Microemulsions, aqueous organic solvents

Enzymes are suspended in hydrated microemulsion surrounded by a monolayer of surfactant molecules dispersed in an apolar solvent [53-60,135] [Fig. 1(b)]. Micelles ( 2 nm sphere) are formed when lyophilized or aqueous preparation of enzymes are introduced with stirring or shaking into a solution of synthetic or natural surfactant in an organic solvent. 

The homocoupling of aryl halide to diaryl compounds, known as Ull-mann coupling, is a synthetically useful reaction and has wide applications in material research. Such couplings have been studied in aqueous conditions. In 1970, arylsulfinic acids were coupled with Pd(II) in aqueous solvents to biaryls (Eq. 6.25).53 However, the reaction required the use of a stoichiometric amount of palladium. In the presence of hydrogen gas, aryl halides homocoupled to give biaryl compounds in moderate yields (30-50%) in an aqueous/organic microemulsion (Eq. 6.26).54... 

Usually, activities of enzymes (hydrogenases included) are investigated in solutions with water as the solvent. However, enhancement of enzyme activity is sometimes described for non-aqueous or water-limiting surroundings, particular for hydrophobic (or oily) substrates. Ternary phase systems such as water-in-oil microemulsions are useful tools for investigations in this field. Microemulsions are prepared by dispersion of small amounts of water and surfactant in organic solvents. In these systems, small droplets of water (l-50nm in diameter) are surrounded by a monolayer of surfactant molecules (Fig. 9.15). The water pool inside the so-called reverse micelle represents a combination of properties of aqueous and non-aqueous environments. Enzymes entrapped inside reverse micelles depend in their catalytic activity on the size of the micelle, i.e. the water content of the system (at constant surfactant concentrations). 

The procedure chosen for the preparation of lipid complexes of AmB was nanoprecipitation. This procedure has been developed in our laboratory for a number of years and can be applied to the formulation of a number of different colloidal systems liposomes, microemulsions, polymeric nanoparticles (nanospheres and nanocapsules), complexes, and pure drug particles (14-16). Briefly, the substances of interest are dissolved in a solvent A and this solution is poured into a nonsolvent B of the substance that is miscible with the solvent A. As the solvent diffuses, the dissolved material is stranded as small particles, typically 100 to 400 nm in diameter. The solvent is usually an alcohol, acetone, or tetrahydrofuran and the nonsolvent A is usually water or aqueous buffer, with or without a hydrophilic surfactant to improve colloid stability after formation. Solvent A can be removed by evaporation under vacuum, which can also be used to concentrate the suspension. The concentration of the substance of interest in the organic solvent and the proportions of the two solvents are the main parameters influencing the final size of the particles. For liposomes, this method is similar to the ethanol injection technique proposed by Batzii and Korn in 1973 (17), which is however limited to 40 mM of lipids in ethanol and 10% of ethanol in final aqueous suspension. 

As is often the case, we have become involved in microemulsions somwehat by accident. In the last five years or so we have been making systematic studies of the thermodynamic properties of aqueous organic mixtures and of electrolytes in these mixed solvents. Of particular interest were our heat capacity measurements. With a differential flow microcalorimeter it is possible to... 

Semiconductor nanoclusters trapped in AOT w/o microemulsions are reported to exhibit longer excited state lifetimes (about 10-100 ns) than those in aqueous solution or in monophasic organic solvents [213]. Clearly the surfactant-nanoparticle interaction is very important not only in restricting growth but also in extending the hfetimes of the excited states. Tata et al. [214] have shown that the removal of water from the micelles leads to a strong increase in fluorescence intensity, and the addition of specific quencher, 4-hydroxythiophenol, leads to variations in quenching efficiencies. 

Water in oil microemulsions with reverse micelles provide an interesting alternative to normal organic solvents in enzyme catalysis with hydrophobic substrates. Reverse micelles are useful microreactors because they can host proteins like enzymes. Catalytic reactions with water insoluble substrates can occur at the large internal water-oil interface inside the microemulsion. The activity and stability of biomolecules can be controlled, mainly by the concentration of water in these media. With the exact knowledge of the phase behaviom" and the corresponding activity of enzymes the application of these media can lead to favomable effects compared to aqueous systems, like hyperactivity or increased stability of the enzymes. 

Biotin-BMCC is insoluble in water and must be dissolved in an organic solvent prior to addition to an aqueous reaction mixture. Preparing a concentrated stock solution in DMF or DMSO allows transfer of a small aliquot to a buffer reaction. The upper limit of biotin-BMCC solubility in DMSO is approximately 33 mM or 17 mg/ml. In DMF, it is only soluble to a level of about 7 mM (4 mg/ml). On addition of an organic solution of the reagent to an aqueous environment (do not exceed 10% organic solvent in the aqueous medium to prevent protein precipitation), biotin-BMCC may form a microemulsion. This is normal and during the course of the reaction the remainder of the compound will be driven into solution as it couples or hydrolyzes. 

Activity and stability are often comparable to values in aqueous media. Many substrates which cannot be made to react in water or in pure organic solvents such as hexane owing to lack of solubility can be brought to reaction in microemulsions. Whereas enzyme structure and mechanism do not seem to change upon transition from water to the microemulsion phase (Bommarius, 1995), partitioning effects often are very important. Besides an enhanced or diminished concentration of substrates in the vidnity of microemulsion droplets and thus of enzyme molecules, the effective pH values in the water pool of the droplets can be shifted in the presence of charged surfactants. Frequently, observed acceleration or deceleration effects on enzyme reactions can be explained with such partitioning effects (Jobe, 1989). 

The solvent emulsification/evaporation method involves lipid precipitation in O/W emulsions. Solid lipids are dissolved in a water-immiscible organic solvent (e.g., cyclohexane) followed by emulsification in an aqueous medium. Upon evaporation of the solvent, the nanoparticle dispersion is formed due to lipid precipitation. Residue of organic solvents is the major problem of this method [94], However, the microemulsion and solvent emulsification/evaporation methods can be performed conveniently in the laboratory without specific apparatuses. 

A great variety of chemical reactions can be advantageously carried out in microemulsions [860-862]. In one of the first papers in this field, Menger et al. described the imidazole-catalyzed hydrolysis of 4-nitrophenyl acetate in water/octane microemulsions with AOT as an anionic surfactant [=sodium bis(2-ethyl-l-hexyl)-sulfosuccinate] [864]. The solubilized water, containing the imidazole eatalyst, is confined in spherical pools encased by surfactant molecules, which have only their anionic head groups (-SOb ) immersed in the aqueous droplets. When the ester, dissolved in water-insoluble organic solvents, is added to this water/octane/AOT/imidazole system, it readily undergoes the catalysed hydrolysis under mild reaction conditions (25 °C). 

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