An altered mechanism of hydrolysis for a metal-complexed phosphate diester

Isotope effects in the nucleophile and in the leaving group were measured to gain information about the mechanism and transition state of the hydrolysis of methyl p-nitrophenyl phosphate complexed to a dinuclear cobalt complex. The complexed diester undergoes hydrolysis about 1011 times faster than the corresponding uncomplexed diester. The kinetic isotope effects indicate that this rate acceleration is accompanied by a change in mechanism. A large inverse 18O isotope effect in the bridging hydroxide nucleophile (0.937 +/- 0.002) suggests that nucleophilic attack occurs before the rate-determining step. Large isotope effects in the nitrophenyl leaving group (18Olg = 1.029 +/- 0.002, 15N = 1.0026 +/- 0.0002) indicate significant fission of the P-O ester bond in the transition state of the rate-determining step. The data indicate that in contrast to uncomplexed diesters, which undergo hydrolysis by a concerted mechanism, the reaction of the complexed diester likely proceeds via an addition-elimination mechanism. The rate-limiting step is expulsion of the p-nitrophenyl leaving group from the intermediate, which proceeds by a late transition state with extensive bond fission to the leaving group. This represents a substantial change in mechanism from the hydrolysis of uncomplexed aryl phosphate diesters.     

Multiple isotope effect study of the acid-catalyzed hydrolysis of methyl formate

Multiple isotope effects have been measured for the acid-catalyzed hydrolysis of methyl formate in 0.5 M HCl at 20 degrees C. The isotope effects in the present investigation include the carbonyl carbon (13k = 1.028 +/- 0.001), the carbonyl oxygen (18k = 0.9945 +/- 0.0009), the nucleophile oxygen (18k = 0.995 +/- 0.001), and the formyl hydrogen ((D)k = 0.81 +/- 0.02). Determination of the carbonyl carbon, carbonyl oxygen, and formyl hydrogen isotope effects was performed via isotopic analysis of residual substrate. However, determination of the oxygen nucleophile isotope effect required analysis of the oxygen atoms of the product (formic acid), which exchange with the solvent (water) under acid conditions. This necessitated measurement of the rate of exchange of these oxygen atoms under the conditions for hydrolysis (k(ex) = 0.0723 min(-1)) and correction of the raw isotope ratios measured during the nucleophile-O isotope effect experiment. These results, along with the previously reported isotope effect for the leaving oxygen (18k = 1.0009) and the ratio of the rate of hydrolysis to that of exchange of the carbonyl oxygen with water (k(h)/k(ex) = 11.3), give a detailed picture of the transition-state structure for the reaction.     

Hydrolysis of phosphotriesters: determination of transition states in parallel reactions by heavy-atom isotope effects

The remote label method was used to measure primary and secondary (18)O isotope effects in the alkaline hydrolysis of O,O-diethylphosphorylcholine iodide (DEPC) and the primary (18)O effect in the alkaline hydrolysis of O,O-diethyl-m-nitrobenzyl phosphate (DEmNBP). Both the leaving group of interest (choline or m-nitrobenzyl alcohol) and ethanol can be ejected during hydrolysis due to the similarity of their pK values. The heavy-atom isotope effects were measured by isotope ratio mass spectrometry. Parallel reaction and incomplete labeling corrections were made for both systems. DEPC has a primary (18)O isotope effect of 1.041 +/- 0.003 and a secondary (18)O isotope effect of 1.033 +/- 0.002. The primary (18)O isotope effect for DEmNBP was 1.052 +/- 0.003. These large effects suggest a highly associative transition state in which the nucleophile approaches very close to the phosphorus atom to eject the leaving group. The large values are also indicative of a large compression, or general movement, on the reaction coordinate.     

A concerted mechanism for the transfer of the thiophosphinoyl group from aryl dimethylphosphinothioate esters to oxyanionic nucleophiles in aqueous solution

Earlier work on the hydrolysis of aryl phosphinothioate esters has led to contradictory mechanistic conclusions. To resolve this mechanistic ambiguity, we have measured linear free energy relationships (beta(nuc) and beta(lg)) and kinetic isotope effects for the reactions of oxyanions with aryl dimethylphosphinothioates. For the attack of nucleophiles on 4-nitrophenyl dimethylphosphinothioate, beta(nuc) = 0.47 +/- 0.05 for phenoxide nucleophiles (pK(a) < 11) and beta(nuc) = 0.08 +/- 0.01 for hydroxide and alkoxide nucleophiles (pK(a) >or= 11). Linearity of the plot in the range that straddles the pK(a) of the leaving group (4-nitrophenoxide, pK(a) 7.14) is indicative of a concerted mechanism. The much lower value of beta(nuc) for the more basic nucleophiles reveals the importance of a desolvation step prior to rate-limiting nucleophilic attack. The reactions of a series of substituted aryl dimethylphosphinothioate esters give the same value of beta(lg) with the nucleophiles HO(-) (beta= -0.54 +/- 0.03) and PhO(-) (beta = -0.52 +/- 0.09). A significantly better Hammett correlation is obtained with sigma(-) than with sigma or sigma degrees , as expected for a transition state involving rate-limiting cleavage of the P-OAr bond. The (18)O KIE at the position of bond fission ((18)k = 1.0124 +/- 0.0008) indicates the P-O bond is approximately 40% broken, and the (15)N KIE in the leaving group ((15)k = 1.0009 +/- 0.0003) reveals the nucleofuge carries about a third of a negative charge in the transition state. Thus, both the LFER and KIE data are consistent with a concerted reaction and disfavor a stepwise mechanism.     

Nucleophilic participation in the transition state for human thymidine phosphorylase

Recombinant human thymidine phosphorylase catalyzes the reaction of arsenate with thymidine to form thymine and 2-deoxyribose 1-arsenate, which rapidly decomposes to 2-deoxyribose and inorganic arsenate. The transition-state structure of this reaction was determined using kinetic isotope effect analysis followed by computer modeling. Experimental kinetic isotope effects were determined at physiological pH and 37 degrees C. The extent of forward commitment to catalysis was determined by pulse-chase experiments to be 0.70%. The intrinsic kinetic isotope effects for [1'-(3)H]-, [2'R-(3)H]-, [2'S-(3)H]-, [4'-(3)H]-, [5'-(3)H]-, [1'-(14)C]-, and [1-(15)N]-thymidines were determined to be 0.989 +/- 0.002, 0.974 +/- 0.002, 1.036 +/- 0.002, 1.020 +/- 0.003, 1.061 +/- 0.003, 1.139 +/- 0.005, and 1.022 +/- 0.005, respectively. A computer-generated model, based on density functional electronic structure calculations, was fit to the experimental isotope effect. The structure of the transition state confirms that human thymidine phosphorylase proceeds through an S(N)2-like transition state with bond orders of 0.50 to the thymine leaving group and 0.33 to the attacking oxygen nucleophile. The reaction differs from the dissociative transition states previously reported for N-ribosyl transferases and is the first demonstration of a nucleophilic transition state for an N-ribosyl transferase. The large primary (14)C isotope effect of 1.139 can occur only in nucleophilic displacements and is the largest (14)C primary isotope effect reported for an enzymatic reaction. A transition state structure with substantial bond order to the attacking nucleophile and leaving group is confirmed by the slightly inverse 1'-(3)H isotope effect, demonstrating that the transition state is compressed by the impinging steric bulk of the nucleophile and leaving group.     

A theoretical investigation of alpha-carbon kinetic isotope effects and their relationship to the transition-state structure of S(N)2 reactions

[reaction: see text] The transition structures and alpha-carbon 12C/13C kinetic isotope effects for 22 S(N)2 reactions between methyl chloride and a wide variety of nucleophiles have been calculated using the B1LYP/aug-cc-pVDZ level of theory. Anionic, neutral, and radical anion nucleophiles were used to give a wide range of S(N)2 transition states so the relationship between the magnitude of the alpha-carbon kinetic isotope effect and transition-state structure could be determined. The results suggest that the alpha-carbon 12C/13C kinetic isotope effects for S(N)2 reactions will be large (near the experimental maximum) and that the curve relating the magnitude of the KIE to the percent transfer of the alpha-carbon from the nucleophile to the leaving group in the transition state has a broad maximum. This means very similar KIEs will be found for early, symmetric, and late transition states and that one cannot use the magnitude of these KIEs to estimate transition-state structure.     

A kinetic isotope effect study on the hydrolysis reactions of methyl xylopyranosides and methyl 5-thioxylopyranosides: oxygen versus sulfur stabilization of carbenium ions.

The following kinetic isotope effects, KIEs (k(light)/k(heavy)), have been measured for the hydrolyses of methyl alpha- and beta-xylopyranosides, respectively, in aqueous HClO(4) (mu = 1.0 M, NaClO(4)) at 80 degrees C: alpha-D, 1.128 +/- 0.004, 1.098 +/- 0.005; beta-D, 1.088 +/- 0.008, 1.042 +/- 0.004; gamma-D(2), (C5) 0.986 +/- 0.001, 0.967 +/- 0.003; leaving-group (18)O, 1.023 +/- 0.002, 1.023 +/- 0.003; ring (18)O, 0.983 +/- 0.001, 0.978 +/- 0.001; anomeric (13)C, 1.006 +/- 0.001, 1.006 +/- 0.003; and solvent, 0.434 +/- 0.017, 0.446 +/- 0.012. In conjunction with the reported (J. Am. Chem. Soc. 1986, 108, 7287-7294) KIEs for the acid-catalyzed hydrolysis of methyl alpha- and beta-glucopyranosides, it is possible to conclude that at the transition state for xylopyranoside hydrolysis resonance stabilization of the developing carbenium ion by the ring oxygen atom is coupled to exocyclic C-O bond cleavage, and the corresponding methyl glucopyranosides hydrolyze via transition states in which charge delocalization lags behind aglycon departure. In the analogous hydrolysis reactions of methyl 5-thioxylopyranosides, the measured KIEs in aqueous HClO(4) (mu = 1.0 M, NaClO(4)) at 80 degrees C for the alpha- and beta-anomers were, respectively, alpha-D, 1.142 +/- 0.010, 1.094 +/- 0.002; beta-D 1.061 +/- 0.003, 1.018(5) +/- 0.001; gamma-D(2), (C5) 0.999 +/- 0.001, 0.986 +/- 0.002; leaving-group (18)O, 1.027 +/- 0.001, 1.035 +/- 0.001; anomeric (13)C, 1.031 +/- 0.002, 1.028 +/- 0.002; solvent, 0.423 +/- 0.015, 0.380 +/- 0.014. The acid-catalyzed hydrolyses of methyl 5-thio-alpha- and beta-xylopyranosides, which occur faster than methyl alpha- and beta-xylopyranosides by factors of 13.6 and 18.5, respectively, proceed via reversibly formed O-protonated conjugate acids that undergo slow, rate-determining exocyclic C-O bond cleavage. These hydrolysis reactions do not have a nucleophilic solvent component as a feature of the thiacarbenium ion-like transition states.     

Intrinsic primary and secondary hydrogen kinetic isotope effects for alanine racemase from global analysis of progress curves

The pyridoxal phosphate dependent alanine racemase catalyzes the interconversion of L- and D-alanine. The latter is an essential component of peptidoglycan in cell walls of Gram-negative and -positive bacteria, making alanine racemase an attractive target for antibacterials. Global analysis of protiated and deuterated progress curves simultaneously enables determination of intrinsic kinetic and equilibrium isotope effects for alanine racemase. The intrinsic primary kinetic isotope effects for Calpha hydron abstraction are 1.57 +/- 0.05 in the D --> L direction and 1.66 +/- 0.09 in the L --> D direction. Secondary kinetic isotope effects were found for the external aldimine formation steps in both the L --> D (1.13 +/- 0.05, forward; 0.90 +/- 0.03, reverse) and D --> L (1.13 +/- 0.06, forward; 0.89 +/- 0.03, reverse) directions. The secondary equilibrium isotope effects calculated from these are 1.26 +/- 0.07 and 1.27 +/- 0.07 for the L --> D and D --> L directions, respectively. These equilibrium isotope effects imply substantial ground-state destabilization of the C-H bond via hyperconjugation with the conjugated Schiff base/pyridine ring pi system. The magnitudes of the intrinsic primary kinetic isotope effects, the lower boundary on the energy of the quinonoid intermediate, and the protonation states of the active site catalytic acids/bases (K39-epsilonNH2 and Y265-OH) suggest that the pKa of the substrate Calpha-H bond in the external aldimine lies between those of the two catalytic bases, such that the proton abstraction transition state is early in the D --> L direction and late in the L --> D direction.     

Comparisons of phosphorothioate with phosphate transfer reactions for a monoester,diester, and triester: isotope effect studies

Phosphorothioate esters are sometimes used as surrogates for phosphate ester substrates in studies of enzymatic phosphoryl transfer reactions. To gain better understanding of the comparative inherent chemistry of the two types of esters, we have measured equilibrium and kinetic isotope effects for several phosphorothioate esters of p-nitrophenol (pNPPT) and compared the results with data from phosphate esters. The primary (18)O isotope effect at the phenolic group ((18)k(bridge)), the secondary nitrogen-15 isotope effect ((15)k) in the nitro group, and (for the monoester and diester) the secondary oxygen-18 isotope effect ((18)k(nonbridge)) in the phosphoryl oxygens were measured. The equilibrium isotope effect (EIE) (18)k(nonbridge) for the deprotonation of the monoanion of pNPPT is 1.015 +/- 0.002, very similar to values previously reported for phosphate monoesters. The EIEs for complexation of Zn(2+) and Cd(2+) with the dianion pNPPT(2-) were both unity. The mechanism of the aqueous hydrolysis of the monoanion and dianion of pNPPT, the diester ethyl pNPPT, and the triester dimethyl pNPPT was probed using heavy atom kinetic isotope effects. The results were compared with the data reported for analogous phosphate monoester, diester, and triester reactions. The results suggest that leaving group bond fission in the transition state of reactions of the monoester pNPPT is more advanced than for its phosphate counterpart pNPP, while alkaline hydrolysis of the phosphorothioate diester and triester exhibits somewhat less advanced bond fission than that of their phosphate ester counterparts.     

Nucleophilic attack on phosphate diesters: a density functional study of in-line reactivity in dianionic, monoanionic, and neutral systems

A density functional study of the hydrolysis reaction of phosphodiesters with a series of attacking nucleophiles in the gas phase and in solution is presented. The nucleophiles HOH, HO-, CH3OH, and CH3O- were studied in reactions with ethylene phosphate, 2'3'-ribose cyclic phosphate and in their neutral (protonated) and monoanionic forms. Stationary-point geometries for the reactions were determined at the density functional B3LYP/6-31++G(d,p) level followed by energy refinement at the B3LYP/6-311++G(3df,2p) level. Solvation effects were estimated by using a dielectric approximation with the polarizable continuum model (PCM) at the gas-phase optimized geometries. This series of reactions characterizes factors that influence the intrinsic reactivity of the model phosphate compounds, including the effect of nucleophile, protonation state, cyclic structure, and solvent. The present study of the in-line mechanism for phosphodiester hydrolysis, a reaction of considerable biological importance, has implications for enzymatic mechanisms. The analysis generally supports the associative mechanism for phosphate ester hydrolysis. The results highlight the importance for the reaction barrier of charge neutralization resulting from the protonation of the nonbridging phosphoryl oxygens and the role of internal hydrogen transfer in the gas-phase mechanism. It also shows that solvent stabilization has a profound influence on the relative barrier heights for the dianionic, monoanionic, and neutral reactions. The calculations provide a comprehensive data set for the in-line hydrolysis mechanisms that can be used for the development of improved semiempirical quantum models for phosphate hydrolysis reactions.     

Kinetics and mechanism of the aminolysis of aryl phenyl chlorothiophosphates with anilines

Kinetic studies of the reactions of aryl phenyl chlorothiophosphates (1) and aryl 4-chlorophenyl chlorothiophosphates (2) with substituted anilines in acetonitrile at 55.0 degrees C are reported. The negative values of the cross-interaction constant rhoXY (rhoXY = -0.22 and -0.50 for 1 and 2, respectively) between substituents in the nucleophile (X) and substrate (Y) indicate that the reactions proceed by concerted SN2 mechanism. The primary kinetic isotope effects (kH/kD = 1.11-1.13 and 1.10-1.46 for 1 and 2, respectively) involving deuterated aniline nucleophiles are obtained. Front- and back-side nucleophilic attack on the substrates is proposed mainly on the basis of the primary kinetic isotope effects. A hydrogen-bonded, four-center-type transition state is suggested for a front-side attack, while the trigonal bipyramidal pentacoordinate transition state is suggested for a back-side attack. The MO theoretical calculations of the model reactions of dimethyl chlorothiophosphate (1') and dimethyl chlorophosphate (3') with ammonia are carried out. Considering the specific solvation effect, the front-side nucleophilic attack can occur competitively with the back-side attack in the reaction of 1'.     

Multiple isotope effect study of the acid-catalyzed hydrolysis of formamide

Multiple isotope effects were measured at the reactive center of formamide during acid-catalyzed hydrolysis in water at 25 degrees C. The mechanism involves a rapid pre-equilibrium protonation of the carbonyl oxygen, followed by the formation of at least one tetrahedral intermediate, which does not appreciably exchange its carbonyl oxygen with the solvent (kh/kex = 55). The pKa for formamide was determined by 15N NMR and found to be about -2.0. The formyl-hydrogen kinetic isotope effect (KIE) is indicative of a transition state that is highly tetrahedral (Dkobs = 0.79); the carbonyl-carbon KIE (13kobs = 1.031) is in agreement with this conclusion. The small leaving-nitrogen KIE (15kobs = 1.0050) is consistent with some step prior to breaking the C-N bond as rate-determining. The carbonyl-oxygen KIE (18kobs = 0.996) points to attack of water as the rate-determining step. On the basis of these results, a mechanism is proposed in which attachment of the nucleophile to a protonated formamide molecule is rate determining.     

Activation of acyl phosphate monoesters by lanthanide ions: enhanced reactivity of benzoyl methyl phosphate

Acyl phosphate monoesters are intermediates in many biochemical acylation reactions, such as those involving aminoacyl adenylates. Benzoyl methyl phosphate, a typical acyl phosphate monoester, is slowly hydrolyzed in neutral solutions but reacts rapidly with amines. Since biochemical processes of acyl phosphate monoesters involve accelerated reactions with oxygen-centered nucleophiles, we sought catalysts for hydrolysis and methanolysis of benzoyl methyl phosphate to mimic the biochemical outcome. Lanthanide ions are particularly effective catalysts, accelerating reactions much more than comparable levels of magnesium ion. Detailed kinetic analysis of the hydrolysis reactions reveals formation of a 1:1 complex, followed by rapid reaction with a nucleophile. The hydroxide-dependent hydrolysis rate in the europium complex is about 10(5) times that of free substrate with hydroxide. A mechanism that accounts for the data and observed behavior involves bidentate coordination of the metal ion by the acyl phosphate through phosphate and carbonyl oxygens, lowering the energy of the tetrahedral addition intermediate and the associated transition states. The dependence of the metal ion catalyzed process on the concentration of hydroxide ion is consistent with coordinated hydroxide acting as a nucleophile. The reaction of benzoyl methyl phosphate with methanol to form methyl benzoate and methyl phosphate is 30 000 times more rapid in the presence of 0.0001 M lanthanum triflate (in the absence of the metal ion k(obs) = 2.1 x 10(-7) s(-1), at 25 degrees C). Thus, the combination of acyl phosphate esters and lanthanide salts appears to be a promising method for biomimetic acylation of hydroxyl groups.     

Experimental and theoretical multiple kinetic isotope effects for an SN2 reaction. An attempt to determine transition-state structure and the ability of theoretical methods to predict experimental kinetic isotope effects

The secondary alpha-deuterium, the secondary beta-deuterium, the chlorine leaving-group, the nucleophile secondary nitrogen, the nucleophile (12)C/(13)C carbon, and the (11)C/(14)C alpha-carbon kinetic isotope effects (KIEs) and activation parameters have been measured for the S(N)2 reaction between tetrabutylammonium cyanide and ethyl chloride in DMSO at 30 degrees C. Then, thirty-nine readily available different theoretical methods, both including and excluding solvent, were used to calculate the structure of the transition state, the activation energy, and the kinetic isotope effects for the reaction. A comparison of the experimental and theoretical results by using semiempirical, ab initio, and density functional theory methods has shown that the density functional methods are most successful in calculating the experimental isotope effects. With two exceptions, including solvent in the calculation does not improve the fit with the experimental KIEs. Finally, none of the transition states and force constants obtained from the theoretical methods was able to predict all six of the KIEs found by experiment. Moreover, none of the calculated transition structures, which are all early and loose, agree with the late (product-like) transition-state structure suggested by interpreting the experimental KIEs.     

Transition state differences in hydrolysis reactions of alkyl versus aryl phosphate monoester monoanions

Although aryl phosphates have been the subject of numerous experimental studies, far less data bearing on the mechanism and transition states for alkyl phosphate reactions have been presented. Except for esters with very good leaving groups such as 2,4-dinitrophenol, the monoanion of phosphate esters is more reactive than the dianion. Several mechanisms have been proposed for the hydrolysis of the monoanion species. (18)O kinetic isotope effects in the nonbridging oxygen atoms and in the P-O(R) ester bond, and solvent deuterium isotope effects, have been measured for the hydrolysis of m-nitrobenzyl phosphate. The results rule out a proposed mechanism in which the phosphoryl group deprotonates water and then undergoes attack by hydroxide. The results are most consistent with a preequilibrium proton transfer from the phosphoryl group to the ester oxygen atom, followed by rate-limiting P-O bond fission, as originally proposed by Kirby and co-workers in 1967. The transition state for m-nitrobenzyl phosphate (leaving group pK(a) 14.9) exhibits much less P-O bond fission than the reaction of the more labile p-nitrophenyl phosphate (leaving group pK(a) = 7.14). This seemingly anti-Hammond behavior results from weakening of the P-O(R) ester bond resulting from protonation, an effect which calculations have shown is much more pronounced for aryl phosphates than for alkyl ones.     

Transition states for glucopyranose interconversion

Glucose is a central molecule in biology and chemistry, and the anomerization reaction has been studied for more than 150 years. Transition-state structure is the last impediment to an in-depth understanding of its solution chemistry. We have measured kinetic isotope effects on the rate constants for approach of alpha-glucopyranose to its equilibrium with beta-glucopyranose, and these were converted into unidirectional kinetic isotope effects using equilibrium isotope effects. Saturation transfer 13C NMR spectroscopy has yielded the relative free energies of the transition states for the ring-opening and ring-closing reactions, and both transition states contribute to the experimental kinetic isotope effects. Both transition states of the anomerization process have been modeled with high-level computational theory with constraints from the primary, secondary, and solvent kinetic isotope effects. We have found the transition states for anomerization, and we have also concluded that it is forbidden for the water molecule to form a hydrogen bond bridge to both OH1 and O5 of glucose simultaneously in either transition state.     

Supermolecule density functional calculations suggest a key role for solvent in alkaline hydrolysis of p-nitrophenyl phosphate

Supermolecule density functional theory calculations show that solvent is responsible for the concerted transition state in alkaline hydrolysis of p-nitrophenyl phosphate suggested by heavy atom kinetic isotope effects.     

The effect of solvent on the structure of the transition state for the S(N)2 reaction between cyanide ion and ethyl chloride in DMSO and THF probed with six different kinetic isotope effects

The secondary alpha- and beta-deuterium, the alpha-carbon, the nucleophile carbon, the nucleophile nitrogen, and the chlorine leaving group kinetic isotope effects for the S(N)2 reaction between cyanide ion and ethyl chloride were determined in the very slightly polar solvent THF at 30 degrees C. A comparison of these KIEs with those reported earlier for the same reaction in the polar solvent DMSO shows that the transition state in THF is only slightly tighter with very slightly shorter NC-C(alpha) and C(alpha)-Cl bonds. This minor change in transition state structure does not account for the different transition structures that were earlier suggested by interpreting the experimental KIEs and the gas-phase calculations, respectively. It therefore seems unlikely that the different transition states suggested by the two methods are due to the lack of appropriate solvent modeling in the theoretical calculations. Previously it was predicted that the transition state of S(N)2 reactions where the nucleophile and the leaving group have the same charge would be unaffected by a change in solvent. The experimental KIEs support this view.     

Proton inventory study of the base-catalyzed hydrolysis of formamide. Consideration of the nucleophilic and general base mechanisms

An NMR study of the rates of hydroxide-promoted hydrolysis of formamide in aqueous media of varying mole fraction D(2)O (n) was performed at [LO(-)] = 1.42 M, T = 25 degrees C, to shed light on whether the mechanism involves a nucleophilic attack of HO(-) on the C=O or HO(-) acting as a general base to remove a proton from an attacking water. The solvent deuterium kinetic isotope effect under these conditions is inverse, k(OH)/k(OD) = 0.77 +/- 0.02 or k(OD)/k(OH) = 1.30 +/- 0.03. Proton inventory analysis of the k(n)() versus n data was undertaken through NLLSQ fits to equations representing four possible mechanisms encompassing nucleophilic and general base ones with waters of solvation on the attacking hydroxide, and with or without waters of solvation on the developing amide hydrate oxyanion. Both nucleophilic and general base mechanisms can be accommodated, but there are restraints on each that are discussed. The preferred mechanism is a nucleophilic one proceeding through a transition state having two solvating waters remaining on the attacking hydroxide and three additional waters attached to the developing amide hydrate oxyanion.     

Kinetic and mechanistic studies of the reaction of a range of bases and metal-hydroxo complexes with the phosphonate ester 2,4-dinitrophenyl ethyl methylphosphonate in aqueous solution

The base hydrolysis and spontaneous water hydrolysis of the phosphonate ester 2,4-dinitrophenyl ethyl methylphosphonate (DNPEMP) has been studied in detail and the activation parameters for these processes determined. The catalytic effect of a series of 19 oxygen and nitrogen bases and metal hydroxo complexes has been studied. The Brønsted plot of pK versus logkN has a slope (β) of 0.47, which is fairly typical for phosphate and phosphonate esters. alpha-Effect nucleophiles such as 2-iodosobenzoate, hypochlorite and the hydroperoxide ion (HO-2) exhibit enhanced reactivity, as does fluoride ion and the bifunctional metal complex [Cu(tmen) (OH2)(OH)]+ (tmen = N,N,N',N',-tetramethylethylenediamine). Reactions involving these species appear to take place by a nucleophilic pathway, while the other oxygen and nitrogen bases act as general base catalysts. The solvent deuterium isotope effect for Et3N catalysis, kH2O/kD2O=1.3, is consistent with general base catalysis. The mechanisms of these reactions are discussed in detail.