Nuclear magnetic resonance spectroscopy isotopically labelled

The mechanism outlined above is supported by experimental findings. An intermediate 5 has been isolated, " and has been identified by and N-nuclear magnetic resonance spectroscopy. Side-products have been isolated, which are likely to be formed from intermediate 4. N-isotope labeling experiments have shown that only the nitrogen remote from the phenyl group is eliminated as ammonia. 

Probably, one of the most valuable advances in this field has dealt with the first chemoenzymatic synthesis of the stable isotope-enriched heparin from a uniformly double labelled 13C, 15N /V-acetylheparosan from E. coli K5. Heteronuclear, multidimensional nuclear magnetic resonance spectroscopy was employed to analyze the chemical composition and solution conformation of N-acety 1 hcparosan, the precursors, and heparin. Isotopic enrichment was found to provide well-resolved 13C spectra with the high sensitivity required for conformational studies of these biomolecules. Stable isotope-labelled heparin was indistinguishable from heparin derived from animal tissues and might be employed as a novel tool for studying the interaction of heparin with different receptors.30... 

Hunger and Wang provide an account of advances in the characterization of solid catalysts in the functioning state by nuclear magnetic resonance spectroscopy. Examples include investigations of zeolite-catalyzed reactions with isotopic labels that allow characterization of transition states and reaction pathways as well as characterization of organic deposits, surface complexes, and reaction intermediates formed in catalyst pores. 

McInnes, A.G., D.G. Smith, L.C. Vining, and L. Johnson Use of in Biosynthetic Studies. Location of Isotope from Labelled Acetate and Formate in the Fungal Tropolone, Sepedonin, by Nuclear Magnetic Resonance Spectroscopy. Chem. Commun. 1971, 325. 

Stereochemical and kinetic analyses of the Brpnsted acid-catalysed intramolecular hydroamination/deuterioamination of the electronically non-activated cyclic alkene (13) with a neighbouring sulfonamide nucleophile have been found to proceed as an anh-addition (>90%) across the C=C bond to produce (15). No loss of the label was observed by and NMR (nuclear magnetic resonance) spectroscopies and mass spectrometry (MS). The reaction follows the second-order kinetic law rate = 2 [TfOH] [13] with the activation parameters being = 9.1 0.5 kcal moP and = -35 5 cal moP An inverse a-secondary kinetic isotope effect of d/ h = (1-15 0.03), observed for (13) deuteration at C(2), indicates a partial CN bond formation in the transition state (14). The results are consistent with a mechanism involving concerted, intermolecular proton transfer from an N-protonated sulfonamide to the alkenyl C(3) position coupled with an intramolecular anti-addition by the sulfonamide group. ... 

Branched fatty acids, known as iso-acids and anteiso-acids, occur normally in small quantities in fats. Their synthesis begins with the amino-acids valine and isoleucine (Figure 3.12). This has been demonstrated with radio-labelled isotopes, by radio-active monitoring and with stable isotopes by C nuclear magnetic resonance spectroscopy or mass spectrometry. Both isobutyric acid and 2-methylbutyric acid are common defensive compounds among insects. Note that a chiral centre is introduced in 2-methylbutyric acid and anteiso acids. 

As the glucose is metabolized, the metabolic derivatives become enriched with the isotopic carbon label and are detected using advanced technologies such as mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy. Importantly, incorporation of isotopes into molecules can alter reaction mechanisms including the rate of a reaction-this is termed isotopic substitution and forms the basis of the kinetic isotope effect. 

The study of biological macromolecules by nuclear magnetic resonance (NMR) spectroscopy has been greatly expanded with the use of isotopic labeling (51). 

The elucidation of the biosynthetic pathway for the production of various metabolites has been extensively examined through the use of techniques that use isotopic labeling (stable isotopes and radioactive isotopes). Initially, radiolabeled precursors were introduced into plants and the resultant radioactive compounds were chemically degraded to identify the positions of the label. As the development of analytical instrumentation advanced, the isotopically labeled natural products were analyzed by mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy instead of chemical degradation. 

The measurement of internal metabolic fluxes is more difficult. The direct measurement of intracellular fluxes is possible with in vivo nuclear magnetic resonance (NMR) spectroscopy. However, the inherent insensitivity of NMR limits its applicability. An improvement over this approach can be found with isotopic tracer techniques [8]. In isotope tracer methods the cells to be studied are provided with a substrate specifically labeled with a detectable isotope (usually or C). The incorporation of label into cellular material and by-products is governed by the fluxes through the biochemical pathways. The quantity and distribution of label is measured and combined with knowledge of the metabolic network to estimate some of the intracellular fluxes. The choices of substrate labeling patterns, as well as which by-products to measure, are guided by careful analysis of the assumed biochemical network. These experiments are usually performed at isotopic steady state so that the flow of isotope into each atom of a metaboHte equals the flux out. For the nth atom of the fcth metabolite the flux balance is [9] ... 

Phosphorous nuclear magnetic resonance ( P-NMR) No isotope labeling is required for P-NMR spectroscopy. The chemical shielding anisotropy, Aa, in P-NMR is comparable to the deuterium quadrupole splitting in H-NMR and can be determined from the edges of the spectrum. 

The usual method of study is to suggest a possible precursor and to feed it to the biosynthesizing system. The precursor has to be labelled in some way to trace it through the sequence of reactions, and that is usually by some isotopic element. It may be a radio-active isotope, such as H, " 0, or that can be followed by its radiation or it can be a stable heavy isotope, such as H, C, N, or 0, that can be traced by mass spectrometry or nuclear magnetic resonance (NMR) spectroscopy (Table 5.1). Another possible way is to use mutant strains of an organism that lack the enzymes to complete a particular synthesis, or to add a specific enzyme inhibitor, so that intermediates accumulate and can be identified. A mutant strain of yeast was important in discovering mevalonic acid and its place in terpene biosynthesis (Chapter 6) and a number of mutants of the bacterium Escherichia coli helped to understand the shikimic acid pathway (Chapter 8). 

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