Multiplicity selection

The insufficiency of a single physiological selection criterion for drought resistance has been indicated repeatedly. The most urgent problem in breeding for drought resistance is the construction of a multiple selection... 

The software tools accompanying the QTRAP MS/MS allow set-up of multiple selected reaction monitoring (SRM) transitions for all likely metabolites after the major product ion transitions for the dosed compound are known. Because QTRAP MS/MS can monitor up to 100 SRM transitions during a single assay, the SRM transitions required for quantitation of the dosed compound and internal standard are obtained along with the possible metabolite transitions. During sample analysis, when a possible metabolite transition exceeds a preset threshold value, the QTRAP MS/MS performs an enhanced product ion (EPI) scan. When the assay is complete, the EPI scans can be used to determine whether the hits are metabolites, and if they are metabolites, what part of the molecule has changed. Thus, one analytical run provides both quantitative and metabolite information. 

The desire to obtain structural and dynamical information from multiply labeled systems stems from the fact that such an approach alleviates the need to produce multiple selectively-labeled molecules. While the use of uniformly labeled samples simplifies the sam-... 

Test methods that analyze individual compounds (e.g., benzene-toluene-ethylbenzene-xylene mixtures and PAHs) are generally applied to detect the presence of an additive or to provide concentration data needed to estimate environmental and health risks that are associated with individual compounds. Common constituent measurement techniques include gas chromatography with second-column confirmation, gas chromatography with multiple selective detectors, and gas chromatography with mass spectrometry detection (GC/MS) (EPA 8240). 

A solution to improve the overall efficiency of selective ID and 2D experiments is described and demonstrated. It is based on the principle of multiple selective excitation with subsequent data processing to disentangle the superimposed responses of the selectively and simultaneously excited spin systems. Several inherent insensitive and correspondingly modified experiments are presented together with typical applications. 

Acquisition scheme for the multiple selective heteronuclear inverse detected ID COSY experiment. Four experiments have to be performed with 3 carbon resonances selected for selective perturbation. The frequencies are set either on-resonance (/i, fi, /j) or off-resonance to the selected resonance frequencies. 

Processing scheme for the multiple selective heteronuclear inverse detected ID COSY experiment. The final spectra A, B, C corresponding to the selective perturbation of three carbons resonating at frequencies fi, fi, and f are obtained by linear combination of the original data a, b, c and d acquired in accordance with the acquisition scheme (table 1). 

The principle of multiple selective excitation has been incorporated into a few ID and 2D experiments, the schemes of which are shown below (fig. 1). Depending on the experiment, either a DANTE pulse train (ID TOCSY [2]), frequency selective 180° pulses (ID NOE [3], ID INADEQUATE [4], ID C/H COSY [5] and 2D TOCSY-COSY [6]) or frequency selective 90° pulses (2D HMBC [11]) are applied to selectively perturb and uniquely label selected spins. Besides the DANTE pulse , composed itself of a series of non-selective rectangular pulses, Gaussian-shaped 180° and... 

Fig. 1. Pulse sequences modified for multiple selective excitation. I ID TOCSY, II het-eronuclear ID NOE, III ID INADEQUATE, IVa heteronuclear ID COSY (optimized to detect Jch), IVb heteronuclear ID COSY (optimized to detect "Jch), V 2D TOCSY-COSY, Via 2D HMBC (designed to detect heteronuclear long-range couplings "Jch only), VIh 2D HMBC (extended pulse sequence to detect both heteronuclear long-range "Jch and...

We have implemented the principle of multiple selective excitation (pulse sequence II in fig. 1) thereby replacing the low-power CW irradiation in the preparation period of the basic ID experiment by a series of selective 180° pulses. The whole series of selective pulses at frequencies /i, /2, , / is applied for several times in the NOE build-up period to achieve sequential saturation of the selected protons. Compared with the basic heteronuclear ID experiment, in this new variant the sensitivity is improved by the combined application of sequential, selective pulses and the more efficient data accumulation scheme. Quantitation of NOEs is no longer straightforward since neither pure steady-state nor pure transient effects are measured and since cross-relaxation in a multi-spin system after perturbation of a single proton (as in the basic experiment) or of several protons (as in the proposed variant) differs. These attributes make this modified experiment most suitable for the qualitative recognition of heteronuclear dipole-dipole interactions rather than for a quantitative evaluation of the corresponding effects. 

Fig. 5a. Part of the spectra obtained with the ID multiple selective INADEQUATE experiment III applied to cembrene 3 dissolved in CDCI3. The spectra were obtained after appropriate processing, showing the responses in the olefinic and the aliphatic regions after selective inversion of carbons...

Fig. 5b. Comparison of the results obtained with the basic (left) and the multiple selective (right) INADEQUATE experiment. The signals of C-10, C-16, C-17 and C-18 are shown and are compared with the responses measured with the modified experiment after perturbation of C-15. Corresponding satellite signals are connected by horizontal lines.

Fig. 6. Spectra of cinnamic aldehyde 4 dissolved in CDClj and obtained with the heteronu-clear multiple selective COSY experiments, (a) Spectra acquired with pulse sequence IVa dedicated to detect and measure Jen connectivities after selective perturbation of carbons C7, C3 and Cl. Spurious TOCSY peaks caused by short purge pulses (not shown in fig. I) are marked with an asterisk, (b) Spectra acquired with pulse sequence IVb dedicated to detect and measure " Jch connectivities after selective perturbation of carbons C-7, C-1 and C-4. Normal H spectra at the bottom.

---