Aggregation of Vitamin C derivatives in water solution

 

The basic molecular architecture of amphiphiles is always based on the simultaneous presence in the same molecule of two or more groups of atoms that possess different affinities for a solvent, with which they establish different interactions, and that is generally defined as a "selective solvent". Usually the solvent is water, and then we distinguish a hydrophilic part (the "polar headgroups"), linked to a hydrophobic block made up of one, two, or more hydrocarbon chains (see Fig. 5).

The polar headgroups can be either neutral, cationic, or anionic residues. Typical neutral surfactants contain functionalities such as -OH, -NR2 (R=alkyl or H), esters, ethers, amides, and so forth. Cationic amphiphiles are for example alkyl-ammonium salts (such as dioctadecyldimethylammonium chloride, usually called DODAC), while anionic tensides can be carboxylates, sulfates, or phosphates. On the other hand, phosphatidylcholines are typical zwitterionic amphiphiles.

An interesting class of molecules are the so-called bolaamphiphiles, where two headgroups are bound by one or two chains (see Fig. 5). In this case monolayered structures are formed. These surfactants are typically present in the membranes of Archaebacteria, primordial microorganisms that live in extreme environments, such as volcanoes or under the oceans, and experience very drastic environmental conditions (pH<2, T>70 C, and high pressures).
Amphiphiles promptly form supramolecular aggregates in water, because of the "hydrophobic effect", that is the formation of a hydrophobic central core and of an external hydrophilic shell. This process reduces the hydrocarbon-water repulsion and then minimizes the total energy of the aggregate.

Depending on the chemical structure of the amphiphile, temperature, ionic strenght of the solution, nature and composition of the solvent, different kinds of aggregates can be obtained, with peculiar properties and structures. As Fig. 5 shows, spreading monolayers, adsorption films, micelles, vesicles (or liposomes), microemulsions, and Langmuir-Blodgett multilayers are different supramolecular structures, but they all originate from the same self-assembly of surfactants in the presence of a selective solvent.



Fig. 5 - Schematic structure of amphiphiles and of their self-assembled supramolecular aggregates.

Although the driving force that leads to the formation of these structures is always the "hydrophobic effect", however each one of the supramolecular assemblies possesses peculiar properties that can be studied with different techniques. Just as an example, monomolecular films are basically studied by measuring the "spreading isotherms", that is the plot that one can obtain by measuring the surface pressure as a function of the surface area; micellar solutions can be regarded as dispersions of small particles (usually spheres or ellipsoids) that can be characterized by surface tension measurements, refractive index, light-scattering, neutron-scattering, viscosity, EPR, NMR, and so forth. Emulsions and microemulsions are of great interest when an intimate mixture of lipophilic and hydrophilic components is desidered, such as in drugs, cosmetics, food processing, paper and textile manufacturing, oil recovery, inks and paintings, and so forth, and for this reason the study of their stability and phase behavior as a function of temperature and composition, i.e. the "phase diagram", is strongly needed.

Fig. 6 shows the spreading isotherms obtained from 6-O-stearoyl ascorbic acid at pH=6, vitamin K1, and vitamin D3 at 25C. As expected, 6-O-stearoyl-ascorbic acid produces more condensed films, because of its long aliphatic side chain, whilst the other two components give monolayers that show a more expanded behavior. The spreading behavior of ascorbyl-palmitate has been studied by Balthasar and Cadenhead (4).


Fig. 6 - Spreading isotherms (surface pressure vs. molecular area) of ascorbyl-stearate, vitamin K1, and vitamin D3.

The formation of self-assembled aggregates in water strongly depends on several factors: surfactant concentration, temperature, ionic strenght, presence of other molecules. When a surfactant is progressively added to water, it will dissolve in the bulk and form an adsorption film at the air/water interface; adding more surfactant will result in the formation of the first aggregates when the concentration equals the "critical micellar concentration", CMC (see Fig. 7).



Fig. 7 - Formation of supramolecular aggregates in a multi-component equilibrium with the surfactant's monomers. The blue spots represent the polar headgroups, and the brown lines indicates the hydrophobic chains.

For c<CMC, the single monomers float in the bulk phase, and begin to produce the so-called adsorption film at the air/water interface, where they orient the hydrophobic chains out in the air, while the polar headgroups are anchored in the aqueous phase. For c>CMC, the monomers' concentration remains equal to the CMC, but the number of aggregates increases (see Fig. 7). In spite of this "static" model, it should be remembered that a micellar system is instead very dynamic, in fact the monomers diffuse all the time from one aggregate to the other, spending some time in the bulk solution as single molecules. The CMC can be easily determined as the crossing point of the two straight lines obtained from the least square fitting of the surface tension vs log c data, as Fig. 8 shows in the case of 6-O-stearoyl-ascorbate water solutions at T=30C and pH=6. CMC can also be measured by other techniques, such as light-scattering, viscosity, conductivity, density, and is generally obtained as the point where the macroscopic parameter suddenly changes, due to the formation of micelles, ultimately of an oil core surrounded by a hydrophilic shell.



Fig. 8 - Calculation of the CMC value from the surface tension vs. concentration plot. The red spots are the experimental data, the black lines are the fitting linear curves. CMC is determined as the intersection point of the two lines.

From this plot it is easy to calculate the area per polar headgroup, A; for a nonionic surfactant the following Gibbs' equation holds:

where R=8.31107 erg/molK, T is the absolute temperature, and NA is the Avogadro number. For the previous plot, A was calculated as 47 2/molecule. When the surfactant is charged, A must be multiplied by a factor 2.

The CMC value is affected by several factors, first of all by the chamical structure of the tenside (hydrophilic/hydrophobic balance, charges, branching groups, unsaturated bonds), by the temperature and by the presence of other molecules and/or ionic species.

The shape and the size of the supramolecular structures - that is the number of monomers per aggregate ("aggregation number", g) - depend on the chemical nature and geometry of the surfactant, on the monomers' concentration and on the temperature.

More recently the formation of large interface aggregates in non-aqueous solvents and with different types of chains has been reported in several papers, provided that the selective solvent plays different interactions with the two incompatible building blocks of the solute. As an example, a semifluorinated n-alkane, bearing a hydrogenated segment linked to a fluorinated block, can form aggregates in a fluorocarbon such as perfluorooctane, because of the well-known mutual immiscibility of hydrocarbons and fluorocarbons.

The self-assembling of surfactants in water and related properties are well described in several books (5).

As already mentioned in the previous section, a large number of lipophilic derivatives of vitamin C can be synthesized, where the polar head group is the ascorbic acid moiety, linked to one or two hydrophobic chains. Assembled in such supramolecular structures, vitamin C derivatives protect degradable materials (particularly unsaturated fats or vitamins): in fact the lipophilic molecules are segregated and protected in the micellar hydrophobic core, whilst the ascorbic acid polar head groups face the water phase and perform their radical-scavenger activity. As a matter of fact, when the vitamin C-based surfactants aggregates in micellar structures, the active ascorbic ring is even more exposed to the facing aqueous medium.
Long chain derivatives of ascorbic acid readily produce monomolecular films at the air/water interface (6-8), and give stable mixed monolayers with some vitamins that possess an amphiphilic structure as well. This feature is particularly important in order to determine the mutual miscibility of ascorbic acid derivatives and some relevant natural compounds with a perspective of producing stabilized systems where the ascorbyl-derivatives protect the other components against oxidation.
The self-assembly properties of 6-O-octanoyl ascorbic acid in water have been recently studied with viscosity, light-scattering and small-angle neutron-scattering measurements. The data show that small, monodisperse, nearly spherical aggregates are formed, with a hydrodynamic radius of about 25 . The oxygen consumption and the reducing activity of this compound have been tested and show that it is at least as powerful as vitamin C (9).

The structure and properties of another derivative, namely the 5,6-octylidene-ascorbic acid, are currently being studied in aqueous beta-octyl-glucoside micelles.

6-O-ascorbic acid esters, such as palmitate, are almost insoluble in water at room temperature, however their solubility increases at higher temperature, above which a clear solution is formed. This is due to the formation of micelles from the saturated solution of monomers, and therefore is referred to as the critical micellization temperature, CMT (1,10). Upon cooling, the micellar solution solidifies into an opaque curd, that contains more than 80% of water, and is a semicrystalline mesophase, usually called "coagel".

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