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Hollow Colloidal Particles Obtained by Nano-Extrusion in the Presence of Phospholipids David J. Lestage and Marek W. Urban* School of Polymers and High Performance Materials, Shelby F. Thames Polymer Science Research Center, The University of Southern Mississippi, Hattiesburg, Mississippi 39406 Received December 7, 2004. In Final Form: March 21, 2005 Using nano- and microsize extrusion, a simple synthetic procedure of preparing hollow monodispersed colloidal particles dispersed in an aqueous phase was developed. Hydrophobic styrene monomer containing 2-hydroxy-2-methyl propiophenone photoinitiator was forced into desired diameter membrane channels and stabilized by the hydrophobic regions of a liposome obtained from 1,2-dilauroyl-phosphocholine phospholipid in an aqueous phase. Such moieties exposed to 254-nm UV radiation polymerize monomers in the hydrophobic zone of the liposome, thus resulting in reinforced hollow vesicles. The size of such particles is controlled by the size of the membrane channels in the extruder and may vary from a few nanometers to micrometers, thus allowing the synthesis of monodisperse hollow colloidal spheres.
Although colloidal particles have been of interest for a number of years and utilized in numerous studies, it has not been realized until recently that their synthesis may be facilitated by amphiphilic bioactive molecules, such as phospholipids.1 Phospholipids, in addition to serving as dispersion stabilizing agents, may be deliberately mobilized to designated areas within polymer matrixes and subsequently aligned depending on concentrations of ionic environments.1 As a result of their structural features, phospholipids are also capable of forming bilayered liposomes, but their inherent weak structural features resulting from hydrophobic interactions require reinforcement.2 Attempts to reinforce spherical structures made from bilayers of phospholipid molecules typically involve polymerization of hydrophobic monomers such as styrenes, methacrylates, isodecyl acrylate, and oleic acid solubilized within the vesicle bilayer which may also contain fluorinated species.3 These efforts relied on thermally or photochemically induced polymerization initiated by either hydrophilic or lipophilic initiators, and some evidence4,5 on the formation of polymer capsules was reported. In a typical experiment, the phospholipid solution was sonicated to obtain desirable liposome sizes, monomer and initiator species were added, and if monomer migrated into the hydrophobic zone and polymerized, polymeric spheres were obtained. However, neither the size nor the particle size distributions are controllable, and such reactions would typically require several hours. An inherent weakness of the hydrophobic loci of polymerization often resulted in parachute-like or semispherical morphologies instead of a spherical shape, because the only controlling variables in this process are structural features of phospholipids, which are also heavily dependent on environmental conditions. The importance of structurally sound liposomes comes from the fact that, under certain conditions, self-assembled bilayered liposomes may serve as lipophilic drug carriers to target sites in aqueous media, 6-10 and when derived from physiological * To whom all correspondence should be addressed. (1) Yacoub, A.; Urban, M. W. Biomacromolecules 2003, 4, 52-56. (2) Murtagh, J.; Thomas, J. K. Faraday Discuss. Chem. Soc. 1986, 81, 127-136. (3) Krafft, M. P.; Schieldknecht, L.; Marie, P.; Giulieri, F.; Schmutz, M.; Poulain, N.; Nakache, E. Langmuir 2001, 17, 2872-2877. (4) Hotz, J.; Meier, W. Langmuir 1998, 14, 1031-1036. (5) Jung, M.; Ouden, I.; Montaoya-Goni, A.; Huber, D. H. W.; Frederik, P. M.; van Herk, A. M.; German, A. L. Langmuir 2000, 16, 4185-4195.
Figure 1. Nano-extruder with desired size membrane channels.
structures, are expected to have increased biocompatibility due to chemical and compositional similarities to their biological counterparts.11 The liposome architecture is inherently fragile, which limits encapsulation properties, thus affecting loads and shelf life stability. To overcome these drawbacks, this letter illustrates a simple synthetic procedure that allows controllable synthesis of hollow colloidal particles with monodispersed hollow particle sizes. We utilized a nano-extruder, and Figure 1 illustrates its schematic diagram. As shown, it consists of two reservoirs separated by a membrane. The latter contains desirable nano- or microdiameter channels through which a liquid between the two reservoirs can be forced back and forth. When hydrophobic monomers at low concentration levels and phospholipid molecules are present in an aqueous environment and passed through the membrane, the size of the liposome consisting of a phospholipid bilayer will in turn be determined by the size of the membrane channels through which the fluid is passed. It is important to keep monomer concentrations low enough to occupy only the hydrophobic zone of the liposome and high enough to be able to polymerize within that zone, and the optimum conditions may vary for each monomer. By pushing such fluid back and forth, the desired size of bilayered liposomes can be obtained with monomer partitioned in the hydrophobic layer. Although (6) Cornelus, C.; Giulieri, F.; Krafft, M.-P.; Riess, J. G. Colloids Surf., A 1993, 70, 233-238. (7) Carpentier, Y. A.; Simoens, C.; Siderova, V.; El Nakadi, I.; Vanweyenberg, V.; Eggerickx, D.; Deckelbaum, R. Nutrition 1997, 13. (8) Freeman, F. J.; Chapman, D. Polymerizable Liposomes: Applications in Biology and Medicine; 1988. (9) Regen, S. L. Polymerized Liposomes as Drug Carriers; 1991. (10) Gregoriadis, G. Liposomes as Drug Carriers: Recent Trends and Progress; John Wiley and Sons: New York, 1988. (11) Westesen, K. Colloid Polym. Sci. 2000, 278, 608-618.
10.1021/la046988i CCC: $30.25 © 2005 American Chemical Society Published on Web 04/12/2005
Letters
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Figure 2. Freeze fracture TEM of a 5-µm hollow p-Sty particle.
the initial droplets are large, to ensure that smaller droplets are not passed through, we stock multiple (typically three) membranes to impart the flow of smaller vesicles. Subsequent monomer polymerization will lead to the formation of hollow particles. Figure 2 illustrates an example of a freeze fracture transmission electron micrograph (TEM) of hollow p-sytrene (p-Sty) spheres obtained using this method. To illustrate the monodispersity of these particles, Figure 3 shows standard TEMs at different magnifications. Using a nano-extruder with different nano-channel membranes allows the synthesis of various sizes of these hollow particles. To obtain hollow p-Sty particles such as shown in Figure 2, 1,2-dilauroyl-phosphocholine phospholipid was solubilized in chloroform (CHCl3) and dried in a rotary evaporator to form a thin lipid film on the wall of the flask. Such films were rehydrated with DDI H2O and vortexed gently to form a lipid solution. Photoinitiator (2-hydroxy-2-methyl propiophenone; Darocur 1173, Ciba Co.) was dissolved in Sty monomer prior to addition to the lipid solution, which was then passed through polycarbonate membranes of the extruder to form monodisperse unilamellar vesicles 5 µm in diameter. The solution was allowed to equilibrate for 1 h under gentle stirring and N2 purge followed by polymerization using 20-V, 254-nm UV radiation.
Figure 3. TEM of a larger population of hollow colloidal particles.
In summary, this one-step procedure allows synthesis of colloidal particles with controlled monodispersed sizes, and a combination of monomers and phospholipids under similarly engineered conditions may provide means for engineering other shapes ranging from nano- to microscales. Because liposomes may be utilized in polymerization of colloidal dispersions, they will be capable of selfassembling during film formation processes, thus providing an outstanding opportunity for the development of new technologies ranging from drug delivery systems to new advances in biotechnologies with stimuli-responsive behaviors which may be tailored for specific applications. Acknowledgment. This work was supported primarily by the MRSEC Program of the National Science Foundation under Award No. DMR 0213883. The authors are also thankful to the National Science Foundation Major Research Instrumentation Program under the Award No. DMR 0315637 for partial financial support of these studies. LA046988I
