Electrooptic Behavior and Structure of Novel Polymer−Vesicle Hybrids

Langmuir 1999, 15, 8849-8855

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Electrooptic Behavior and Structure of Novel Polymer-Vesicle Hybrids D. H. W. Hubert,*,† P. A. Cirkel,‡ M. Jung,† G. J. M. Koper,‡ J. Meuldijk,§ and A. L. German† Laboratory of Polymer Chemistry, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven, The Netherlands, Leiden Institute of Chemistry, Leiden University, Gorlaeus Laboratories, P.O. Box 9502, 2300 RA Leiden, The Netherlands, and Process Development Group, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven, The Netherlands Received June 8, 1999. In Final Form: August 11, 1999 It has been previously shown that polymerization of styrene in DODAB vesicles gives rise to the formation of unique parachute-like structures in which a polymer bead is confined to the bilayer at one pole of the vesicle. In this paper, quantitative data on the size and the shape of these hybrid particles and of the bare DODAB vesicles are obtained by combination of cryo-TEM, transient electrooptic birefringence (Kerr effect), and dynamic light scattering. Also, this study qualitatively assesses the influence of the confined polymer bead on the electrooptic behavior. It turns out that the presence of the bead gives rise to a special response in the transient electrooptic birefringence, typical of the existence of permanent dipoles. Unlike other vesicles, the DODAB vesicles and the polymer/vesicle hybrids do not display a deformation step in their electrooptic response. As such, these particles act as rigid particles under the action of an electric field. The thermal behavior of the DODAB templates and parachutes is investigated and explained in terms of intrinsic vesicle properties.

Introduction Controlled synthesis of organized matter is the subject of many publications in the field of polymer science1 and materials science.2 Chemists and materials scientists are engaged in developing techniques to control the morphologies and structural organization of materials from the microscale to the macroscale.3 Many approaches are based on performing reactions (organic4,5 or inorganic6,7) in organized media or in the presence of superstructuring agents. To this end, lyotropic ordered phases are frequently exploited. These agents or media are intended to act as templates, to imprint their morphology and structure on the synthesized matter. Polymerization of hydrophobic monomers inside the bilayer of a surfactant vesicle is an example of this approach.8-10 In our previous study on * Corresponding author. † Laboratory of Polymer Chemistry, Eindhoven University of Technology. ‡ Leiden Institute of Chemistry, Leiden University. § Process Development Group, Eindhoven University of Technology. (1) Paleos, C. M. Polymerization in Organised Media; Gordon and Breach Science Publishers: Philadelphia, 1992. (2) Mann, S.; Burkett, S. L.; Davis, S. A.; Fowler, C. E.; Mendelson, N. H.; Sims, S. D.; Walsh, D.; Whilton, N. T. Chem. Mater. 1997, 9, 2300-2310. (3) Schu¨th, F. Curr. Opin. Colloid Interface Sci. 1998, 3, 174-180. (4) Antonietti, M.; Hentze, H.-P. Colloid Polym. Sci. 1996, 274, 696702. (5) Chew, C. H.; Li, T. D.; Gan, L. H.; Quek, C. H.; Gan, L. M. Langmuir 1998, 14, 6068-6076. (6) Kresge, C. T.; Leonowicz, M. E.; Roth, W. J.; Vartuli, J. C.; Beck, J. S. Nature 1992, 359, 710-712. (7) Liu, J.; Kim, A. Y.; Wang, L. Q.; Palmer, B. J.; Chen, Y. L.; Bruisma, P.; Bunker, B. C.; Exarhos, G. J.; Graff, G. L.; Rieke, P. C.; Fryxell, G. E.; Vriden, J. W.; Tarasevich, B. J.; Chick, L. A. Adv. Colloid Interface Sci. 1996, 69, 131-180. (8) Murtagh, J.; Thomas, J. K. Faraday Discuss. Chem. Soc. 1986, 81, 127-136. (9) Morgan, J. D.; Johnson, C. A.; Kaler, E. W. Langmuir 1997, 13, 6447-6451. (10) Hotz, J.; Meier, W. Langmuir 1998, 14, 1031-1036.

Figure 1. Cryo-TEM micrograph of a 1.0 × 10-2 kmol/m3 DODAB surfactant vesicle solution, that is template vesicles (bar represents 100 nm).

polymerizations in bilayers of unilamellar vesicles, we focused on a qualitative morphological characterization.11 By means of cryo-electron microscopy we were able to study the morphology of both the template vesicles and the polymer-containing products. We have shown that this polymerization gives rise to the formation of interesting new parachute-like structures in which a polymer bead is confined to the bilayer of the vesicles, collapsed at one pole, see Figure 2. We concluded that incompatibility (11) Jung, M.; Hubert, D. H. W.; Bomans, P. H. H.; Frederik, P. M.; Meuldijk, J.; van Herk, A. M.; Fischer, H.; German, A. L. Langmuir 1997, 13, 6877-6880.

10.1021/la9907275 CCC: $18.00 © 1999 American Chemical Society Published on Web 10/19/1999

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Figure 2. Cryo-TEM micrograph of the polymer-vesicle hybrids (parachutes) as obtained after polymerization of styrene in the bilayers of the template vesicles (bar represents 100 nm).

between the synthesized polymer and the bilayer matrix is the driving force for the formation of these peculiar hybrids and that, as a consequence, templating does not occur. The parachute-like polymer-vesicle hybrids represent a new class of polymer colloids with highly interesting characteristics. These submicron colloidal structures exhibit multicompartmental features: The bilayer, the confined polymer bead, and the enclosed aqueous interior represent solubilization or encapsulation sites with distinct properties. A further characterization of these structures is believed to be interesting and is expected to reveal fundamental insight into the effect of the confinement of a polymer bead on the vesicle (bilayer) characteristics. The aim of the work presented in this paper is to obtain quantitative information on the shape, size, and electrical properties of the vesicular structures. For this, a study of the electrooptic birefringence was conducted, supplemented with data from light scattering. These obtained data are compared to earlier described cryo-electron microscopic results.11 Electrooptic birefringence (EB), or synonymously the Kerr effect, can be used to characterize macromolecular solutions and colloidal systems.12,13 The temporal behavior of the electrooptic signalsrise and decay, when switching the electric field on and offsis controlled by the rotational diffusion of the colloidal particles. Since this diffusion is dependent on the size and the shape of the particles, this technique enables quantification of these parameters. The magnitude of the equilibrium birefringence is a measure of the electrical and optical properties of the particles. The technique of transient electrooptic birefringence has been successfully applied to measure size, polydispersity, and dynamics for a variety of colloidal systems, for example, aqueous dispersions of disklike bentonite particles,14 surfactant solutions of rodlike micelles,15 or elongated inverted micelles.16,17 Transient electrooptic birefringence (TEB) studies on surfactant vesicles are reported only scarcely.18-22 These reports deal with this technique as a rapid and sensitive method to measure the (12) Schorr, W.; Hoffmann, H. In Physics of Amphiphiles: Micelles, Vesicles and Microemulsions; Degiorgio, V., Corti, M., Eds.; NorthHolland: Amsterdam, 1985; p 160. (13) Stoylov, S. P. Colloid Electrooptics: Theory, Techniques, Applications; Academic Press: London, 1991. (14) Peikov, V.; Sasai, R.; Stoylov, S. P.; Yamaoka, K. J. Colloid Interface Sci. 1998, 197, 78-87. (15) Schorr, W.; Hoffmann, H. J. Phys. Chem. 1981, 85, 3160-3167. (16) Mantegazza, F.; Degiorgio, V.; Giardini, M. E.; Price, A. L.; Steytler, D. C.; Robinson, B. H. Langmuir 1998, 14, 1-7. (17) Cirkel, P. A.; Koper, G. J. M. Langmuir 1998, 14, 7095-7103.

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size and the optical polarizability of vesicles. The observation that vesiclessalthough spherical in shapesexhibit orientational birefringence is explained by assuming small departures from true sphericity. These departures are due to thermal fluctuations or are induced by the electric field. Typically, in these measurements relaxation processes due to vesicle deformation are observed. Consequently, transient electrooptic birefringence allows determination of the size of the vesicles and represents a means to obtain information on the elasticity of the vesicle bilayer. Interestingly, the vesicles studied in this paper behaved in a particular way. It turned out that the amphiphile used gives rise to the formation of nonspherical vesicles with strong angularities; see Figure 1. Also, signs of relaxation processes due to vesicle deformation were absent. Both conducts are most likely related to a high bilayer rigidity. What is more, the parachute-like polymervesicle hybrids exhibit an electrooptic response that can be explained by the occurrence of a permanent dipole, whereas the bare vesicles orient due to the rise of an induced dipole. Apparently, the polymer concentration at one pole of the vesicles gives cause to this peculiar behavior. Experimental Section Materials. Unilamellar vesicles were prepared by extrusion of 10 mM dioctadecyldimethylammonium bromide (DODAB, Fluka) through a stack of three polycarbonate filters (200 nm pores, Millipore) at 60 °C (overpressure of 5 bar) in three subsequent passes. The extruded solution was kept at this elevated temperature for 12 h before being allowed to cool to room temperature. An opaque solution resulted from this procedure. In this solutionsreferred to as the template stock solutionsthe mass fraction of DODAB amounts to φm ) 0.63%. The vesicle-polymer hybridssreferred to as the parachute stock solutionsresulted from the following: monomer (20 mM styrene, Merck) and the photoinitiator (0.24 mM 2,2-dimethoxy2-phenylacetophenone, DMPA, Ciba-Geigy) were added to the solution to obtain overall concentrations as indicated. The monomer and initiator were absorbed by the vesicles after stirring overnight at room temperature. Photoinitiated polymerizations were performed at 25 °C using a pulsed excimer laser (XeF, λ ) 351 nm) at a 2 Hz pulse frequency and 30 mJ of energy per pulse. Conversion of the monomer was X ) 91%, as determined by HPLC. The weight-averaged molecular weight of the produced polymer was found to be Mw ) 35 kg/mol by size exclusion chromatography. Cryo-electron Microscopy. The colloidal stock solutions of the template and the parachute vesicles were visualized by means of cryo-TEM, a cryogenic microscopic technique.23 This technique allows the most direct and least perturbing morphological characterization at a detailed level, without the intervention of staining agents or laborious sample preparation (e.g. etching, fracture, or sublimation). Essentially, the preparation of a sample for cryo-TEM involves the physical fixation of a sample contained in a thin film. This fixation comprises the fast vitrification of the sample by rapid cooling through plunging into liquid coolant (ethane). Micrographs of the vitrified film were taken under low dose conditions after transfer to a cryo-holder and introducing the holder in the microscope. Vitrification was performed using a controlled environment vitrification system. (18) Ruderman, G.; Jennings, B. R.; Lyle, I. G. Int. J. Macromol. 1984, 6, 99-102. (19) Ruderman, G.; Jennings, B. R.; Dean, R. T. Biochim. Biophys. Acta 1984, 776, 60-64. (20) Wu¨rtz, J.; Hoffmann, H. J. Colloid Interface Sci. 1995, 175, 304317. (21) Scho¨nfelder, E.; Hoffmann, H. Ber. Bunsen-Ges. Phys. Chem. 1994, 98, 842-852. (22) Asgharian, N.; Wu, X.; Meline, R. L.; Derecskei, B.; Cheng, H.; Schelly, Z. A. J. Mol. Liquids 1997, 72, 315-322. (23) Frederik, P. M.; Stuart, M. C. A.; Bomans, P. H. H.; Lasic, D. D. In Handbook of Nonmedical Applications of Liposomes; Theory and Basic Sciences; Lasic, D. D., Barenholz, Y., Eds.; CRC Press: Boca Raton, FL, 1996; Chapter 15.

Novel Polymer-Vesicle Hybrids Transient Electrooptic Birefringence (TEB). Transient electrooptic birefringence experiments were performed with a conventional setup13,17 as follows: light from an 8 mW heliumneon laser (λ ) 632.8 nm) travels through a high-quality Glen Thompson polarizer. This polarizer is oriented in such a way that linearly polarized light is obtained with its axis of polarization at π/4 with respect to the electric field. After passing through the sample, the beam travels first through a polarizer before its intensity is converted to a voltage by a photomultiplier tube. The voltage of this tube and the voltage applied on the sample are measured simultaneously by a digital oscilloscope (LeCroy 9450). The voltage of the tube can be converted to obtain the birefringence signal ∆n(t). The sample is contained in a quartz cuvette with two parallel platinum electrodes. The electrodes are held together at a distance of 2.3 mm by two Teflon spacers. The optical path length is 49.3 mm. The temperature is kept constant by circulating water from a thermostated bath through a jacket that holds the cell. The sample was allowed to equilibrate at the set temperature for at least 2 h prior to the measurements. The voltage is supplied by a function generator (Hameg HM 8130) in combination with a high-speed power amplifier (NF Electronic Instruments 4020). In this way, a single pulse could be generated as well as a train of pulses. The response time of this setup is better than 1 µs. The investigations were restricted to the Kerr regime, where the induced stationary birefringence (∆n∞) is proportional to the square of the applied electric field E (E2 < 108V2/m2). The Kerr constant (K) is defined as this proportionality constant: K ) ∆n∞/E2. The pulse-duration lengths were chosen such that the induced birefringence reached a stationary value. Dynamic Light Scattering (DLS). Dynamic light scattering experiments were performed using a Malvern multiangle DLS apparatus (Malvern 4700, λ ) 488 nm). Stock solutions were diluted (dilution factor > 50, φm < 0.01%) before measuring the intensity autocorrelation function. Due to these dilutions, it was assumed that the measured diffusion coefficient by DLS represents the self-diffusion coefficient of the particles. The method of cumulant analysis was employed to calculate an intensityaveraged diffusion coefficient 〈D〉I orsequivalentlysa form-factorweighted z-averaged diffusion coefficient.24 This value represents the z-averaged diffusion coefficient for sufficiently small particles, or for particles of any size or shape in the limit θ f 0 (where the form factor reduces to 1).25,26 The 〈D〉I values as obtained from the cumulant analysis were measured for a range of scattering angles (10° e θ e 90°). The z-averaged diffusion coefficient 〈D〉 was calculated by linear extrapolation to an angle of 0°. The corresponding z-averaged hydrodynamic radius 〈R〉 then can be calculated by means of the Stokes-Einstein relation for spheres.24 The cumulant analysis also allowed determination of the socalled polydispersity index. This quantity measures the broadness of the decay rate distribution.24 More precisely, it represents the squared value of the relative standard deviation of the latter distribution; for a perfectly monodisperse sample, it should therefore be zero. Measurements were performed at 25 °C after enabling thermal equilibration of the sample.

Results The template vesicles (the bare vesicles, DODAB) and the parachute vesicles (the polystyrene-containing product vesicles) were both examined in terms of cryo-TEM, transient electrooptic birefringence, and dynamic light scattering. Cryo-TEM was utilized to visualize the shape and the structural details of the colloidal entities.11 Par excellence, this cryogenic technique allows direct and unperturbed morphological characterization at a detailed level. Figures 1 and 2 are micrographs of stock solutions of the template and the parachute vesicles (φm ) 0.63%). Both micrographs show that the vesicular structures are unilamellar in (24) Selser, J. C.; Yeh, Y.; Baskin, R. J. Biophys. J. 1976, 16, 337356. (25) Doi, M.; Edwards, S. F. The Theory of Polymer Dynamics; Clarendon Press: Oxford, 1986. (26) Ruf, H.; Georgalis, Y.; Grell, E. Methods Enzymol. 1989, 172, 364-390.

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Figure 3. Electrooptic birefringence (∆n∞) as a function of the square of the applied electric field (T ) 20 °C). The linear dependencies show that the systems obey the Kerr law.

nature; that is, one single bilayer builds a vesicle. Interestingly, the micrograph of the template vesicles displays ellipsoidal structures with strong angularities. Although this behavior is known for DODAB, it is yet not completely understood.11,27 Remarkably, the morphology of the parachute vesicles is less angular. A striking observation is to be made when looking at the difference in the diversities of the morphologies. The template vesicles exhibit a wider variety of structures than the parachute ones. Transient Electrooptic birefringence. First, the Kerr regime was searched for. At electric field strengths of E < 104 V/m the stationary electrooptic birefringence (∆n∞) linearly depends on the field strength squared, as in accordance with the Kerr law. This regime roughly coincides with the field strengths reported for previous vesicle studies.22 Figure 3 shows this linear behavior for both the bare vesicles (templates) and the polymercontaining vesicles (parachutes) at 25 °C (after 50× dilution, φm ) 1.3 × 10-4). The Kerr constants K were calculated to be 2.8 × 10-18 m2/V2 and 1.4 × 10-15 m2/V2 for the template vesicles and the parachute vesicles, respectively. Typical pulse lengths δ were 15 × 10-3 s. To investigate possible influences of particle-particle interaction, samples with different degrees of dilution were measured. For samples with 2 × 10-5 < φm < 6 × 10-4, the Kerr constant was shown to linearly depend on φm. Both for higher and lower values, the measured Kerr constant was found to be lower than what would be expected from extrapolation. This aberration for φm < 2 × 10-5 is likely to be due to a surface adsorption effect, as a simple calculation shows that the surface area of the electrodes and the cuvette is then comparable to the total area of the vesicles. The deviations for φm > 6 × 10-4 may be accounted for by emerging intervesicular interactions or even clustering. Further, in a series of experiments, the temporal behavior of the induced electrooptic birefringence was examined for in and out of field responses; see Figure 4. These measurements were performed at 25 °C (φm ) 1.2 × 10-4). The relaxation processes of the in and out of field birefringence of the template vesicles could be fitted with a one-exponential function.

∆ndecay(t) ) ∆n∞ exp(-t/τ) or ∆nrise(t) ) ∆n∞[1 - exp(-t/τ)] (27) Laughlin, R. G.; Munyon, R. L.; Burns, T. W.; Coffindaffer, T. W.; Talmon, Y. J. Phys. Chem. 1992, 96, 374-383.

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Figure 4. Temporal behavior of the birefringence for both samples at E ) 6.0 kV/m and T ) 25 °C. Table 1. Relaxation Time for the Rise and Decay of the Electrooptic Birefringence (E ) 6.0 kV/m and T ) 25 °C) sample templates parachutes

rise [in field] τr (10-3 s)

decay [out of field] τd (10-3 s)

1.35 ( 0.02

1.05 ( 0.02 1.72 ( 0.02

Table 2. Translational and Rotational Diffusion Coefficients as Obtained from Analysis of Dynamic Light Scattering and of the Rise and Decay Electrooptic Birefringence, Respectively (T ) 25 °C) Drot (s-1) sample

rise

decay

〈Dtrans〉 (10-12 m2/s) extrapolated

templates parachutes

124 ( 2 112 ( 2

159 ( 3 97 ( 2

2.32 ( 0.03 2.37 ( 0.03

In Figure 4, for reasons of better visual comparison, the out of field response is depicted after transformation by subtraction of ∆n∞ and inversion with respect to the time axis. For infinite dilution, where there are no interparticle interactions, rotation diffusion coefficients can be obtained from the measured relaxation times of transient electrooptic birefringence.13,25 The theory for small colloidal particles predicts that the relaxation times for rise and decay are identical in the case of pure induced dipole moment orientation:

Drot )

1 6τ

Since the rise and decay of the electrooptic birefringence for the template vesicles can be fitted with a single relaxation time constant, it is straightforward to calculate the corresponding rotational diffusion coefficient. The results are shown in Tables 1 and 2. For the parachute vesicles, the out of field response was characterized by a single exponential as well. However, the rise of the signal for these parachute structures could not be described by a single-exponential rise function. This pointed out that a completely different orientation mechanism governs the rise of the birefringence. It was found that the rise of the signal could well be fitted by a two-exponential function that holds for transient rise electrooptics effects for colloids also bearing a permanent dipole moment:25

[

3R exp(-2Drott) + 2(R + 1) µ2 R-2 exp(-6Drott) with R ) ∆RkBT 2(R + 1)

∆n(t) ) ∆n∞ 1 -

]

where R represents the ratio of the permanent and induced

Figure 5. Influence of field reversal on the measured optic birefringence signal of the parachute vesicles at T ) 25 °C (the arrows indicate the onset of the field reversal).

dipole moments term, µ respresents the permanent dipole, and ∆R is the anisotropy of the electric polarizability. The fit resulted in R ) 7 ( 1 and Drot ) 112 ( 2 s-1. This latter value corresponds reasonably well with the rotation diffusion coefficient obtained from the decay of the electrooptic effect; see Tables 1 and 2. The interpretation in terms of a permanent dipole is further supported by field-reversal experiments; see Figure 5. The arrows indicate the onset of the field reversal. The observation that upon field reversal the electrooptic birefringence reduces strongly before it is restored to its initial stationary value corresponds to what is expected for permanent dipoles, as the particles are forced to realign in the opposite direction.28 In contrast, the birefringence of the template vesicles is unaffected by field reversal, which permits the conclusion that these particles couple to the electric field by induced dipole moments. This induced dipole behavior is further supported by the fact that the rise and decay functions are virtually identical; see Figure 4. An important factor in the explanation for the large difference in magnitude of the birefringence (see Kerr constants) between the templates and the parachutes is this intrinsic difference in coupling mechanism. Another factor that contributes to the higher magnitude is the larger refractive index difference between the particle and the medium12,16 in the case of the parachutes caused by the presence of the polymer (refractive indexes of the water, the bilayer, and the polystyrene bead: 1.33, ∼1.4, and 1.6, respectively, at 20 °C and λ ) 589 nm29,30). In additional experiments, the Kerr constants were determined as a function of temperature; see Figure 6. In these experiments, the samples were equilibrated for at least 2 h at each temperature. As a check for reversibility, the samples were allowed to cool to room temperature, and the Kerr constants were determined again at this temperature. It was noticed that the Kerr constant for the solution of the template vesicles recuperated to its initial value, whereas the constant for the solution of parachute vesicles was remarkably reduced to ∼50% of its original value. Dynamic light scattering (DLS) measurements were performed to determine the translation diffusion coefficient of the template and the parachute vesicles. The apparent diffusion coefficient (the form-factor weighted z-averaged diffusion coefficient 〈D〉I) and the polydispersity (28) Fredericq, E.; Houssier, C. Electric Dichroism and Electric Birefringence; Clarendon Press: Oxford, 1973. (29) Weast, R. C.; Astle, M. J.; Beyer, W. M. CRC Handbook of Chemistry and Physics; CRC Press: Boca Raton, FL, 1985. (30) Brandrup, J.; Immergut, E. H. Polymer Handbook; WileyInterscience: New York, 1989.

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Figure 6. Kerr constant (K) as a function of the cell temperature.

index were determined at different scattering angles using the method of cumulant analysis. Linear extrapolation of the apparent diffusion coefficients to zero angle resulted in the z-averaged values for the diffusion coefficients 〈D〉; see Table 2. The polydispersity indexes were measured to be 0.19 and 0.10 for the templates and the parachutes, respectively. Discussion Size and Shape. The micrographs (Figures 1 and 2) allow rough estimations of the size and the shape of the templates and the parachutes. Judging from these photos, template vesicles seem to display a wider variety in shape and size than parachute vesicles. Roughly, the templates are characterized by a long axis of ∼150 nm. The parachutes are less angular, and more spherical, and measure a radius of ∼100 nm. A more quantitative and macroscopic characterization is obtained by DLS and transient electrooptic birefringence. Dynamic light scattering shows that the polydispersity index for templates is almost twice as large as that for parachutes. This parameter quantifies the qualitative conclusion of a wider variety of structures in the template stock solution. Next, the equivalent sphere radii of the vesicles may be obtained by using the averaged rotation and translation diffusion coefficient from the transient electrooptic birefringence relaxation and the dynamic light scattering data, respectively. This is done in Table 3 by interpreting these motional relaxations due to independent monodisperse particles with a spherical shape; see Appendix. For both templates and parachutes, these radii are in accordance with rough estimates from the micrographs. According to Table 3, there is a difference in radii obtained by measurements of electrooptic birefringence and dynamic light scattering. Apart from the interparticle interactions which were excluded (see Results), two reasons can account for this difference: the anisotropy and the polydispersity of the particles. A quantification of the anisotropy can be addressed by the following: As stated before, the rotation diffusion is dependent on the size and geometry of the particles. The same holds for translation diffusion. Both types of diffusion hinge differently on the size and shape. Therefore, from the combination of both diffusion coefficients, size and shape parameters can be determined. In our approach we assume that the shape of the particles is amenable to being modeled as ellipsoids. The prolate spheroid is a rodlike shape generated by rotating an ellipse around its long semiaxis a, with the two shorter semiaxes b identical. The oblate spheroid is a disklike shape generated by rotating an ellipse about its short semiaxis a, the two long

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semiaxes b being identical. We define the axial ratio as p ) a/b (p > 1 and p < 1 for prolate and oblate ellipsoids, respectively). This assumption and description leaves us with two independent parameters that define the shape of the particles (the long semiaxis and the axial ratio). The rotation and translation diffusion constants are functions of these two parameters, for each spheroid in a different way; see Appendix. Resolving the corresponding equations with the use of the determined diffusion coefficients results in the shape parameters as summarized in Table 3. Assuming prolate geometry, the bare vesicles are described by ellipsoids with the long semiaxis equal to 106 nm and an axial ratio of 1.4, whereas the parachute vesicles are characterized by a long semiaxis of 151 nm and the ratio 2.6. When assuming oblate geometry, these values read 132 nm (p ) 0.43) and 166 nm (p ) 0.0050) for templates and parachutes, respectively. When comparing these results with the micrographs, it can be concluded that calculated dimensions for the templates comply rather well, whereas for the parachutes they deviate significantly from what can be estimated from the photos; see Figure 2. This difference might well be related to particle polydispersity in both size and shape. The measured translation diffusion coefficient represents a z-averaged value.26 The kind of average obtained from transient electrooptic birefringence is difficult to assess, since it depends on the way the particles couple to the electric field. The parachute particles couple differently to the field than the templates do (vide infra). This may explain why, although the light scattering results and micrographs show a higher degree of polydispersity in the templates compared to the parachutes, the effect seems to induce a larger deviation in the model parameters for the parachutes. Electric Field Response. The template vesicles seems to couple to the electric field by an induced dipole moment. This result has also been found in previous electrooptic birefringence studies on vesicles.18-22 However, in these studies the electric field response (both the in and out field behavior) could not be fitted to a single-exponential function. Typically, a biexponential function was used. In some of these studies cryo-electron micrographs were taken, revealing vesicles of an almost perfectly spherical shape.20,21 In order for such a vesicle to be able to make the solution birefringent upon orientation, the vesicle should become nonspherical. The biexponential decay was therefore explained as being the result of a combination of a deformation and an orientation mechanism. Figure 1 shows that our choice of surfactant gives rise to template vesicles of a nonspherical shape with strong angularities. Consequently, deformations are not a prerequisite to display birefringence anymore. The observation of a singleexponential decay indicates that the vesicles resist deformation under the action of the field. A plausible explanation for the apparent lack of deformations can be given in terms of the phase structure of the bilayer at the examined temperatures. Typically, surfactant vesicles exhibit a phase transition from an ordered gel-like state to a more liquid-crystalline-like state at the so-called phase transition temperature Tm.31 This transition in phase structure is accompanied by an increase in the mobility of the individual molecules in the bilayer and a decrease in the bending elastic modulus of the bilayer.32 The systems (31) Seddon, J. M.; Templer, R. H. In Structure and Dynamics of Membranes; Sackmann, E., Lipowski, R., Eds.; Elsevier: Amsterdam, 1995; Chapter 3. (32) Sackmann, E. In Structure and Dynamics of Membranes; Sackmann, E., Lipowski, R., Eds.; Elsevier: Amsterdam, 1995; Chapter 5.

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Table 3. Equivalent Sphere Radius 〈R〉 and Equivalent Parameters for a Prolate Shape Calculated Using the Rotational and Translational Diffusion Coefficients in Table 2 sample

〈R〉 [from 〈Drot〉] [10-9 m]

〈R〉 [from 〈Dtrans〉] [10-9 m]

long semiaxis [10-9 m]

axial ratio p ) a/b

templates parachutes

109 ( 19 121 ( 12

105 ( 3 103 ( 2

106 151

1.4 2.6

studied by others (natural lipids and short chain surfactants) are expected to display a Tm below or close to room temperature, whereas our system (DODAB) exhibits this transition at T ) 43 °C.33 The solid-like state of the DODAB bilayer at room temperature is likely to resist bilayer deformations (high bending modulus).32 The parachute-shaped polymer-vesicle hybrids, although having an outside contour which is rather spherical, are optically anisotropic due to the concentration of polymer at one pole. The orientation of such a particle is therefore also likely to give rise to birefringence, without any deformation being necessary. However, the mechanism by which these parachutes couple to the electric field is clearly different from the templates. Three observations lead to the conclusion that the difference originates from a permanent dipole on the parachutes: (1) The shape of the in and out field response (Figure 5), (2) the result of the field inversion experiments (Figure 6), and (3) the magnitude of the Kerr effect on these hybrids (about three orders higher compared to the templates). Where does this permanent dipole come from? The dipole induced on vesicles in an electric field is believed to arise from a redistribution of charges, both along the surfactant bilayer and in the counterion cloud. The confinement of a polymer bead to one pole of the vesicle seems to create a similar type of charge redistribution. Now that we have a qualitative understanding about the mechanism by which the vesicles couple to the electric field, we could try to assess how the nonuniformity of the particles affects the birefringence signals. The results from dynamic light scattering suggest a considerable amount of size polydispersity. Moreover, the electron micrographs show a variation in the particle morphology especially for the templates. In principle, this would lead to a deviation from a single-exponential decay. The degree of this deviation, however, depends on the shape of the particles, as well as on the nontrivial way in which the particles couple to the electric field. Apparently, for both templates and vesicles, the deviation from a single-exponential decay becomes too small to be determined. Nonetheless, the relaxation time and the rotational diffusion coefficient calculated from these signals will be averages in which vesicles of different shapes and sizes will have a different weight. This can explain why the discrepancy, outlined in the previous paragraph, between the particle sizes from light scattering and transient electrooptic birefringence is larger for the parachutes when compared to the templates. The overestimation of the parachutes’ size by transient electrooptic birefringence can be explained by the fact that large vesicles carry a large dipole moment and thus have a high weight in the average rotational diffusion coefficient. Temperature Dependence. Finally, the temperature dependence of the Kerr constant is intrinsically different for the two types of vesicles; see Figure 6. The template vesicles show a reversible and steady behavior of the Kerr constant with respect to temperature. In contrast, the temperature response of the parachute vesicles displays an abrupt decrease between 35 and 40 °C and the original

Kerr constant value is not restored again when cooling to the initial temperature. Irreversible structural changes that are induced at higher temperatures could account for this observation. We suggest that this structural change is due to a partial release or expulsion of the polymer beads, as was supported by cryo-electron micrographs (photos not shown). This release will result in a decrease in birefringence, since the bare vesicles that result from this transformation exhibit significantly smaller birefringence. The fact that the general tendency of the Kerr constant decreases with temperature is expected, since the orientation processes are opposed by the increased Brownian motion due to the higher thermal energy of the system.

(33) Nascimento, D. B.; Rapuano, R.; Lessa, M. M.; Carmona-Ribeiro, A. M. Langmuir 1998, 14, 7387-7391.

The rotation and translation diffusion coefficients are functions of the geometric parameters. In the case of

Conclusions The combination of transient electrooptic birefringence, cryo-electron microscopy, and dynamic light scattering allowed a detailed quantitative analysis and characterization of the vesicular systems. Both the template and the parachute vesicles were characterized, and the results were compared. The fact that the measured transient electrooptic birefringence signals did not display relaxations that could be attributed to a vesicle deformation step confirms the anisotropic shape of the vesicles as seen in the micrographs and can be explained by their intrinsic rigidity. To the best of our knowledge, this is the first vesicular system that acts as rigid particles under the action of an electric field. The solid-like phase structure of the bilayer at the experimental temperatures is suggested to account for this phenomenon. It was found that template vesicles orient in an electric field due to the rise of an induced dipole. The parachute vesicles with the confined bead, however, seem to possess a permanent dipole. Measurements on the reversibility of the Kerr effect with respect to temperature pointed out that irreversible structural changes take place in the case of the parachute vesicles, whereas the template vesicles seem reversible in their thermal behavior. This irreversible structural change is expected to originate from the partial expulsion of the polymer beads. To conclude, the presence of a polymer bead inside the bilayer of the vesicles strongly affects the dynamics and electrooptic behavior of the vesicles to which it is confined. The parachute vesicles represent an interesting class of polymer colloids regarding their composition, morphology, and electric/optic behavior. Acknowledgment. This work was supported by funding from the Strategic Research Fund of Imperial Chemical Industries (ICI, U.K.). M.J. would like to thank The Netherlands Foundation for Chemical Research (NWO/CW). The authors kindly acknowledge Dr. P. M. Frederik and P. H. H. Bomans (Maastricht University, The Netherlands) for their valuable cryo-TEM work. Appendix

Novel Polymer-Vesicle Hybrids

Langmuir, Vol. 15, No. 26, 1999 8855

prolate spheroids, the rotation diffusion is given by25,34,35

Drot )

1)1/2) + 1 - p2]-1

kBT with ζrot ) ζrot

( )[

]

2 16π 3 1 (2p - 1) ηa 1 - 4 ln(p + (p2 - 1)1/2) - 1 2 1/2 3 p p(p - 1)

-1

whereas the translation diffusion is described by

Dtrans )

ζrot ) 16πηb3(1/p - p3)[3(1/p2 - 1)1/2 arctan((1/p2 -

kBT with ζtrans ) ζtrans 6πηa(1 - 1/p2)1/2[ln(p + (p2 - 1)1/2)]-1

where ζrot, ζtrans, and η represent the rotation friction constant, the translation friction constant, and the dynamic viscosity, respectively. A similar set of equations can be found to describe oblate ellipsoids. The relevant friction constants for these oblate structures depend on the relevant geometric parameters as follows:34 2 1/2

2

for the rotation friction constant. The formula expression for the rotation friction constant was derived from expressions for rotation friction constants36 in the three axial directions, assuming equipartition between these axial directions. If both types of diffusion constants are known, the geometric parameters a (or b) and p can be easily resolved when presuming a certain shape. It has to be realized that this approach assumes reasonable monodispersity of the particles and accuracy of the data. Additionally, equivalent sphere radii 〈R〉 of the particles may be obtained by using the averaged rotation diffusion coefficient from the transient electrooptic birefringence relaxation data, with

Drot )

kBT 8πη〈R〉3

1/2 -1

ζtrans ) 6πηb(1 - p ) [arctan((1/p - 1) )] for translation and

when interpreting this motional relaxation due to independent monodisperse particles with spherical shape. LA9907275

(34) Probstein, R. F. Physicochemical Hydrodynamics; Wiley-Interscience: New York, 1994. (35) Hunter, R. J. Foundations of Colloid Science; Clarendon Press: Oxford, 1987; Vol. 1.

(36) Lyklema, J. Fundamentals of Interface and Colloid Science, Academic Press: London, 1991; Vol. 1.