Kinetic isotope experiments

On the other hand, the involvement of vinyl cationic species in the reaction cannot be ruled out in some cases, as shown in Scheme 4. In this context, it was found that the reaction of 3-butyn-2-one with mesitylene can occur without Pd(OAc)2, clearly indicating the involvement of vinyl cations generated from alkynes and H+ in this reaction.47 1 The yield difference in the presence and in the absence of Pd(OAc)2 may be explained by the competition between [Pd(n)02CF3]+ and vinyl cationic species in the electrophilic substitution of aromatic G-H bonds. Recent kinetic isotope experiments suggest a mechanism involving alkyne coordination to Pd(n) followed by electrophilic aromatic substitution.476... 

In a collaboration between the Abelson and Hecht labs [56b], a series of noncoded amino acids were introduced into dihydrofolate reductase (DHFR) to probe substrate binding and the requirement of an aspartic acid residue for catalytic competence. When aspartic acid analogs mono- or disubstituted at the )0-carbon were substituted for the active site aspartic acid residue, the mutant DHFRs were still able to catalyze the NADPH-dependent reduction of dihydrofolate to tetrahydrofolate at 74 - 86 % of the wild-type rate. While hydride transfer from NADPH is not the rate-limiting step for the wild-type enzyme at physiological pH, a kinetic isotope experiment with NADPD indicated that hydride transfer had likely become the rate-limiting step for the mutant containing the )0,)0-dimethylaspartic acid. 

Let us consider the classical secondary kinetic isotope experiments on lysozyme-catalyzed hydrolysis of glycosides. Earlier X-ray work indicated that the enzyme... 

Modifying the selectivity for a particular product is a more challenging task. To understand why Ag is the most selective catalyst for ethylene epoxidation, an highly important reaction practiced industrially for decades, Linic et al. performed detailed spectroscopic and kinetic isotope experiments and DFT calculations, and they concluded that the selectivity between the partial and total oxidation of ethylene on Ag(l 11) is controlled by the relative stability of two different transition states (TS s) that are both accessible to a common oxametallacycle intermediate One results in the closure of the epoxide ring and ethylene oxide (EO), while the other leads to acetaldehyde (AC) via intra-molecular H shift and eventually combustion. The authors... 

There is an important experimental distinction between and kinetic isotope experiments. Deuterium isotope effects are usually measured by comparing reciprocal plots from separate experiments with deuterium-labeled reactants and with unlabeled reactants, which allows determination of the V isotope effect from the ratio of intercepts and the V/K one from the ratio of slopes. In contrast to that, tritium isotope effects are measured by the internal competition method, that is, by changes in specific activity in product or residual substrate, and thus provide only the isotope effect on V/K. The kinetic isotope effects on V with tritium can be measured only with the aid of carrier-free tritiated molecules. 

Cases of this kind have been investigated in studies of anodic hydrocarbon oxidation/ While a priori calculations of the values of electrochemical kinetic Isotope effects, particularly with complex organic molecules, cannot be regarded as quantitatively reliable owing to the difficulty of evaluation of the relevant reactant and transition state partition functions for particles in solution, nevertheless sufficient experimental data exist which, coupled with theoretical evidence of trends of values of the isotope effect for different types of reaction pathway, enable useful distinctions in mechanism to be made on the basis of H/D kinetic isotope experiments on electrochemical reactions. 

In their paper describing the direct hydroxylation of arene sp C—H bonds with a ruthenium catalyst, Rao and co-workers demonstrated that a simple thiophene was also compatible with these reaction conditions (Scheme 10.21). It is proposed that under acidic conditions, [RuCl2(p-cymene)]2 facilitates C—H bond cleavage of 67 via an orthometalation process throngh chelation with the ester carbonyl group (kinetic isotope experiments also support a kinetically relevant C—H metalation step). Snbseqnent reductive elimination afforded the hydroxylated thiophene 68 in 41% yield. 

Solvolytic experiments specifically designed to test Bartell s theory were carried out by Karabatsos et al. (1967), who were primarily interested in an assessment of the relative contributions of hyperconjugation and non-bonded interactions to secondary kinetic isotope effects. Model calculations of the (steric) isotope effect in the reaction 2- 3 were performed, as well as that in the solvolyses of acetyl chloride... 

Labelling experiments provided the evidence that the Fe1- and Co1-mediated losses of H2 and 2H2 from tetralin are extremely specific. Both reactions follow a clear syn- 1,2-elimination involving C(i)/C(2) and C(3)/C(4), respectively. In the course of the multistep reaction the metal ions do not move from one side of the rr-surface to the other. The kinetic isotope effect associated with the loss of the first H2 molecule, k( 2)/k(Y)2) = 3.4 0.2, is larger than the KIE, WFLj/ATHD) = 1.5 0.2, for the elimination of the second H2 molecule. A mechanism of interaction of the metal ion with the hydrocarbon n-surface, ending with arene-M+ complex 246 formation in the final step of the reaction, outlined in equation 100, has been proposed241 to rationalize the tandem MS studies of the unimolecular single and double dehydrogenation by Fe+ and Co+ complexes of tetraline and its isotopomers 247-251. 

The results from these experiments also allowed Hannon and Traylor to determine the primary and secondary hydrogen deuterium kinetic isotope effects for the hydride abstraction reaction. If one assumes that there is no kinetic isotope effect associated with the formation of 3-deutero-l-butene, i.e. that CH2=CHCHDCH3 is formed at the same rate (k ) from both the deuterated and undeuterated substrate (Scheme 25), then one can obtain both the primary (where a deuteride ion is abstracted) and the secondary deuterium... 

Some care must be exercised in setting up the crossover equipment to account for kinetic isotope effects associated with abstraction or insertion into a carbon-hydrogen (deuterium) bond. In general, abstraction is expected to exhibit a larger isotope effect than insertion, and this appears to be the case (see below). To accommodate this, and to increase the sensitivity of the experiment, it is often necessary to employ a smaller amount of the hydrocarbon than of its deuteriated analog. 

Figures 7-9 show the fractional conversion of methanol in the pulse as a function of temperature for the three catalysts and the three methanol feeds. Evidently the kinetic isotope effect is present on all three catalysts and over the complete temperature range, indicating that the rate limiting step is the breaking of a carbon-hydrogen bond under all conditions. From these experiments, the effect cannot be determined quantitatively as in the case of the continuous flow experiments, but to obtain the same conversion of CD,0D, the temperature needs to be 50-60° higher. This corresponds to a factor of about three in reaction rate. The difference in activity between PfoCL and Fe.(MoO.), is larger in the pulse experiments compared to tHe steady stateJ results.

A suggested reaction mechanism based on kinetic and isotope experiments, product distributions, and the effect on these of addition of water to the feed or leaving oxygen out is shown in Figure 13. 

Experiments have implied that quantum mechanical effects can be involved in the proton and hydride transfer of several biological processes. Specifically, large kinetic isotope effects (KIE) observed for... 

Reduction of aldonolactones and their derivatives with isotopically modified reducing agents leads to sugars labeled at the anomeric center. Glycosides substituted with deuterium or labeled with tritium are widely employed for kinetic isotope-effect measurements, mechanistic studies, isotope-tracing experiments, and so on. 

Table 3 collects the results obtained by different authors on the reaction of 0s04(NH3) with ethylene. Though the methods were not identical, the results are very similar, and in all cases there is a clear preference of the [3+2] over the [2+2] pathway. Additional confirmation was provided by a combined experimental and theoretical study using kinetic isotope effects (KIEs) to compare experiment and theory. Kinetic isotope effects were measured by a new NMR technique [25] and compared to values, which are available from calculated transition states. It showed that indeed only the [3+2] pathway is feasible [10]. 

As mentioned earlier, products of disproportionation often accompany the rearrangement products. This reaction is also acid-catalysed and it is a reasonable assumption that reaction proceeds via the protonated species. Experiments with the 4,4 -diiodohydrazobenzene (19) showed that there were significant nitrogen and para-carbon kinetic isotope effects23. This implies that disproportionation must take place after C—C bonding has occurred, i.e. that the intermediate must be the quinonoid form 20 (and cannot, for example, be a jr-complex), which is then believed to react with another reactant molecule to give the disproportionation products (Scheme 4). 

As for all trace-level analyses, sample preparation and handling are of crucial importance. In addition to all the usual problems of GC-MS, measurements of isotope ratios must ensure that none of these steps introduce any isotope discrimination. Any chemical reactions, including conversion of the organic sample molecules to the simple gases which are those actually analyzed, must be quantitative (100% conversion) to avoid kinetic isotope effects [627]. Until relatively recently, all gas IRMS experiments employed a dual-inlet system to permit switching between sample and standard C02 contained in two bellows containers. The pressures in the two bellows are adjusted to be equal and,... 

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