Biomacromolecules 2003, 4, 360-365
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Self-Organization of Amphiphilic Polymer in Vesicle Bilayers Composed of Surfactant Mixtures Sang-Yoon Kang,† Baek-Seok Seong,‡ Young Soo Han,‡ and Hee-Tae Jung*,† Department of Chemical and Biomolecular Engineering, Korea Advanced Institute of Science and Technology, 373-1 Guseong-dong, Yuseong-gu, Daejeon 305-701, Korea, and Neutron Physics Department, Hanaro Center, Korea Atomic Energy Research Institute, P.O. Box 105, Yuseong-gu, Daejeon 305-600, Korea Received October 19, 2002
Cryogenic transmission electron microscopy (cryo-TEM) and small angle neutron scattering (SANS) are used to investigate the association of amphiphilic polymers consisting of a double-chain hydrophobic tail attached onto poly(ethylene glycol) (PEG) polymer chains into two different systems of equilibrium vesicles. For cetyltrimethylammonium bromide (CTAB)/sodium perfluorohexanoate (FC5) vesicle bilayers, the size distribution of the vesicles slightly becomes narrow in the presence of the polymers, suggesting that the wedge-shaped polymers increase the spontaneous curvature of the vesicles. In contrast, the confinement of polymer molecules inside the CTAB/sodium perfluorooctanoate (FC7) vesicles that are stabilized by spontaneous curvature causes an abrupt decrease in the bilayer rigidity. By an analysis of vesicle size distribution, it is found that the membrane elasticity of CTAB/FC7 vesicles is varied considerably from 6kBT to 0.3kBT, implying the transition of stabilization mechanism from spontaneous curvature to thermal fluctuation in the presence of polymer. The polymer incorporation mechanism into the bilayers is understood, in the comparison of the vesicle radius and size distribution before and after adding polymer, as that the polymer is anchored into the vesicle bilayer owing to hydrophobic property after the adsorption on the surface of the bilayer. Introduction Mixtures of anionic and cationic surfactants self-assemble into various microstructures such as micelle, vesicle, lamellar, columnar, and cubic mesophases, depending on the strength of intra- and intermolecular interactions, the relative fractions of different groups within the molecules, and the shape of the molecules.1-4 Among the mesophases, there has been much recent interest in studying the microstructure and properties of vesicle phase because of their possible applications in exceptionally selective membranes, drug-delivery, microreactors for production of colloidal particles, and cosmetic applications.5 In such applications, the control of vesicle stability is crucial to avoid the aggregation or the fusion of bilayers. It is well-known that spontaneous equilibrium vesicles can be formed from anion-cation surfactant pairs acting as double-tailed zwitterionic surfactants.6 Such pseudo-double-tailed surfactants are expected to form bilayers. However, the stability of the spontaneous vesicles against aggregation and the existence of highly swollen lamellar phases require the presence of additional repulsive interactions in addition to conventional double-layer electrostatics.7-9 To date many investigators have attempted to figure out the unknown mechanism of stabilizing vesicles. Several theoretical works have shown that the large swellings of the † ‡
Korea Advanced Institute of Science and Technology. Korea Atomic Energy Research Institute.
lamellar phases and the spontaneous vesicles may be stabilized by Helfrich-type undulations indicative of low bending modulus bilayers.10-12 The previous experimental results have shown that the stability of equilibrium unilamellar vesicles is determined by thermal membrane fluctuations or spontaneous curvature.13 In other words, the spontaneous vesicles are stable either when repulsive potential energy due to bilayer fluctuation dominates the van der Waals attraction between bilayers or when the stiffness of bilayers are enough to avoid addition of second bilayer to the vesicle. An interesting approach is lately directed toward vesicle membranes consisting of surfactants, water, and polymers so that it may provide more understanding of the protein interactions with liposome in cell membrane, as well as may help the development of highly stable drug delivery materials.14-18 It is predicted that the addition of a small amount of polymer alters the microstructure and the elastic properties of vesicle bilayer. In the previous theoretical works18,19 the mechanism of polymer penetration into vesicle is proposed such that polymer in a vesicle phase creates a new effective polymer-mediated phase after the incursion of polymer depending on the architecture of the polymer and on the polymer/bilayer interactions. However, it is not well understood how the polymer is incorporated into the bilayers and how the anchoring of polymer affects the vesicle structure.
10.1021/bm025704a CCC: $25.00 © 2003 American Chemical Society Published on Web 12/21/2002
Self-Organization of Amphiphilic Polymer
Biomacromolecules, Vol. 4, No. 2, 2003 361
Thus, experimentally we examine the morphology obtained from cryogenic transmission electron microscopy (cryoTEM) and small angle neutron scattering (SANS) to investigate the difference between before and after the addition of polymer to vesicles by estimating the radius distribution and the bilayer elastic properties of the vesicles. This analysis gives useful information to understand microstructure variation, as well as to identify how vesicles are stabilized. Here polymer-surfactant macromolecules are anchored into two different systems of equilibrium vesicles. We observe that the polymer molecules are fully incorporated into those surfactant bilayers of unilamellar vesicle phase along with the resultant variation of microstructures depending on the bending modulus of surfactant mixture. Experimental Section Materials and Sample Preparations. Spontaneous vesicle phase was obtained from the mixtures of cationic and anionic single-tailed surfactants in aqueous solutions. Cetyltrimethylammonium bromide (CTAB, Aldrich), as a cationic surfactant, was recrystallized three times from a 50/50 mixture of acetone and ethanol. For anionic surfactants, sodium perfluorooctanoate acid (FC7) and sodium perfluorohexanoate acid (FC5) from TCI America were neutralized with sodium hydroxide to give the sodium salts. Two different systems of spontaneous unilamellar vesicles (ULV) were used in this study. One is made up of CTAB and FC5 in deionized water, and the other is a mixture of CTAB and FC7. The subscripts 7 and 5 represent the number of carbons in the fluorocarbon chain. The stock solutions allowed a week for complete equilibration at room temperature. The two different surfactant mixtures self-assemble into a variety of phases in water. Unilamellar vesicles form only in the water-rich corner of the phase diagrams, and the increase of total concentration leads to the formation of the liquid crystalline lamellar LR and multilamellar vesicle (MLV) phase over the wide range of composition.20 The ULV phase exists on the FC7 (and FC5)-rich side at concentrations between about 2 and 4 wt % surfactant and for mixing ratios greater than 70% FC7 (and FC5). The cationic and anionic surfactants ratio was fixed to isolate the effect of amphiphilic polymer on the structure and bending rigidity of the spontaneous vesicles. The polymer-surfactant macromolecule used here is composed of a dimyristoylphosphatidylethanolamine (DMPE) lipid at the one end of a water-soluble poly(ethylene glycol). The PEG-coated liposomes (DMPE), which we refer to as PEG-DMPE here,21,22 were purchased from Avanti Polar Lipids (Alabaster, AL) and used as received. PEG-DMPE of two different molecular weights were used in this experiment containing PEG with the number-average molecular weights, Mn, of ∼2000 and ∼5000 obtained by gel permeation chromatography (GPC) in chloroform. The polydispersity (Mw/Mn) of the samples are 1.13 and 1.11, respectively.22 The polymers were added in the catanionic vesicle solutions to total surfactant weight ratio. The molecular structure and schematic of vesicle bilayer composed of cationic-anionic surfactants and PEG-DMPE are shown in Figure 1.
Figure 1. Schematic representation of unilamellar vesicle membranes composed of catanionic surfactant, PEG-DMPE (poly(ethylene glycol)-dimyristoylphosphatidylcholine). Mn ≈ 2000 for N ) 45, Mn ≈ 5000 for N ) 113.
cryo-TEM. A thin layer (
