Sensitivity enhancement methods pulse sequence

The INEPT (Insensitive Nuclei Enhanced by Polarization Transfer) experiment [6, 7] was the first broadband pulsed experiment for polarization transfer between heteronuclei, and has been extensively used for sensitivity enhancement and for spectral editing. For spectral editing purposes in carbon-13 NMR, more recent experiments such as DEPT, SEMUT [8] and their various enhancements [9] are usually preferable, but because of its brevity and simplicity INEPT remains the method of choice for many applications in sensitivity enhancement, and as a building block in complex pulse sequences with multiple polarization transfer steps. The potential utility of INEPT in inverse mode experiments, in which polarization is transferred from a low magnetogyric ratio nucleus to protons, was recognized quite early [10]. The principal advantage of polarization transfer over methods such as heteronuclear spin echo difference spectroscopy is the scope it offers for presaturation of the unwanted proton signals, which allows clean spec-... 

The former is a protein of 14.7 kDa involved in the multienzyme nucleotide excision repair (NER) pathway with a determined NMR solution structure . In this protein, the Zn + possesses rather a structural than a catalytic role. Zn NMR spectra were acquired using a rather sophisticated probe (for details, see Reference 87) and operating at temperatures 5-250 K. Data acquisition was performed with the application of spin-echo methods for enhanced sensitivity . Specifically, experiments were carried out at 25 K using a combination of CP (cross-polarization) and spikelet echo pulse sequences which provide a considerable increase in signal-to-noise ratio (of the order of 30) relative to a classical quadrupole echo pulse sequence. The proton field strength applied to the above measurements was 60 kHz with a matching field of 20 kHz for zinc and a contact time... 

Pulse sequences for non-selective polarization transfer, not only useful for signal enhancement but also for multiplicity selection, are referred to as INEPT [54], abbreviated from Insensitive Nuclei Enhanced by Polarization Transfer . An improved method denoted as Distortionless Enhancement by Polarization Transfer or DEPT" [55] permits the cleanest multiplicity selection known so far, with full enhancement and low sensitivity to individual CH coupling constants. In addition, fully enhanced and undistorted coupled spectra can be recorded. Finally, subspectra for CH, CH2 and CH3 groups can be generated. 

The development of carbon-13 NMR during the last eight years has been characterized by a continual increase in the sensitivity and quality of spectra. A reduction in measuring time - equivalent to an enhancement in sensitivity has been achieved mainly by cryomagnet technology. The efficiency with which NMR information can be obtained has been substantially improved by new computer-controllable pulse sequences for one-and two-dimensional NMR experiments. A selection of these new methods, in particular, those used for multiplicity analysis and homo- or heteronuclear shift correlations, is presented in chapter 2 of this edition. 

The second (sine) term produced by the evolution delay has all the information we need—it is antiphase 13C coherence labeled with the 13C chemical shift in t —but it is lost because its phase (S ) causes it to be unaffected by the 90° 13C pulse at the end of t. Effectively we are throwing away half of our signal at this point. A new method was developed to save this wasted signal and boost the sensitivity of HSQC and many other experiments. These modified pulse sequences are called sensitivity enhanced or sensitivity improved (Bruker adds si to the pulse sequence name) and the strategy is called preservation of equivalent pathways (PEP) because the two terms are equivalent except for their phase. 

Other strategies that show great promise in reducing NMR acquisition time utilise methods to obtain multiple sets of data from one experiment through a concept known as time-shared evolution. An example of this process that should find utility in natural products elucidation was demonstrated by a pulse sequence called CN-HMBC.93 Traditionally, a separate 13C-HMBC and 15N-HMBC were acquired independently. However, the CN-HMBC allows both 13C- and 15N-HMBC spectra to be obtained simultaneously. By acquiring both data sets simultaneously, an effective 50% time reduction can be achieved.93 This approach has also been demonstrated for a sensitivity-enhanced 2D HSQC-TOCSY (heteronuclear multiple bond correlation total correlation spectroscopy) and HSQMBC (heteronuclear single quantum... 

The projection-reconstruction approach is a technique unrelated to covariance processing which can provide data typically inaccessible to the natural product chemist. For example, 13C-15N correlation spectra were obtained for vitamin B12 at natural abundance.104 Compared with a conventional three-dimensional 13C-15N correlation experiment, the projection-reconstruction method provides a sensitivity enhancement of two orders of magnitude. The final 13C-15N spectrum was reconstructed from data obtained from ll l5N and H- C correlations acquired using a time-shared evolution pulse sequence that allowed all the information to be obtained in one experiment.104 Martin and co-workers also demonstrated the ability to generate 13C-15N correlation spectra using unsymmetrical indirect covariance NMR with vinblastine as an example.105 In the latter case, 13C-15N correlation spectra were obtained from - C HSQC data and H-1sN HMBC data that were acquired separately. Both methods provide access to correlations that would be inaccessible for most natural products at natural abundance. 

It is 9 years since the publication of the first edition of this book and in this period the discipline of NMR spectroscopy has continued to develop new methodology, improve instrumentation and expand in its applications. This second edition aims to reflect the key developments in the field that have relevance to the stmcture elucidation of small to mid-sized molecules. It encompasses new and enhanced pulse sequences, many of which build on the sequences presented in the first edition, offering the chemist improved performance, enhanced information content or higher-quality data. It also includes coverage of recent advances in NMR hardware that have led to improved instrument sensitivity and thus extended the boundaries of application. Many of the additions to the text reflect incremental developments in pulsed methods and are to be found spread across many chapters, whereas some of the more substantial additions are briefly highlighted below. 

While the SPT method, as the name indicates, is a selective experiment, techniques derived from the principle of population transfer of a non-selective nature are also availahle. The first of these to appear was the INEPT experiment [38]. The pulse sequence schematics for INEPT and refocused INEPT are shown in Pig. 8.1. As with the SPT experiment described above, INEPT and its successor, the DEPT ex-p)eriment, both operate with enhanced sensitivity through the perturhation of the Boltzmann populations. This prototypical non-selective experiment is now relatively seldom used as more refined variants have been developed. The INEPT experiment (r = Fig. 8.1A) generates antiphase responses analogous to... 

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