Intermolecular Isotope Effects Research Articles

Microbial models of mammalian metabolism: involvement of cytochrome P450 in the N-demethylation of N-methylcarbazole by Cunninghamella echinulata.

1. As previously reported (Yang and Davis 1992), N-methylcarbazole (NMC) is converted to N-hydroxymethylcarbazole (NHMC), and 3-hydroxy-N-hydroxymethylcarbazole (3-OH-NHMC), two relatively stable carbinolamine metabolites by the fungus Cunninghamella echinulata (ATCC 9244). Decomposition of these two carbinolamines yields the corresponding dealkylated metabolites, carbazole and 3-hydroxycarbazole. In the present study, the possible involvement of cytochrome P450 in the requisite N-alkyl hydroxylation reaction was examined. 2. Carbon monoxide, a classical P450 inhibitor, markedly inhibited the formation of NHMC, as did potassium cyanide. 1-Benzylimidazole, piperonyl butoxide and SKF-525A inhibited the formation of both NHMC and 3-OH-NHMC, while beta-naphthoflavone (5,6-benzoflavone) induced their formation. 3. The source of the oxygen atom in the metabolite NHMC was examined by GC/MS analysis of NHMC formed during incubation of NMC in H218O-enriched medium which resulted in no incorporation of labelled oxygen into the metabolite. 4. An intermolecular isotope effect was not observed for the formation of NHMC suggesting that C-H bond cleavage is not a rate limiting step in the formation of this metabolite under the conditions examined. 5. It was concluded that P450 enzymes may be involved in the N-demethylation of NMC catalyzed by this fungal model of mammalian metabolism, and provides further support for biochemical and mechanistic parallels between mammalian metabolism and microbial systems catalyzing phase-1 biotransformations.

On the mechanism of oxidation of cholesterol at C-7 in a lipoxygenase system.

Incubation of [7-2H2]cholesterol with soybean lipoxygenase and linoleic acid in the presence of oxygen gave a mixture of 5-cholestene-3 beta,7 alpha-diol, 5-cholestene-3 beta,7 beta-diol, 3 beta-hydroxy-5-cholesten-7-one,5 alpha,6 alpha-epoxycholestan-3 beta-ol, and 5 beta,6 beta-epoxycholestan-3 beta-ol. The conversion into the 7-oxygenated products was associated with a very high intermolecular isotope effect (KH/KD = 15-17), suggesting that the rate-limiting step in the overall conversion is likely to be the abstraction of hydrogen at C-7 in a radical reaction. Evidence that linoleic acid is to some extent directly involved was obtained with the use of [7-3H]cholesterol. Incubation of [7-3H]cholesterol resulted in a significant incorporation of 3H in the reisolated linoleic acid fraction. The isotope effect associated with conversion of [7 alpha-2H]cholesterol into 7-oxygenated products in the lipoxygenase system was 2-3, indicating that the extraction of hydrogen is nonstereospecific. Incubation of [7-2H2]cholesterol with 13-hydroperoxy-9,11-octadecadienoic acid gave the above 7-oxygenated products with relatively small isotope effects (KH/KD = 3-4). It is concluded that the most important mechanism for oxidation of cholesterol at C-7 in the lipoxygenase system involves participation of radicals and that a carbon-centered linoleic acid radical can extract hydrogen directly from cholesterol. Fatty acid hydroperoxides and their secondary products seem to be less important as initiators in connection with oxidation of cholesterol.

ChemInform Abstract: C‐H Insertion and π‐Complex Formation Reactions of (η5‐C5Me5)(PMe3)Ir with Ethylene: An Intra‐ and Intermolecular Isotope Effect Study.

AbstractThermolysis of (I) in the presence of ethylene (II) leads to a mixture of (III) ( P21/n, Z=4) and (IV).

ChemInform Abstract: Kinetic H/D Isotope Effects for Gas Phase Hydroxylation of Benzene and Chlorobenzene Between 520‐1080 K. Hydroxyl Radical versus O(3P) Atom Attack

Gas phase slow combustion of (chloro)benzene in O2/N2 mixtures, and induced by addends such as tert butylhydroperoxide, cyclohexane, or methanol, leads to (chloro)-phenol as the only important aromatic product. Using C6H6/C6D6 mixtures, formation of phenol/perdeuterophenol was studied between 520–1080 K. The temperature dependence of this product ratio was found to obey the Arrhenius expression for the intermolecular isotope effect log kH/kD = −0.14 ± 0.03 + (1240 ± 80)/2.303RT (R in cal/mol K). Essentially the same result was obtained for the intramolecular isotope effect, measuring the change in isomer distribution for the chlorophenols formed from p-deuterio-chlorobenzene versus those for chlorobenzene. These results are in accordance with H(D)-abstraction by ·OH, via a linear transition state, as the first and (relative) rate determining step. Whereas above 1000 K, at reduced pressure, the intramolecular isotope effect continues to prevail, C6H6/C6D6 do not show differences in rate of formation of C6H5OH/C6D5OH. Under these conditions, the only effective reaction of arene to phenol appears to be set in by addition of O(3P).

Carbon-hydrogen insertion and .pi.-complex formation reactions of (.eta.5-C5Me5)(PMe3)Ir with ethylene: an intra- and intermolecular isotope effect study

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTCarbon-hydrogen insertion and .pi.-complex formation reactions of (.eta.5-C5Me5)(PMe3)Ir with ethylene: an intra- and intermolecular isotope effect studyPage O. Stoutland and Robert G. BergmanCite this: J. Am. Chem. Soc. 1988, 110, 17, 5732–5744Publication Date (Print):August 1, 1988Publication History Published online1 May 2002Published inissue 1 August 1988https://doi.org/10.1021/ja00225a025RIGHTS & PERMISSIONSArticle Views436Altmetric-Citations112LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (2 MB) Get e-Alerts Get e-Alerts

Kinetic H/D isotope effects for gas phase hydroxylation of benzene and chlorobenzene between 520–1080 K. Hydroxyl radical versus O(3P) atom attack

AbstractGas phase slow combustion of (chloro)benzene in O2/N2 mixtures, and induced by addends such as tert butylhydroperoxide, cyclohexane, or methanol, leads to (chloro)‐phenol as the only important aromatic product. Using C6H6/C6D6 mixtures, formation of phenol/perdeuterophenol was studied between 520–1080 K. The temperature dependence of this product ratio was found to obey the Arrhenius expression for the intermolecular isotope effect log kH/kD = −0.14 ± 0.03 + (1240 ± 80)/2.303RT (R in cal/mol K). Essentially the same result was obtained for the intramolecular isotope effect, measuring the change in isomer distribution for the chlorophenols formed from p‐deuterio‐chlorobenzene versus those for chlorobenzene.These results are in accordance with H(D)‐abstraction by ·OH, via a linear transition state, as the first and (relative) rate determining step. Whereas above 1000 K, at reduced pressure, the intramolecular isotope effect continues to prevail, C6H6/C6D6 do not show differences in rate of formation of C6H5OH/C6D5OH. Under these conditions, the only effective reaction of arene to phenol appears to be set in by addition of O(3P).

Regioselectivity and Deuterium Isotope Effects in Geraniol Hydroxylation by the Cytochrome P‐450 Monooxygenase from Catharanthus roseus (L.) G. DON

AbstractThe hydroxylation of geraniol (8) by cytochrome P‐450 (P‐450Cath.) from the subtropical plant Catharanthus roseus (L.) G. DON was optimised to give 8‐hydroxygeraniol (9) as the single product in 35% yield. Incubations of different 13C‐ and 2H‐labelled geraniols revealed that H‐abstraction is completely regioselective in favour of the CH3 group trans to the chain at C(6) of 8. An intramolecular isotope effect kH/kD = 8.0 was determined, suggesting that H‐abstraction is one of the major rate‐contributing steps; however, the intermolecular isotope effect was surprisingly inverse at low conversion kH/kD = 0.50, indicating the existence of rate‐contributing steps preceding the first irreversible, isotope‐sensitive reaction in the sequence.

Spin-orbit state-selected reactions of Kr+(2P3/2 and 2P1/2) with H2, D2, and HD from thermal energies to 20 eV c.m.

Spin-orbit state-selected reactions of Kr+(2PJ), J=3/2 and J=1/2, with isotopic molecular hydrogen (H2, D2, and HD) to form KrH+ and KrD+ are investigated using guided ion beam techniques. Reaction cross sections for each spin-orbit state are measured as a function of the relative translational energy of the reactants from near thermal energies up to 20 eV c.m. At low energies, the cross sections for reaction with H2 and D2 are each about 2.6 times larger for the 2P3/2 ground spin-orbit state of krypton ion than for the 2P1/2 excited state. For the HD reaction, the 2P3/2 reactivity is enhanced by a factor of about 4.2. A higher-energy process, which exhibits an apparent activation energy of about 1 eV, is present only for the 2P3/2 spin-orbit state. The Kr+(2P3/2) spin-orbit state exhibits a strong intermolecular isotope effect, showing the unusual dependence σ(HD)>σ(H2)>σ(D2) for the total reaction cross sections at low energies. Cross sections for the Kr+(2P1/2) state show a much smaller dependence on the hydrogen isotope. The intramolecular isotope effect for the individual KrH+ and KrD+ channels in the reaction with HD shows several reversals over the energy range studied. These results are discussed in terms of the potential energy surfaces of the krypton–hydrogen system.

C+(2P)+H2(D2,HD)→CH+(CD+)+H(D). I. Reaction cross sections and kinetic isotope effects from threshold to 15 eV c.m.

Total cross sections for the reactions of carbon(1+) ions with isotopic molecular hydrogen (H2, D2, and HD) to form methyliumylidene (CH+ and CD+) have been measured using guided ion beam techniques. Cross sections are reported as a function of the translational energy of the reactants from the reaction threshold up to 15 eV c.m. The true cross sections are shown to rise sharply from thresholds given by the thermochemical endothermicities. Inter- and intramolecular isotope effects in the threshold region can be attributed to the different endothermicities due to vibrational zero-point energies. At higher energies, an unexpected intermolecular isotope effect is found. Thermal reaction rates (300 K) derived from the data are 1.2×10−16 for H2, 2.3×10−17 for D2, and 1.2×10−16 for HD (17% CH+, 83% CD+), all in units of cm3 s−1.

Recoil tritium reactions with molecular hydrogen. Part 3.—Isotope effects

Scavenger and moderation studies on T + HD and T + H2/D2 mixtures in the presence of ICl scavenger and the moderators He, Ar and Kr are reported. Kinetic-theory analysis of the product yields leads to similar conclusions to those reported for the T + H2 and T + D2 systems. However, the isotope effects are found to depend on the nature of the moderator species, and the intermolecular isotope effect is found to be significantly smaller (ca. 1) than that determined from the separate, single-reactant systems, or that predicted from theoretical excitation functions.

Determination of intra- and intermolecular tritium isotope effects in the reaction of thymine 7-hydroxylase

Different [7- 3H]thymine preparations have been used to determine the inter- and intramolecular isotope effects of the 2-oxoglutarate-dependent thymine hydroxylation, catalyzed by thymine 7-hydroxylase (thymine, 2-oxoglutarate:oxygen oxidoreductase, EC 1.14.11.6). Specific activity ratios of products, viz., 3H 2O and 5-hydroxymethyluracil, and remaining substrate to initial substrate have been determined. The influence on these ratios of intra- and intermolecular isotope effects at different degrees of tritium substitution has been analyzed. An intramolecular isotope effect with a k H k T of about 6.5 has been found. No intermolecular isotope effect of T V K could be detected when oxygen concentration was varied from 0.4 to 0.01 mM. This agrees with a mechanism in which 2-oxoglutarate is irreversibly changed before the bond-breaking in thymine takes place.

Kinetic isotope effects in cytochrome P-450-catalyzed oxidation reactions. Intermolecular and intramolecular deuterium isotope effects during the N-demethylation of N,N-dimethylphentermine.

Two N,N-dimethylphentermine (N,N-dimethyl-2-amino-2-methyl-3-phenylpropane) substrates differing in deuterium substitution have been used to determine the intermolecular and intramolecular isotope effects associated with the cytochrome P-450-dependent N-demethylation of this substrate. No intermolecular isotope effect was observed in Vmax or Vmax/Km when the reaction rates for this substrate were compared to those for the substrate in which both N-methyl groups contained deuterium. In contrast, identical isotope effects of 1.6 to 2.0 were observed in both Vmax and Vmax/Km when this reaction was studied with a substrate in which only one of the two N-methyl groups was substituted with deuterium. Furthermore, both the intermolecular and intramolecular isotope effects were independent of the cytochrome P-450/NADPH-cytochrome P-450 reductase mole ratio. From these data, it is concluded that: 1) the carbon-hydrogen bond cleavage step does not contribute significantly to Vmax; 2) the contribution of the carbon-hydrogen bond cleavage step to Vmax is not detectably increased through changes in the cytochrome P-450/NADPH-cytochrome P-450 reductase mole ratio; 3) the N-methyl groups are free to exchange at the enzyme active site. The basis for these conclusions is the proposal of a new kinetic model for interpretation of intramolecular isotope effects which shows that intramolecular isotope effects are not necessarily equal to intrinsic isotope effects and, in fact, may be much smaller.

Intramolecular 13C kinetic isotope effects in the decarboxylation of oxalic acid in dioxane solution

Intramolecular 13C kinetic isotope effects in the decomposition in dioxane solution of oxalic acid to carbon dioxide and formic acid were determined at several temperatures between 102 and 141°C. The isotope effect is small (approximately 0.5%) and any temperature dependence is masked by the experimental imprecision (which averages about 0.09%). The method of three-element reaction coordinates (with ν1‡=0) was employed to expose possible properties of the transition state. Relatively good definition of transition state properties is obtained with several models. But, when one requires simultaneous fit of the model calculations to the results recorded here and to the intermolecular isotope effects reported earlier (’’double correspondence’’), the parameters are severly confined. The best double correspondence is obtained when: (a) transition state force constants are set to correspond to bond orders (near the C atom which will appear in carbon dioxide product) of C–C near 0.1, of C–O near 1.9, with that of O–H set at 0.5; (b) the relative internal coordinate displacements comprising the reaction coordinate eigenvector are C–C extension, 0.50 to 0.75; C–O contraction, 0.15 to 0.18; and O–H extension, 0.35 to 0.07. Anharmonicity and other small corrections are neglected without substantial effect on the conclusions described.

ChemInform Abstract: SECONDARY DEUTERIUM ISOTOPE EFFECTS ON CARBONYL YLIDE INTERMEDIATES

The formation of vinyldihydrofuran (6) and dihydrooxepine (5) is sensitive to deuterium substitution at the terminal methylene of 2,3-divinyloxirane. Intra- and intermolecular isotope effects have been examined and suggest that 5 is produced on a competitive basis with 6 from a common intermediate.

Secondary Deuterium Isotope Effects on Carbonyl Ylide Intermediates

The formation of vinyldihydrofuran (6) and dihydrooxepine (5) is sensitive to deuterium substitution at the terminal methylene of 2,3-divinyloxirane. Intra- and intermolecular isotope effects have been examined and suggest that 5 is produced on a competitive basis with 6 from a common intermediate.

ChemInform Abstract: REACTION OF SINGLET OXYGEN WITH 2‐METHYLNORBORN‐2‐ENE, 2‐METHYLIDENENORBORNANE, AND THEIR 7,7‐DIMETHYL DERIVATIVES, THE TRANSITION STATE GEOMETRY FOR HYDROPEROXIDATION

AbstractDie Bestrahlung des Norbornens (Ia) in Acetonitril in Gegenwart von Sauerstoff und Methylenblau als Sensibilisator und nachfolgende Reduktion mit NaBH4 liefert hauptsächlich das exo‐Norbornanol (IIa) und daneben sein endo‐Isomeres (IIIa).

Reaction of Singlet Oxygen with 2‐Methylnorborn‐2‐ene, 2‐Methylidenenorbornane, and their 7,7‐Dimethyl Derivatives. The transition state geometry for hydroperoxidation

AbstractThe dye‐sensitized photo‐oxygenation of 2‐methylnorbonr‐2‐ene (3), 2‐methylidene‐norbornane (4) and their 7,7‐dimethyl derivatives (5 and 6) has been studied. In all cases allylically rearranged hydroperoxides were formed, except that 4 also gave a little norbornanone (presumably from the dioxetane) and 5 gave some endo‐3,7,7‐trimethylnorbornan‐2‐one as a secondary photo‐product. It was found that the exo/endo attack ratios by singlet oxygen on 3 and 5 are 66 and 0.19. By exploiting the C (3) monodeuteriated derivatives, 4 and 6 showed ratios of 28 and 0.67.Rates of reactivity of the olefins 3 and 4 were compared to methylidenecyclopentane, 1‐methylcyclopentene and 1‐methylcyclohexene as monocyclic standards. Additionally, comparative rates between the 7,7‐dimethyl olefins and their parents were measured. When further comparison was made of the rate ratios partitioned for exo and endo attack, it was seen that oxygen experienced a 500‐ to 1000‐fold rate retardation on approach to the endo side of 3 compared to that for its monocyclic analogue. Exo rates between the parent norbornene 3 and its 7,7‐dimethyl derivative 5 showed a 250‐fold decrease. Although four times smaller than the difference reported for epoxidation, the evidence clearly pointed to a one‐step cyclic process as the rate determining step for photo‐oxygenation. The steric evidence, taken with the low values found for the intermolecular isotope effects of 1.14 ± 0.01 and 1.02 ± 0.01 observed for exo and endo‐3‐deuterio‐2‐methylidene‐norbornanes, permits the deduction that the transition state is largely dipolar. In the early stages of the addition bonding between one end of the oxygen molecule and the terminal vinyl carbon is advanced. At the same time positive charge is dispersed by hyperconjugation between the central carbon atom and the allylic carbon‐hydrogen bond. At a later stage the anionic oxygen atom abstracts the loosened allylic hydrogen atom to create the hydroperoxide. No evidence for the formation of a discrete perepoxide intermediate was obtained.

Intramolecular kinetic isotope effect in the reaction of fluorine atoms with methane-d2 (CH2D2)

The relative rates of the two competing isotopic reactions: F + CH2D2 → HF + CHD2 and F + CH2D2 → DF + CH2D, were measured in a fast flow system by direct mass spectrometric analysis of the reaction products in the temperature range of 159° to 298°K. The ratio of rates can be expressed as an Arrhenius type equation: kF+HCHD2/kF+DCH2D = (0.81 ± 0.03) exp[(275 ± 15)/R T]. The relation between the intramolecular isotope effect kF+HCHD2/kF+DCH2D and the intermolecular isotope effect kF+CH4/kF+CD4 is discussed with regard to secondary isotope effects in the fluorine atom + methane reaction.

Effect of partial deuteration on triplet-state lifetimes

Studies of the effect of the number and position of deuterium substituents on the phosphorescence lifetime reveal a significant dependence of the rate of radiationless decay on the positions as well as the number of protons. A theoretical interpretation of the effect of the number and position of deuteration, the intermolecular isotope effect, and the solvent effect on the triplet state lifetimes is given. The experimental results on benzene, naphthalene, and rare earth ions are discussed. For benzene, and naphthalene, the positions of protons of the triplet state are not equivalent and hence the corresponding CH stretching modes are not equivalent. These modes are grouped into two classes; one class is more effective in the electronic relaxation than the other. Under certain conditions the non-radiative constant is linear with respect to the number of modes of each class.

Deuterium Isotope Effects on the Lifetime of the Phosphorescent Triplet State of Naphthalene

The effect of the number and position of deuterium substituents on the phosphorescence lifetime has been studied for eight different isotopic naphthalenes at 76°K. These studies reveal a significant dependence of the rate of radiationless decay to the ground state on the positions of the protons in the molecule. This conclusion is contrary to the conclusion of Lin and Bersohn that the radiationless decay is independent of the positions of the protons. It is found that protons in the α positions are about 1.6 times as effective as protons in the β positions in promoting radiationless transitions. A simple empirical relation is found to give a reasonable fit to the data. A small intermolecular isotope effect on the triplet-state lifetime of naphthalene-d8 in ethanol is also found upon deuteration of the solvent. The experimental results are discussed in terms of existing theories of radiationless processes.