Kinetics of the Hydride Reduction of an NAD⁺ Analogue by Isopropyl Alcohol in Aqueous and Acetonitrile Solutions: Solvent Effects, Deuterium Isotope Effects, and Mechanism

Abstract

The rate constants of the hydride-transfer reactions from isopropyl alcohol (i-PrOH) to an NAD(+) model, 9-phenylxanthylium ion (PhXn(+)), in acetonitrile (AN) and in water containing AN (80% H(2)O/20% AN) were determined over a temperature range from 36 to 67 degrees C. The reactions follow second-order rate laws. In the latter solution, formation of the water adduct of PhXn(+) was observed as a side-equilibrium (K). The observed inverse solvent kinetic isotope effect (k(H(2)O)(obs)/k(D(2)O)(obs) = 0.54), the larger than unity equilibrium isotope effect (K(H(2)O)/K(D(2)O) = 2.69), and the results of acid effect on the observed rate constants of the reactions are consistent with the "side-equilibrium mechanism". Kinetic isotope effects at all three H/D positions of i-PrOH for the net hydride-transfer process were determined in both solutions at 60 degrees C: KIE(alpha-D)(H) = 3.2(AN), 3.2(H(2)O); KIE(beta-D6)(H) = 1.05(AN), 1.16(H(2)O); KIE(OD)(H) = 1.08(AN), 1.04(H(2)O). These KIE values are consistent with the presence of the positively charged alcohol moiety in the transition state (TS) for cleavage of the alpha-C-H bond, the delocalization of the positive charge over the alpha-C-OH group, and the stepwise hydride and proton transfer processes. Comparison of the activation parameters for the reactions in the two solvent systems as well as those in the i-PrOH/AN (1:1 v/v) reported earlier suggests that the AN medium promotes the reaction by activating the ground-state alcohol reactant through weak interactions with the electron pairs on alcohol O, while water and parent alcohol media facilitate the reaction by H-bonding stabilization of the alcohol moiety of the TS. Results suggest that in the alcohol dehydrogenases without a Zn(II) cofactor in the active sites alcohols would be oxidized via hydride transfer to NAD(+) coenzyme followed by the rapid deprotonation to the nearby basic species in the active site of the enzymes.

Structure–reactivity relationship for alcohol oxidations via hydride transfer to a carbocationic oxidizing agent

“…In order to further confirm the general hydride‐proton sequential transfer mechanism for this kind of hydride‐transfer reactions, KIEs at the α‐H(D), β‐CH 3 (CD 3 ), and OH(D) positions of the 1‐phenylethanol were determined by comparing the pseudo‐first‐order rate constants of the reactions of alcohols containing both isotopes under the same conditions. Values of primary KIE α‐D , secondary KIE β‐D3 , and secondary KIE OD are listed in Table 1, consistent with the corresponding KIEs observed for the 2‐propanol reaction in the same solvent (also listed in Table 1 for comparison)33 and indicating a stepwise mechanism (1) (see the subsequent discussion). Additionally, this research group is currently working on the study of temperature dependence of α‐D KIEs for some of the reactions studied in this paper, and has found that the KIE α‐D are all primary, further suggesting a rate‐limiting hydride transfer mechanism for this class of reactions.…”

“…We have recently reported the kinetic and mechanistic study of the oxidation of 2‐propanol via hydride‐transfer to carbocationic oxidants (R + ) in various solvent systems to form the corresponding ketone product, the hydride reduction products of carbocations (RH) and a proton (Eqn (1), R 1 = R 2 = CH 3 ) 31–33. R + used include PhXn + and 10‐methylacridinium ion.…”

“…In this paper, kinetics of hydride‐transfer reactions from alcohols to PhXn + in AN were determined spectroscopically (UV–Vis) by following the decay of the latter reactant at 373 nm with time. The detailed method to determine the pseudo‐first‐order rate constants ( k obs ) of the reaction of 2‐propanol in AN can be found in our previous work 33. Kinetics of the reactions of all alcohols with PhXn + (1.0 × 10 −3 M) were determined under pseudo‐first‐order conditions with alcohol concentrations being in large excess (0.015–0.5 M).…”

“…Product analysis for the reaction of 1‐phenylethanol was carried out; corresponding PhXnH and acetophenone products were isolated and their structures were characterized by 1 H NMR spectroscopy. Here, absorption decay at 373 nm was due to the PhXn + consumption and absorption rise at 285 nm due to the formation of PhXnH 32, 33. Representative kinetic scans for the reactions of other alcohols are shown in the Supplemental Information (Figs.…”

Imbalanced tunneling ready states in alcohol dehydrogenase model reactions: rehybridization lags behind H-tunneling

“…[24][25][26][27] Here, kinetics of the hydride-transfer reactions from alcohols to Xn + ClO 4 À in MeCN were similarly determined by following the decay of the Xn + UV-Vis absorption (Fig. S1, ESIw).…”