Cationic Reverse Micelles Create Water with Super Hydrogen‐Bond‐Donor Capacity for Enzymatic Catalysis: Hydrolysis of 2‐Naphthyl Acetate by α‐Chymotrypsin

Reverse micelles (RMs) are very good nanoreactors because they can create a unique microenvironment for carrying out a variety of chemical and biochemical reactions. The aim of the present work is to determine the influence of different RM interfaces on the hydrolysis of 2‐naphthyl acetate (2‐NA) by α‐chymotrypsin (α‐CT). The reaction was studied in water/benzyl‐n‐hexadecyldimethylammonium chloride (BHDC)/benzene RMs and, its efficiency compared with that observed in pure water and in sodium 1,4‐bis‐2‐ethylhexylsulfosuccinate (AOT) RMs. Thus, the hydrolysis rates of 2‐NA catalyzed by α‐CT were determined by spectroscopic measurements. In addition, the method used allows the joint evaluation of the substrate partition constant K_p between the organic and the micellar pseudophase and the kinetic parameters: catalytic rate constant k_cat, and the Michaelis constant K_M of the enzymatic reaction. The effect of the surfactant concentration on the kinetics parameters was determined at constant W₀=[H₂O]/[surfactant], and the variation of W₀ with surfactant constant concentration was investigated. The results show that the classical Michaelis–Menten mechanism is valid for α‐CT in all of the RMs systems studied and that the reaction takes place at both RM interfaces. Moreover, the catalytic efficiency values k_cat/K_M obtained in the RMs systems are higher than that reported in water. Furthermore, there is a remarkable increase in α‐CT efficiency in the cationic RMs in comparison with the anionic system, presumably due to the unique water properties found in these confined media. The results show that in cationic RMs the hydrogen‐bond donor capacity of water is enhanced due to its interaction with the cationic interface. Hence, entrapped water can be converted into “super‐water” for the enzymatic reaction studied in this work.     

Investigation on the basic hydrolysis of crystal violet in mixed reverse micelles formed with AOT and nonionic surfactants

 The kinetics and thermodynamics of the basic hydrolysis of crystal violet (CV) in mixed reverse micelles formed with anionic surfactant AOT and nonionic surfactants have been investigated. It was found that the mixed reverse micelles had inhibitory effects on CV hydrolysis compared with the normal aqueous solution, and the equilibrium constant K of the reaction in mixed reverse micellar systems is smaller than that in pure water. The influence of water content and surfactant composition in reverse micelles on the second-order rate constant k ₁ of the positive reaction, on the first-order rate constant k _-1 of the reverse reaction, as well as on the equilibrium constant K of the reaction has been studied, and the results obtained were interpreted in terms of the nature of surfactants and the properties of microenvironment where the reaction took place. Received: 24 October 1997 Accepted: 18 March 1998     

Complexation of Al(III) by 8-Hydroxyquinoline and Drastic Fluorescence Enhancement in Reverse Micelles

Complexation of Al^IIIby 8-hydroxyquinoline and fluorescence behavior of the quinolinate(s) were studied in reverse micellar systems at low water content, and compared to aqueous media. Two surfactants were used: one was cationic (CTAC: cetyltrimethylammonium chloride) and the other was anionic (AOT: sodium bis(2-ethylhexyl)sulfosuccinate). The results obtained in the CTAC/dichloromethane system (W= [H₂O]/[surfactant] = 0.9) showed that complexation occurred very likely in the oil phase and no micellar effect was observed. On the contrary, in the presence of AOT, specific micellar effects were observed due to the presence of the anionic polar heads: stabilization of the positively charged 1:1 and 1:2 chelates, at the expense of the neutral water-insoluble 1:3 chelate which is formed in aqueous solutions under similar conditions;drastic fluorescence enhancement factorsof 120 and 100 in AOT/heptane (W= 1.5) and AOT/dichloromethane (W= 1.6), respectively. Such factors have never been reported so far in either hydroorganic or direct micellar systems. In return, the length of time for the production of the complex(es) is increased because of the microheterogeneity of the medium and the small sizes of the water pools.     

Micropolarity of sodium bis(2-ethylhexyl) sulfosuccinate reverse micelles prepared in supercritical ethane and near-critical propane

The micropolarity of sodium bis(2-ethylhexyl) sulfosuccinate (AOT) reverse micelles prepared in supercritical ethane and near-critical propane has been determined in terms of a solvent polarity parameter, E _T(30) values, by using absorption probes, 1-ethyl-4-methoxycarbonyl pyridinium iodide and 2,6-diphenyl-4-(2,4,6-triphenylpyridinio)-phenolate as a functions of pressure and the molar ratio of water to AOT, W ₀, at a constant temperature of 310 K. The micropolarity comparable to that of ethanol was observed for reverse micelles containing water of W ₀ = 2. The micropolarity increased with the water content and became independent of pressure after the system changed to a one-phase reverse micelle solution. For a given W ₀ value, no difference in the micropolarity was noticed in the micelles prepared in ethane and propane. Phase behaviour investigations have revealed that complete dissolution of 50 mM AOT occurred at 20 MPa in supercritical ethane, while a much lower pressure of 1 MPa was required in near-critical propane. The amount of water solubilized in reverse micelles formed in supercritical ethane was relatively low, reaching a W ₀ value of 7 at 36 MPa. In contrast, the amount of water solubilized in near-critical propane reverse micelles was W ₀ = 11 at a much lower pressure of 6 MPa. A higher pressure was required to solubilize larger amount of water in reverse micelles prepared in both ethane and propane. Received: 9 October 1998 Accepted in revised form: 12 February 1999     

PNPP Cleavage Catalyzed by Schiff Base Mn(III) Complexes Containing Polyether Side Chains in CTAB Micellar Solutions

Two Schiff base Mn(III) complexes containing polyether side chain were synthesized and characterized. The catalytic hydrolysis of p‐nitrophenyl picolinate (PNPP) by the two complexes in the buffered CTAB micellar solution in the pH range of 6.60–8.20 was investigated kinetically in this study. The influences of acidity, temperature, and structure of complex on the catalytic cleavage of PNPP were also studied. The mechanism of PNPP hydrolysis catalyzed by Schiff base manganese(III) complexes in CTAB micellar solution was proposed. The relative kinetic and thermodynamic parameters were determined. Comparied with the pseudo‐first‐order rate constant (k ₀) of PNPP spontaneous hydrolysis in water, the pseudo‐first‐order rate constants (k _obsd) of PNPP catalytic hydrolysis are 1.93×10³ fold for MnL¹ ₂Cl and 1.06×10³ fold for MnL² ₂Cl in CTAB micellar solution at pH=7.00, T=25°C, and [S]=2.0×10^?4mol · dm^?3, respectively. Furthermore, comparing the k _obsd of PNPP catalytic hydrolysis by metallomicelles with that of PNPP hydrolysis catalyzed only by metal complexes or CTAB micelle at the above‐mentioned condition, metallomicelles of MnL₂(L=L¹, L²) Cl/CTAB exhibit notable catalytic activities for promoting PNPP hydrolysis, and MnL¹ ₂Cl/CTAB system is superior in promoting cleavage of PNPP relative to MnL² ₂Cl/CTAB system under the same experimental conditions. The results indicate that the rate of PNPP catalytic cleavage is influenced by the structures of the two complexes, the acidity of reaction systems, and the solubilization of PNPP in CTAB micelles.     

Kinetic Studies on of Veratryl the Lignin Peroxidase Catalyzed Oxidation Alcohol by H2O2 in AOT/Isooctane/ Toluene/Water Reverse Micelles

The steady state kinetics of the lignin peroxidase （LIP） catalyzed oxidation of veratryl alcohol （VA） by H2O2 in a sodium bis（2-ethylhexyl） sulfosuccinate （AOT）/isooctane/toluene/water reverse micellar medium was studied and a comparison with the corresponding aqueous medium was made to understand the effect of the reverse micellar medium on the catalytic mechanism and kinetic parameters. Results indicated that the model reaction in the AOT reverse micelle followed the ping-pong mechanism with true kcat, Km,VA and KmH2O2 being 59.6min^-1, 13.9 mmol· L^-1 and 94.8 μmol·L^-1, respectively; inhibition of high level of H2O2 on LiP followed the reversible competitive pattern with Ki being 0.140 mmol·L^-1. The reaction mechanism and inhibition pattern in the AOT reverse micellar medium were the same as those in bulk aqueous medium, but the kinetic parameters except KmH2O2 were greatly different in the two media. The kcat and Ki values in the reverse micelle were approximately 2 and 20 times smaller than the corresponding values in the aqueous solution, but the Michaelis constant of VA was approximately 100 times greater than that in the aqueous solution. The above mentioned differences in the kinetic parameters were caused by the microheterogeneity and the interface of the AOT reverse micelle, which resulted in the partitioning of VA and H2O2, and by the changes of the conformation of LiP and the reactivity of the substrates.     

Kinetics of the Oxidation of Phenylhydrazine by [Fe(CN)6]3− in Water‐in‐Oil Microemulsions

The oxidation reaction of phenyl hydrazine (Phh) by hexacyanoferrate ([Fe(CN)₆]^3?) has been studied in water‐in‐oil (w/o) microemulsion media. The kinetic profile of the reaction was investigated as a function of [Phh], droplet size, and droplet concentration. Comparison of the kinetic profiles of the reaction in microemulsion, water‐urea, and water‐AOT‐urea media indicates that the kinetic profile of the reaction in microemulsion shows a behavior similar to that of the reaction in water‐AOT‐urea medium at 4 M urea. An initial increase and then a decrease in k_obs is observed with increasing molar ratio, W_o(=[H₂O]/[AOT]) at constant [AOT] (=0.4 M), whereas k_obs decreases upon increasing the AOT concentration at constant molar ratio.     

Improvement in enzyme activity and stability by addition of low molecular weight polyethylene glycol to sodium bis(2-ethyl-L-hexyl)sulfosuccinate/isooctane reverse micellar system

The activity and stability of Chromobacterium viscosum lipase (glycerolester hydrolase, EC 3.1.1.3)-catalyzed olive oil hydrolysis in sodium bis (2-ethyl-1-hexyl)sulfosuccinate (AOT)/isooctane reverse micelles is increased appreciably when low molecular weight polyethylene glycol (PEG 400) is added to the reverse micelles. To understand the effect of PEG 400 on the phase behavior of the reverse micellar system, the phase diagram of AOT/PEG 400/water/isooctane system was studied. The influences of relevant parameters on the catalytic activity in AOT/PEG 400 reverse micelles were investigated and compared with the results in the simple AOT reverse micelles. In the presence of PEG 400, the linear decreasing trend of the lipase activity with AOT concentration, which is observed in the simple AOT reverse micelles, disappeared. Enzyme entrapped in AOT/PEG reverse micelles was very stable, retaining>75% of its initial activity after 60 d, whereas the half-life in simple AOT reverse micelles was 38 d. The kinetics parameter maximum velocity (V _max)exhibiting the temperature dependence and the activation energy obtained by Arrhenius plot was suppressed significantly by the addition of PEG 400.     

Effect of the Media on the Quantum Yield of Singlet Oxygen (O2(1Δg)) Production by 9H‐Fluoren‐9‐one: Microheterogeneous Systems

The quantum yield (Φ_Δ) of singlet oxygen (O₂(¹Δ_g) production by 9H‐fluoren‐9‐one (FLU) is very sensitive to the nature of the solvent (0.02 in a highly polar and protic solvent, such as MeOH, to 1.0 in apolar solvents). This high sensitivity has been used for probing the interaction of FLU with micellar media and microemulsions based on anionic (sodium dodecyl sulfate, SDS; bis‐(2‐ethylhexyl)sodium sulfosuccinate, AOT), cationic (cetyltrimethylammonium chloride, CTAC) and nonionic (Triton X‐100, TX) surfactants. Values of Φ_Δ of FLU vary in a wide range (0.05–1.0) in both microheterogeneous media and neat solvent, and provide information on the microenvironment of FLU, i.e., on its localization within organized media. In ionic and nonionic micellar media, as well as in four‐component microemulsions, FLU is, to various extents, exposed to solvation by the polar and protic components of the microheterogeneous systems (water and/or butan‐1‐ol) in the micellar interfacial region (Φ_Δ=0.05–0.30). In contrast, in AOT reverse micelles (consisting of AOT as surfactant, cyclohexane as hydrophobic component, and water), FLU is located in the hydrophobic continuous pseudophase, and is totally separated from the micellar water pools (Φ_Δ≈1.0).     

Specifics of the formation of J-aggregates of cyanine dyes in solutions of AOT/water/hexane reverse micelles

The behavior of a cyanine dye (3,3′-di-(gamma-sulfopropyl)-4,5,4′,5′-dibenzo-9-ethylthiacarbocyanine betaine pyridinium salt) was studied in AOT/water/hexane reverse micelles over a wide range of W at various concentrations of the dye, AOT, and reverse micelles. The processes occurring during the formation of the AOT/water/hexane micellar solution were studied in detail. It has been shown that, before the formation of the stable microemulsion, the dye aggregation processes occur by virtue of the interaction of the dye with the AOT anion. The amount of J-aggregates is proportional to the logarithm of the ratio of the amount of AOT molecules to the amount of dye molecules. The time behavior of J-aggregates after the formation of a micellar structure depends on the concentration of reverse micelles, thereby indicating an important role of intermicellar exchange.     

Kinetics for the Reaction between Potassium Ferricyanide and Cobalt Chloride in Aqueous Solution and Microemulsion

The kinetics for the reaction between potassium ferricyanide (K₃Fe(CN)₆) and cobalt chloride (CoCl₂) in aqueous solution and water/bis(2-ethylhexyl) sodium sulfosuccianate (AOT)/isooctane microemulsions were studied by three-wavelength spectrophotometry at 298.2 K. The second-order rate constants (k₂) were calculated from the time dependence of the concentration of reactant K₃Fe(CN)₆. The result showed that the reaction rates in water/AOT/isooctane microemulsions were slower than that in the aqueous solution, and k₂ decreased with molar ratio (ω) of water to AOT in microemulsions, which was interpreted by the transition state theory and confirmed that the reaction took place at the interfaces of the microemulsion water pools.     

Determination of the Water/AOT/Isooctane Reverse Micelles Size Parameters from Their Refractive Index Data

The present paper aims to study the proprieties of water confined in water/AOT/isooctane reverse micelles from their refractive indices at 298.5 K. The refractive indices of the microemulsions were investigated at increasing water concentration and at different micellar volume fractions, Φ _d. The refractive index of micellar water was deduced for a large water to AOT molar ratio, W ₀. The refractive index of interfacial water was also calculated. Then, the molar faction of interfacial water, α, was deduced for different W ₀ values and compared with those measured by NMR as reported in the literature. The results show qualitative agreement. Finally, the average aggregation number, n _agg, the area per surfactant headgroup, σ, as well as the interfacial thickness, d _i, of the reverse micelles were determined for the W ₀ values studied. The values of these structural parameters also show good qualitative agreement with NMR and small angle X-ray scattering data reported in the literature. Thus, the refractive index method can be a cheap and fast alternative for these two methods.     

Hydrolysis of PNPP Catalyzed by Cu (II), Ni (II) Schiff Base Complexes in CTAB Micellar Solution

The kinetics of hydrolysis of p‐nitrophenyl picolinate(PNPP) catalyzed by metallomicelles formed from Cu (II), Ni (II) Schiff base complexes (CuL, NiL) and CTAB micelle were investigated in the pH range of 6.0–9.0 at 30°C. For the Cu (II) Schiff base complex CuL, the apparent rate constants (k _obsd) of PNPP hydrolysis initially increased with the increasing pH of reaction media, then fell off. For the Ni (II) Schiff base complex NiL, the k _obsd always increased with the increasing pH. The kinetic and thermodynamic parameters were calculated. The hydrolysis rate of PNPP catalyzed by Cu (II) complex was much larger than that by Ni (II) complex in CTAB micellar solution. The catalytic mechanism of the PNPP hydrolysis was discussed in detail, and the possibly active specie for the catalytic hydrolysis of PNPP was the monohydroxo metal complex.     

Micellar Catalysis of Composite Reactions II. The Effect of CTAB Micelles on the Alkaline Fading of Malachite Green

The alkaline fading of malachite green, which is interpreted as parallel first order and second order reactions, has been studied in cetyltrimethylammonium bromide (CTAB) micellar solution at 25°C using spectrophotometry. A micellar catalytic model is proposed in this paper for constant concentration of hydroxideion. For this model, the first order and the second order rate constants in CTAB micellar phase,k_1m and k_2m have been obtained.

The experimental results indicate that the first order reaction of malachite green cation with water is catalysed by CTAB micelles while the second order reaction of malachite green cation with hydroxide ion is inhibited by CTAB micelles. The first order rate constant in CTAB micellar phase, k_1m , is 210 times of that in the bulk phase, but the second order rate constant in CTAB micellar phase, k_2m , is 0. 166 time of that in the bulk phase. The results are interpreted mostly in relation to the micellar micropolarity and electrostatic interaction. @Keywords: Micelle, Micellar catalysis, Parallel first order and seond order reactions, Malachite green     

Enthalpies of transfer of ubiquinone-0 from water to aqueous surfactant solutions at various temperatures

Mixing and dilution enthalpies of aqueous solutions of 2,3-dimethoxy-5-methyl-1,4-benzoquinone (UQ₀) and of N,N-dimethyl-dodecylamine-N-oxide (DDAO) were measured and used to calculate the enthalpies of transfer δH(W →W +S) of UQ₀ from water to DDAO aqueous solutions at 20‡, 25‡, 30‡, and 35‡C. From the dependence of δH(W →W +S) on surfactant concentration, the distribution constant between aqueous and micellar phases and the standard transfer enthalpy of UQ₀ from water to DDAO micelles were evaluated along with the standard transfer free energy and entropy. The approach used required knowledge of the CMC and micellization enthalpy at each temperature. Thus, the thermodynamics of micellization of DDAO was studied by means of dilution enthalpy measurements at the several temperatures.     

Small‐Angle X‐Ray Scattering and Near‐Infrared Vibrational Spectroscopy of Water Confined in Aerosol‐OT Reverse Micelles

The state of water confined in Aerosol‐OT–hydrocarbon–water reverse micelles with cyclohexane, n‐pentane, n‐octane, and n‐dodecane as apolar solvents is investigated by small‐angle X‐ray scattering and near‐infrared vibrational spectroscopy of the first overtone of the OH stretching mode of water. The experiments focus on water/AOT molecular ratios W₀=2–20, where water is strongly affected by the confinement and surface–water interactions. The pair‐distance distribution functions derived from the small‐angle scattering patterns allows a detailed characterization of the topology of these systems, and they indicate deviations from monodisperse, spherical water pools for some of these hydrocarbon systems. In contrast to a common assumption, the pool size does not scale linearly with W₀ in going from dry reverse micelles (W₀→0) to essentially bulk‐like water (W₀>20). The first overtone of the OH‐stretching vibration exhibits highly structured spectra, which reveal significant changes in the hydrogen bonding environment upon confinement. The spectra are rationalized by a core/shell model developed by Fayer and co‐workers. This model subdivides water into core water in the interior of the micelle and shell water close to the interface. Core water is modelled by the properties of bulk water, while the properties of shell water are taken to be those of water at W₀=2. The model allows the representation of the spectra at any hydration level as a linear combination of the spectra of core and shell water. Different approaches are critically reviewed and discussed as well.     

Catalytic effect of CTAB reverse micelles on the kinetics of dissociation of bis(2,4,6–tripyridyl-s-triazine) iron(II)

The kinetics of dissociation of bis(2,4,6–tripyridyl-s-triazine) iron(II), ([Fe(TPTZ)₂]²⁺) has been studied in CTAB/chloroform/hexane reverse micellar medium. In the absence of acid, the reaction is immeasurably slow and does not go to completion in conventional aqueous medium but is markedly accelerated and takes place with a rate constant equal to 55.3 × 10^?3 s^?1 and goes to completion in reverse micelles. The significant increase in rate is attributed to the special properties of the water pool in the reverse micelles like low dielectric constant, nucleophilic effect of Br^- ion, and favorable partitioning of TPTZ in the organic phase. The rate of the reaction decreases with increase in W (=[H₂O]/[CTAB]) at constant CTAB concentration and remains constant with increase in CTAB at fixed W. The results are compared with other closely related systems.     

Calorimetric investigation of reaction Ca(NO3)2 + Na3PO4 in water and microemulsions saturated with CO2

The reactions of Ca(NO₃)₂ + Na₃PO₄ in water and water/sodium bis(2‐ethylhexyl) sulfosuccinate (AOT)/hydrocarbon microemulsions saturated CO₂ with various molar ratios of water to surfactant R, oil phases, and surfactant concentrations were investigated by isothermal titration calorimetry. The product of the reaction was confirmed to be sodium‐and‐carbonate‐substituted hydroxyapatite (NaCO₃HAP) by Fourier transform infrared spectra (FTIR), energy dispersive spectrometry (EDS), and X‐ray diffraction (XRD). From calorimetric measurements, the molar enthalpies of solution of water in the AOT/n‐dodecane system, and the molar enthalpies, the rate constants, and the activation energies of the reactions were determined. It was found that the enthalpy of solution of water in AOT/n‐dodecane micells and the molar enthalpy of the reaction in the microemulsions increased with the decreases of R until R = 7; below that they kept almost constant. It may be attributed to the increase of the ratio of the bound water to the free water with the decrease of R until there was no free water when R < 7. However, the reaction rate constant k₁ was affected by the ionic strength of the medium and log k₁ showed a linear dependence on 1/R in the whole range of R we investigated. It was also observed that the rate constant and the enthalpy of the reaction remained almost unchanged when the surfactant concentration and the nature of oil phase varied. © 2011 Wiley Peiodicals, Inc. Int J Chem Kinet 43: 322–330, 2011     

Terahertz absorption spectroscopy of protein-containing reverse micellar solution

Terahertz time-domain spectroscopy has been carried out for AOT/isooctane reverse micellar solution with myoglobin at the water-to-surfactant molar ratios (w₀) of 0.2 and 4.4. The amplitude of the absorption spectrum increases with increasing the protein concentration at w₀ = 0.2, whereas it decreases at w₀ = 4.4. The molar extinction coefficients of the protein-filled reverse micelle, and the constituents, i.e., myoglobin, water, and AOT, have been derived by use of the structural parameters of the micellar solution. The experimental results are interpreted in terms of hydration onto the protein and surfactant in the reverse micelle.     

Interfacial Synthesis,Vol. Ill,Recent Advances,Charles E. Carraher,Jr. and Jack Preston,editors, Marcel Dekker,Inc., New York and Basel, 1982, $65.00

Hydrolyses of p‐nitrophenyl picolinate (PNPP) and p‐nitrophenyl acetate (PNPA) mediated by the micellar catalytic systems of two types of cationic surfactants [cetyltrimethylammonium bromide (CTAB), Gemini dimethylene‐1,2‐bis(cetyltrimethylammonium bromide) (16‐2‐16, 2Br^?)] were investigated spectrophotometrically in the pH range of 7.0–9.0 and 25°C. Also, the effects of several kinds of additives, such as ethanol, cyclodextrins (CDs), on the hydrolytic reactions of PNPP and PNPA were studied systematically. It is noteworthy that: (1) double chain Gemini surfactant micellar system enhanced the hydrolyses of carboxylic acid esters notably compared with single chain surfactant (CTAB) micellar solutions under the same reaction conditions; (2) the apparent rate constants (k _obsd) of PNPP and PNPA hydrolyses increased with the increasing in pH values of reaction media; (3) as additives, ethanol has effect on both PNPP and PNPA hydrolyses, and moreover, the k _obsd for hydrolyses decreased with the increasing contents of ethanol (≤5%) at 25°C and pH 9.00; (4) the presence of CDs [α‐cyclodextrin (α‐CD), β‐cyclodextrin (β‐CD), γ‐cyclodextrin (γ‐CD)], as additives, showed different effects on PNPP and PNPA hydrolyses in different reaction systems.