Phosphate dianions, hydrolysis

D. Branduardi, M. De Vivo, N. Rega, V. Barone, and A. Cavalli,/. Chem. Theory Comput., 7(3), 539-543 (2011). Methyl Phosphate Dianion Hydrolysis in Solution Characterized by Path Collective Variables Coupled with DFT-Based Enhanced Sampling Simulations. 

A kinetic isotope effect 160/180 of 2% in the spontaneous hydrolysis of the 2,4-dinitrophenyl phosphate dianion, whose ester oxygen is labeled, suggests a P/O bond cleavage in the transition state of the reaction, and thus also constitutes compelling evidence for formation of the metaphosphate 66,67). The hydrolysis behavior of some phosphoro-thioates (110) is entirely analogous 68). 

Yet another situation is observed in the 2,4-dinitrophenyl phosphate dianion. A significant effect of amines on the rate of decomposition is admittedly observed however, typical 2nd order kinetics, lower enthalpy of activation compared with spontaneous hydrolysis, and strongly negative AS values (see Table 3) indicate an Sn2(P) reaction. Surprisingly, the reaction rate remains unaffected by the basicity of the amine, even when its pKa value changes by 8 units. 

Table 3. Activation parameters of spontaneous hydrolysis and second-order reactions with amines of the 2,4-dinitrophenyi phosphate dianion...

The highly electrophilic character of the POf ion would suggest a very unselective phosphorylation behavior. For example, the ratio of alkyl phosphate to inorganic phosphate obtained in hydrolyses of phosphoric esters in water/alcohol mixtures should reflect the molar ratio of water and alcohol. This is indeed found in numerous cases, e.g. in the hydrolysis of phenyl and 4-nitrophenyl phosphate monoanions 97) or of 4-nitrophenyl phosphate dianions 65) at 100 °C in methanol/ water mixtures of various compositions, as also in the solvolysis of the acetyl phosphate dianion at 37 °C 97) or of phosphoenol pyruvate monoanions 82). Calculations of the free energy of the addition reactions of water and ethanol to the POf ion support the energetic similarity of the two reactions 98) (Table 4). 

Reactions of 2,4-dinitrochloro-benzene and -naphthalene are speeded by DDDAOH and the corresponding chloride -I- NaOH (Cipiciani et at., 1984). The rate/surfactant concentration profiles and the rate constants are very similar to those for reactions in solutions of the corresponding C16 single chain surfactants which form normal micelles. The spontaneous hydrolysis of 2,4-dinitrophenyl phosphate dianion is also speeded by DDDAC1 and rates reach plateau values in very dilute surfactant (Savelli and Si, 1985). 

The iron(II)-iron(III) form of purple acid phosphatase (from porcine uteri) was kinetically studied by Aquino et al. (28). From the hydrolysis of a-naphthyl phosphate (with the maximum rate at pH 4.9) and phosphate binding studies, a mechanism was proposed as shown in Scheme 6. At lower pH (ca. 3), iron(III)-bound water is displaced for bridging phosphate dianion, but little or no hydrolysis occurs. At higher pH, the iron(III)-bound OH substitutes into the phosphorus coordination sphere with displacement of naphthoxide anion (i.e., phosphate hydrolysis). The competing affinity of a phosphomonoester anion and hydroxide to iron(III) in purple acid phosphatase reminds us of a similar competing anion affinity to zinc(II) ion in carbonic anhydrase (12a, 12b). 

Proton transfer may proceed directly or via a six-membered cyclic transition state involving a molecule of water. A calculation of the intermediate zwitter-ionic concentration for the hydrolysis of methyl phosphate monoanion, based on the pKa values for methanol and methyl phosphate dianion, predicts the first-order rate coefficient for zwitterion decomposition to be ca. 10 sec-1 at 100°C. This value is in good agreement with the observed rate of hydrolysis and, considering the assumptions involved, with the rate of P-O bond fission of the presumed zwitterionic intermediate (2) formed in the Hg(II) catalyzed solvolysis of phosphoenolpyruvic acid, a model reaction for pyruvate kinase10. 

Catalysis may also be observed via transition states or intermediates which are more than six-membered. An example is the hydrolysis of glucose-6-phosphate dianion which surprisingly is more rapid (5 times) than the monoanion. Presumably the relatively acidic 1-hydroxyl group of glucose (p/Ca = 10.8, 100°C) can act as a general-acid catalyst of phosphate group expulsion (58),3 > even though the required chair conformation has all substituents axial, viz. 

The pH-rate profile for unbuffered hydrolysis of glyceraldehyde-3-phosphate (6-3-P) has been attributed to hydrolysis of the monoanion of the phosphate monoester at pH < 4, spontaneous formation of glyceraldehyde from the phosphate dianion at pH 7-8, and, at higher pH, hydroxide-catalysed methylglyoxal formation. Reaction of the dianion is not subject to a solvent isotope effect and is believed to occur by the irreversible ElcB mechanism whereby an enediolate intermediate, formed on rate-determining C(2) deprotonation, subsequently expels phosphate trianion by C—0 bond breaking. The diethylacetal and 2-methyl-G-3-P do not hydrolyse under the same conditions.5... 

The electrostatic model for the micellar effect on the hydrolysis of phosphate monoesters is also consistent with the results of inhibition studies (Bunton et al., 1968, 1970). The CTAB catalyzed hydrolysis of the dinitrophenyl phosphate dianions was found to be inhibited by low concentrations of a number of salts (Fig. 9). Simple electrolytes such as sodium chloride, sodium phosphate, and disodium tetraborate had little effect on the micellar catalysis, but salts with bulky organic anions such as sodium p-toluenesulfonate and sodium salts of aryl carboxylic and phosphoric acids dramatically inhibited the micelle catalysis by CTAB. From equation 14 and Fig. 10, the inhibitor constants, K, were calculated (Bunton et al., 1968) and are given in Table 9. The linearity of the plots in Fig. 10 justifies the assumption that the inhibition is competitive and that incorporation of an inhibitor molecule in a micelle prevents incorporation of the substrate (see Section III). Comparison of the value of for phenyl phosphate and the values of K for 2,4-and 2,6-dinitrophenyl phosphates suggests that nitro groups assist the... 

Robinson, 1969a). It is probable that the hydrophobic nature of the phenyl groups of p-nitrophenyl diphenyl phosphate results in deep penetration of the neutral ester in the Stern layer, thus shielding the phosphoryl group from nucleophilic attack. Unlike other reactions between nucleophiles and neutral substrates catalyzed by cationic micelles (Bunton and Robinson, 1968, 1969a) and the hydrolysis of dinitrophenyl phosphate dianions in the presence of cationic micelles (Bunton et al., 1968), the catalysis of the hydrolysis of -nitrophenyl diphenyl phosphate by CTAB arises from an increase in the activation entropy rather than from a decrease in the enthalpy of activation. The Arrhenius parameters for the micelle-catalyzed and inhibited reactions are most probably manifestations of the extensive solubilization of this substrate. However, these parameters can be composites of those for the micellar and non-micellar reactions and the eifects of temperature on the micelles themselves are not known. Interpretation of the factors which affect these parameters must therefore be carried out with caution. In addition, the inhibition of the micelle-catalyzed reactions by added electrolytes has been observed (Bunton and Robinson, 1969a Bunton et al., 1969, 1970) and, as in the cases of other anion-molecule reactions and the heterolysis of dinitrophenyl phosphate dianions, can be reasonably attributed to the exclusion of the nucleophile by the anion of the added salt. 

Like other esters, phosphates undergo hydrolysis to the parent acid and alcohol. Here, the acidity of —OH attached to phosphorus has several effects. In the first place, since acidic phosphate esters can undergo ionization, there may be many species present in the hydrolysis solution. A monoalkyl ester, for example, could exist as dianion, monoanion, neutral ester, and protonated ester any or all of these could conceivably be undergoing hydrolysis. Actually, the situation is not quite that complicated. From the dissociation constants of these acidic esters, one can calculate the fraction of ester in each form in a given solution. The dependence of rate on acidity of the solution often shows which species is the principal reactant. 

The data and mechanistic conclusions summarized above come from work with aryl phosphomonoesters as predicted by the steep jSlg value, alkyl ester dianions react at very slow rates. A recent study of methyl phosphate found the rate of the dianion hydrolysis to be below the threshold of detectability, with an estimated rate constant of 2 x 10 20 s 1 at 25 °C.3 Since this value is close to the rate predicted from an extrapolation of the Bronsted plot of aryl phosphomonoester dianions, a similar mechanism is likely to be followed for alkyl and aryl esters. 

---