Apparent isotope effects

Figure 5A shows experimentally derived profiles of pH vs rate for reactions in H2O and D2O [30, 50, 71]. The magnitude of the apparent isotope effect (ratio of rate constants in H2O and D2O) is 4.4 and the profiles appear to support the possibility that a proton is transferred from (Mg -bound) water molecules. However, careful analysis led us to conclude that a metal ion binds directly to the 5 -oxygen. Since the concentration of the deproto-nated 2 -oxygen in H2O should be higher than that in D2O at a fixed pH, we must take into account this difference in pKa, namely ApKa (=pKa °-pKa ), when we analyze the solvent isotope effect of D2O [30, 50, 68, 71]. We can estimate the pKa in D2O from the pKa in H2O using the linear relationship shown in Fig. 5B [30, 68, 73-75]. If the pKa for a Mg -bound water molecule in H2O is 11.4, the ApKa is calculated to be 0.65 (solid line in Fig. 5B). Then, the pKa in D2O should be 12.0. Demonstrating the absence of an intrinsic isotope effect (kH2o/kD20=l)> the resultant theoretical curves closely fit the experimental data, with an approximate 4-fold difference in...

In addition to the reaction sequence shown, the substrate may have to mutarotate non-enzymically to the form favored for affording the first enzyme-substrate complex. In such multi-stage reactions, the observed isotope-effects for the overall reaction are usually low. The situation is further complicated by possible differences in Km between a deuterated substrate and the unlabeled substrate, so that, unless a full kinetic analysis of the reaction is performed, the values determined for the apparent isotope-effect may vary. 

Small isotope-effects can be detected by double-labeling techniques, in which the carbon skeleton is labeled with 14C, and the ratio of 14C to tritium is measured both in the substrate and the product. Care must be taken in the observation and interpretation of isotope effects determined from the hydrogen-isotope content of the product. Just as in non-enzymic reactions (see p. 154), discrimination against the substrate containing deuterium or tritium leads to an increase in the isotopic content of the substrate, and this decreases the apparent isotope-effect towards the end of the reaction. 

The apparent isotope effect a in Eq. (78) is related to the isotope effects in forward and reverse reactions by... 

It can be seen that the apparent isotope effect on V/K varies inversely with the ratio between values of 1 and the true intrinsic isotope effect. When... 

The mechanism for this reaction is not of the Langmuir-Hinshelwood form, but rather involves the reaction between a surface-bonded NO molecule with an NO molecule in the solution phase along with a simultaneous electron transfer. The Tafel slope in acidic solution is (120 mV), which implies that the first electron-transfer step is rate-determining (see Addendum to this chapter). The reaction is first order in H+ and shows no apparent isotope effect. The following mechanism has been proposed ... 

Bowman WR, Bruce IT, Kirby GW (1969) Biosynthetic incorporatitm of [p- C, 3,5- H2, 4- H]-cinnamic acid into capsaicin and norpluviine lack of an apparent isotope effect following an NIH shift. J Chem Soc Chem Commun 1075-1077... 

What is the apparent isotope effect on for the reaction mechanism given in Exercise 6.9.4 ... 

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