set with array dimensions set at 16k X Ik instead of 4k X 4k. 

In some instances, we may find that examination of a cross peak close to the diagonal (5j is almost equal to 82) is difficult because of distortion of the cross peak due to intensity on the diagonal of the spectrum. The double-quantum filtered COSY (DQF-COSY) experiment fortunately provides a method for suppression of most of the signal intensity found on the diagonal. 

Any phase-sensitive COSY experiment has a minimum phase cycle (usually 4 scans per h time increment), and calibration of the 90° pulse is strongly recommended. Setting the receiver gain in any COSY experiment should be carried out when collecting the 1-D spectrum (and not when we digitize the first FID of the 2-D COSY, because the signal intensity in the COSY experiment builds up and reaches a maximum only after a number of FIDs have been digitized. 

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Figure 5.37 (a) Conventional phase-sensitive COSY spectrum of basic pancreatic trypsin inhibitor, (b) Double-quantum filtered (DQF) phase-sensitive COSY spectrum of the same trypsin inhibitor, in which singlet resonances and solvent signal are largely suppressed. Notice how clean the spectrum is, especially in the region near the diagonal line. (Reprinted from Biochem. Biophys. Res. Comm. 117, M. Ranee, et al., 479, copyright (1983) with permission from Academic Press, Inc.)... 

Phase-sensitive NOESY spectra are generally significantly superior in resolution to absolute-mode spectra. As in phase-sensitive COSY spectra. 

Another disadvantage is that for solving certain stereochemistry problems, it is necessary to be able to not only establish connectivity but to measure couplings fairly accurately so that the data can be used in conjunction with the Karplus curve. Whilst this is possible using a phase sensitive COSY (Note -... 

The mechanism of singlet oxygen detoxification involves quenching by Vitamin Be (215a). After silylating this compound, in order to make it soluble in CD2CI2 to work at low temperature, the quenching process proceeds as shown in equation 75 (Section V.C.4). The main product is endoperoxide 217, probably in equilibrium with hydroperoxide 216. The structure of 217 can be established from the H and NMR spectra, e.g. 5h = 6.06 ppm (IH, d, 7 = 4.4 Hz, HCOO), 4.19 (IH, broad, NH), 5c = 96.8 ppm (MeCOO), 86.6 ppm (HCOO), and DEPT, phase-sensitive COSY, HMQC and HMBC spectra . 

Note that with a non-DQ-filtered, phase sensitive COSY experiment the cross peaks are again purely absorptive while diagonal peaks irrespective of the phase correction will have both absorptive and dispersive character. Unlike most other 2D spectra, it is therefore best to phase correct a non-DQ-filtered phase sensitive COSY spectrum while examining the cross rather than the diagonal peaks. 

Load the raw data of the 2D phase sensitive COSY experiment of glucose D NMRDATA GLUCOSE 2D HH GHHCODF 001001. SER. Calculate the spectrum and proceed as above, but use the Syma command for symmetrization. Compare the results obtained with/without symmetrization. 

The determination of the conformation of 1,6-linkages requires that the conformation of the C5, C6 bond is established, in addition to conformation of the glycosidic bond. Besides the standard techniques for obtaining the coupling constants interpretation of the cross peak patterns in phase sensitive COSY spectra proved to be very valuable [29, 30]. 

A detailed inspection of the coupling constants 3J5 6, and 3 J5 6 of several manno-sidic oligosaccharides utilizing phase sensitive COSY spectra has been reported [29]. The population analysis based on these coupling constants shows the preference of the gg conformation over gt while tg is not found within the precision of the analysis. 

For more complex spectra, the data are usually presented as a series of contours just as hills and valleys are represented on a topological map. We see this representation of the data in the right part of Figure 5.6. Projections of the data are often included in 2-D spectra, which is equivalent to shining a light on the peak to reveal its shadow, which is obviously 1-D. Often these projections are replaced with actual 1-D spectra that have been acquired separately. So long as there are no negative peaks (e.g., phase sensitive COSY, not covered in this book), we use this method without comment. 

Several of the spectra in this workbook were obtained using techniques such as proton-detected C-H shift correlation and multiple-quantum-filtered phase-sensitive COSY which were not covered in detail in our main text because they have come into general use since it was written. This is a measure of the rate at which practical NMR is progressing but presents no problem in the interpretation of the spectra. Various field strengths and modes of presentation, ranging from continuous-wave traces to phase-sensitive two-dimensional contour plots, were used for the spectra. In part, this reflects the history of individual problems, but it is also intentional. It is important to be able to extract the essential message of a spectrum independently of the way it is presented. 

Several of the spectra in this workbook were obtained using techniques such as proton-detected C—H shift correlation and multiple-quantum-filtered phase-sensitive COSY which were not covered in detail in our main text because they have come into general use since it was written. 

Figure 6-15 Phase-sensitive COSY diagram for two spins, with the diagonal peaks in dispersion mode and the cross peaks in antiphase absorption mode. The ID spectrum is on the right. (Reproduced from F. J. M. van de Ven, Multidimensional NMR in Liquids, VCH, New York, 1995, p. 171.)...

Double Quantum Filtered COSY The double quantum filtered COSY experiment (DQF-COSY, Section 6-1 and Figure 6-16) is similar to COSY, with three 90° pulses in the sequence 90°-/i-90°-T-90°-f2 (acquire). The DQF-COSY experiment is performed in the phase-sensitive mode, but, unlike the situation in the phase-sensitive COSY experiment, in the DQF-COSY both diagonal and cross peaks can be phased as absorptive signals. This difference not... 

Figure 5.19. The phase-sensitive COSY for a coupled two-spin AX system. Diagonal peaks have broad, in-phase douh e-dispersion lineshapes (D) whereas crosspeaks have narrow, antiphase double-absorption lineshapes (A), as further illustrated in the row extracted from the spectrum.

For the phase-sensitive COSY experiment both p = 1 pathways must be preserved during ti. Recall that for a pure-phase 2D spectrum the signal detected in ta must be amplitude-modulated as a function of tt. Such a signal can only be obtained if both of the counter-rotating coherences are retained since the retention of only one of these unavoidably leads to a phase-modulated... 

Figure 5 Jl. The coherence level diagrams and coherence transfer pathways for (a) P-type, (b) N-type and (c) phase-sensitive COSY.

Phase-sensitive COSY-90 High-resolution display due to absorptive lineshapes. Crosspeak fine structure apparent J measurement possible. Diagonal peaks have dispersive lineshapes which may interfere with neighbouring crosspeaks. Requires high digital resolution to reveal multiplet structures. 

Previous sections have already made the case for acquiring COSY data such that it may be presented in the phase-sensitive mode. The pure-absorption lineshapes associated with this provide the highest possible resolution and allow one to extract information from the fine-structure within crosspeak multiplets. However, it was also pointed out that the basic COSY-90 sequence suffers from one serious drawback in that diagonal peaks possess dispersion-mode lineshapes when crosspeaks are phased into pure absorption-mode. The broad tails associated with these can mask crosspeaks that fall close to the diagonal, so there is potential for useful information to be lost. The presence of dispersive contributions to the diagonal may be (largely) overcome by the use of the double-quantum filtered variant of COSY [37], and for this reason DQF-COSY is the experiment of choice for recording phase-sensitive COSY data. 

Figure 5>t2. The double-quantum filtered COSY spectrum (right) provides greater clarity close to the diagonal peaks than the basic phase-sensitive COSY (left) as it does not suffer from broad, disposive diagonal peaks. 

Figure 5.46. The crosspeak structures from the phase-sensitive COSY spectrum of a three-spin AMX system. The arrows indicate splittings due to the labelled couplings. The spectrum was simulated with Jam = 18, Jax = 12 and Jmx = 6 Hz and the final digital resolution was 2.3 Hz/pt in each dimension.

In this magnitude COSY experiment it is possible to obtain sine and cosine modulated data with co-addition using a simple two step phase cycle and consequently the minimum number of scans per increment is 2 or for long acquisitions a multiple of 2. The simplicity of this pulse program is in contrast to the phase sensitive COSY experiment and other Bruker two dimension pulse programs which utilize phase increment commands such as ipl to achieve frequency discrimination in fl. 

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