7130
J. Phys. Chem. 1996, 100, 7130-7134
Counterion Effects on Colloid Stability of Cationic Vesicles and Bilayer-Covered Polystyrene Microspheres L. R. Tsuruta and A. M. Carmona-Ribeiro* Departamento de Bioquı´mica, Instituto de Quı´mica, UniVersidade de Sa˜ o Paulo, CP 26077, Sa˜ o Paulo SP, Brazil ReceiVed: August 30, 1995; In Final Form: NoVember 27, 1995X
The major factor responsible for the very low colloid stability (W) of dioctadecyldimethylammonium (DODA) chloride, bromide, or acetate vesicles in the presence of monovalent salt is identified. Large DODAAc vesicles prepared in 0.6 M D-glucose remain stable over the entire range of NaAc concentrations tested (0-150 mM NaAc). The pH measured for this highly concentrated D-glucose solution is 5.1-5.3. Because acetic acid is a neutral and small molecule that can freely permeate the DODAAc vesicle, upon addition of an acidic NaAc plus D-glucose solution to the vesicle outside, acetic acid forms and reaches the vesicle interior acting as an acetate carrier; acetate binds to the inner vesicle surface and eliminates any charge asymmetry between the inner and outer surface. Counterions such as chloride or bromide do not cross the membrane to reach the inside. Thereby asymmetry of the charge distribution generated upon external addition of NaCl or NaBr, respectively, to DODACl or DODABr vesicles cannot be relaxed, causing the hydrophobic defects that decrease W. For DODA vesicles and bilayer-covered polystyrene microspheres of similar sizes, W is systematically higher for the covered particles. For the large bilayer-covered particles, salt-induced aggregation is reversible upon salt removal, whereas for the small ones it is not. From bromide to acetate, counterion size and hydration increases and so does W. DODAAc vesicles at low pH and covered particles are the most stable dispersions. DODAAc from vesicles adsorbs with high affinity onto sulfate latexes. An adsorption maximum at intermediate supernatant concentrations typically points out a competition between vesicle/vesicle adhesion and vesicle/ latex interaction as DODAAc concentration increases in the supernatant.
Introduction Charged synthetic amphiphile vesicles are perfectly rigid spherical shells surrounding an aqueous compartment.1 They are highly charged and have a much smoother interface than other colloids. Because of these properties, they should eventually behave as model colloids. They have indeed been used to test theoretical models.2,3 However, their polydispersity is still a disadvantage compared with highly monodisperse systems such as polystyrene microspheres.4 On the other hand, the possibly hairy, rough, or conducting surface of these microspheres often represents a problem. Thus, the interaction between synthetic amphiphile vesicles and oppositely charged polystyrene microspheres has been proposed as a possible route to produce an ideal model colloid: homodisperse and smooth bilayer-covered polystyrene microspheres.4,5 In contrast to single-chained surfactants, dioctadecyldimethylammonium (DODA) salts are effective flocculants or stabilizers for oppositely charged dispersions in solution at very low concentration.4-7 Adsorption isotherms for DODA onto sulfate polystyrene are of high affinity, and bilayer deposition onto the latexes is a conspicuous event .4,5,8 However, depending on DODA counterion, after bilayer deposition, increasing vesicle concentration may cause vesicle adhesion (with or without vesicle rupture) to the bilayer-covered latex.8 Returning to the vesicle structure problem, this adhesion was shown to be due to interdigitation in the vesicle membrane; interdigitation being associated with a low extent of specific counterion binding at low ionic strength.8,9 Possibly, the best type of bilayer to cover the microspheres and produce an ideal colloid is that obtained * To whom correspondence should be addressed. E-mail: mcribeir@ iq.usp.br. X Abstract published in AdVance ACS Abstracts, March 15, 1996.
0022-3654/96/20100-7130$12.00/0
from the interaction between the bromide derivative (DODABr) and sulfate polystyrene microspheres.5 The specific binding of bromide at the DODA bilayer surface prevents interdigitation in the vesicle structure, so that increasing vesicle concentration does not lead to vesicle adhesion to the bilayer-covered latex over a relatively broad range of DODABr concentration in the supernatant.8 For latexes covered with DODABr bilayers, a maximum in colloid stability as a function of particle size was observed5 which was qualitatively consistent with predictions from DLVO models that take into account aggregation at a secondary minimum.10 In this work, the issue of compared colloid stability of DODA cationic vesicles and DODA-supported bilayers is addressed over a range of monovalent salt concentration for DODA with acetate, chloride, and bromide as counterions and two different supporting particle or vesicle sizes. The results shed new light on the importance of transmembrane symmetry of charge to obtain a nondefective and stable synthetic amphiphile membrane. Material and Methods Dioctadecyldimethylammonium bromide, DODABr (99.9% pure), was obtained from Sigma Chemical Co. (St. Louis, MO) and used as such. DODACl was obtained by ion exchange from DODABr using Amberlyst A-26 in the chloride form from E. Merck (Darmstadt, Germany). After the bromide/chloride exchange, DODACl in methanol was precipitated, filtered, washed, recrystallized from 1:1 ether/acetone, and dried as previously described.11,12 DODAAc was obtained by ion exchange from DODABr using Amberlyst A-26 in the acetate form and purified by recrystallization from ether.12 All other © 1996 American Chemical Society
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TABLE 1: Properties of Sulfate Polystyrene Microspheres in Water at 25 °C mean diam (nm)
area per charge group (nm2)
SSA (cm2 g-1)
76 249
7.1 7.7
748 316 228 401
reagents were analytical grade and used without further purification. Water was Milli-Q quality. Large DODABr, DODACl, or DODAAc vesicles (LV) were prepared by injecting a chloroform solution of the amphiphile into a 0.6 M D-glucose solution, 70 °C.13 Small vesicles (SV) were prepared by sonication with tip.14 DODA concentration in the dispersions was determined by solubilization of a dye/ amphiphile complex in non-ionic micelles15 or by microtitration.16 Charged polystyrene microspheres stabilized by negative sulfate charges and described as ultraclean by the supplier were obtained from Interfacial Dynamics Co. (Portland, OR) and used as supplied. The main characteristics of the microspheres as specified from the supplier are given in Table 1. Mean diameters were obtained by the supplier using an electron microscope. Microspheres were diluted in the same solution where vesicles were prepared. These solutions were pure water, 0.6 M D-glucose, 1 mM NaAc, or 0.6 M D-glucose and 1 mM NaAc depending on the vesicles preparation media. Interactions between vesicles and microspheres was induced by adding the vesicles to the polystyrene. The proportion between total surface areas for vesicles and particles (Av/Ap) was kept inside the limits that assure bilayer deposition and minimize subsequent vesicle adhesion,5 i.e., 5 > Av/Ap > 1. The proportion Av/Ap is specified for each experiment. Mixtures were thermostated at 25 °C for 1 h before W or adsorption measurements were carried out. For the DODAAc adsorption measurements, mistures were centrifuged at 14 000g for 1 h at 15 °C to separate particles from vesicles. Aliquots of supernatant and original vesicle preparation were used to determine DODAAc concentration.15 Total surface area on the polystyrene (Ap) was calculated from the particle number density and the mean particle radius given by the supplier. Total surface area on the vesicles (Av) was calculated from the area per monomer for DODAAc at an air/water interface, i.e., 0.65 nm2,17 the amphiphile concentration in the vesicle/particle mixture, the final volume of the mixture, and the fully interdigitated monolayer model for the vesicle structure.9 Adsorption was expressed as the number of amphiphile molecules adsorbed per square meter of polystyrene. The colloid stability of the covered particles after monovalent salt addition was characterized by measuring turbidity at 400 nm in a Hitachi U-2000 spectrophotometer as a function of time. The salt was NaBr, NaCl, or NaAc added to DODABr, DODACl, or DODAAc bilayer-covered microspheres, respectively. The time lag between mixing and the start of the register was usually smaller than 6 s. The initial flocculation rate (V0) was calculated from the turbidity kinetics. Mean values were calculated from, at least, two independent kinetics for each salt concentration. The inverse of the initial flocculation rate was taken as directly related to the experimental stability factor of the dispersions (W), the proportionality constant being calculated from rapid flocculations (W ) 1).18 Colloid stability for vesicles was obtained using the same procedure described above. Osmotically nonresponsive SV1 were prepared in water and salt solutions were added to have their colloid stability determined. On the contrary, LV, which are responsive to osmotic gradients, were prepared in D-glucose 0.6 M and kept at isoosmotic conditions (0.66 Osm/kg) after salt addition. Thus, salt solutions to be added to LV contained always the complementary amount of D-glucose necessary to
keep osmolarity at 0.66 Osm/kg. pH measured for all solutions containing D-glucose was equal to 5.1-5.3 and therefore, very close to the apparent pK for dissociation of acetic acid that is equal to 4.8. Reversibility (R) of aggregation19 was measured for DODABr covered microspheres with 76 and 249 nm of mean diameter previously aggregated upon salt addition over a range of concentrations (0-0.15 M NaCl). The turbidity (400 nm) increase after salt addition was then followed for 10 min. The turbidity at 10 min is A1. As a control, the turbidity of covered microspheres after water addition (A1c) was also measured. Thereafter, samples (with salt) and control (without salt) were submitted to 2 h dialysis against 1 L of DODABr dispersion in water at the same DODABr concentration in the mixture. To prevent any eventual DODABr uptake by the dialysis bag, dialysis membranes were incubated overnight at the DODABr concentration in the sample and control mixtures prior to dialysis. After dialysis, the turbidity at 400 nm of the samples and control dialysates was measured: A2 and A2c, respectively. Because a small amount of covered microspheres remained attached to the dialysis bag after dialysis, turbidity measurements had to be corrected by a dilution factor given by A1c/A2c. R was taken as
R ) 1 - [(A1c/A2c)(A2 - A2c)/(A1 - A1c)] Results and Discussion 1. Colloid Stability for Vesicles and Supported Bilayers: Size and Counterion Effects. DODA cationic vesicles, though much studied,1 are poorly characterized from the point of view of counterion effects on vesicle properties.8,9 Among these is their colloid stability. In general, the most studied vesicles were those made up of DODACl, and these presented a very low colloidal stability in the presence of monovalent salt due to the occurrence of hydrophobic defects in the bilayer.19 Later on, evidence for interdigitation in the amphiphile bilayers was presented8,9 and deemed as a possible cause for the additional attractive force between vesicles resulting in a low colloid stability. Under these circumstances, our rationale for the present work was that the interaction between these cationic vesicles and the sulfate polystyrene microspheres by formation of ion pairs4 might orient deposition of a less interdigitated bilayer onto the latexes, and this would appear as an increase in colloid stability for the covered microspheres. Overall, the results in Figure 1 corroborate the reasoning above since W (Figure 1) and CCC (Table 2) values are sistematically higher for covered microspheres in comparison with W for vesicles of similar sizes. One should compare Figure 1A for small covered microspheres with Figure 1C for SV, and Figure 1B for large covered microspheres with Figure 1D for LV. However, in the case of DODAAc LV and covered particles, the contrary was observed. DODAAc LV prepared in a 0.6 M D-glucose solution with or without 1 mM NaAc are extremely stable upon NaAc addition (Figure 2 A,B, respectively). At first it was difficult to envisage how this high colloid stability could have been achieved for this particular vesicle type. A few possible explanations were available. The fully interdigitated nature of DODAAc vesicles9 might result in a highly stable dispersion not affected by addition of large amounts of NaAc to the vesicle outside because acetate would not bind at all even when present in large amounts. This possibility was quickly eliminated since DODAAc SV, which are also interdigitated (see the last subsection), are very unstable (Figure 1C). Acetate in the form of acetic acid might have been permeating the vesicle membrane from the outside, dissociating inside and binding
7132 J. Phys. Chem., Vol. 100, No. 17, 1996
Figure 1. Colloid stability (W) as a function of monovalent salt concentration (C) for bilayer-covered polystyrene microspheres with 76 (A) and 249 nm of mean diameter (B) and for small (C) or large DODA vesicles (D). The monovalent salt used to induce flocculation was NaAc, NaCl, and NaBr for DODAAc, DODACl, and DODABr covered particles or vesicles, respectively. Details on experimental conditions are given in Table 2.
similarly to the vesicle inner and outer surface. This last mechanism would be unique for acetate but would be relevant only at pH values not too far from the pKa for acetic acid, which is equal to 4.8. This led us to measure the pH of all D-glucose or D-glucose plus NaAc solutions: it was 5.1-5.3. Therefore, a substantial amount of acetic acid is present to carry acetate to the vesicle interior. Only acetate among the three counterions can form a weak acid by protonation and permeate the vesicle via its neutral form. The consequence of this permeation would be the relaxation of charge asymmetry between the outer and the inner membrane surface. For chloride or bromide, addition of the corresponding salt to the vesicle outside would result in specific counterion binding at the outer vesicle surface that would not be balanced by an equivalent counterion binding to the inner membrane surface. Hence hydrophobic defects would appear in the bilayer leading to a low colloid stability for DODACl and DODABr LV (Figure 1D). Additionally, a repulsive short-ranged hydration force was described for interacting DODAAc layers12 that is absent for DODABr.11 Thus some binding of the large and hydrated acetate anion to the DODAAc membrane could well account for stabilization via development of an hydration shell. Contrasting with the very high stability of LV (Figure 2), W for DODAAc SV is rather low (Figure 1C). Not only are preparation media different (DODAAc SV are prepared in water, and LV in 0.6 M D-glucose) but also, and more important, sonication is a drastic procedure for dispersing amphiphiles and forming vesicles. Membrane fragments and/or quite small strained vesicles that spontaneously fuse along time are produced.1 Specific counterion and size effects on colloid stability of vesicles and supported bilayers are summarized in Table 2. The highest and lowest critical coagulation concentration (CCC) values are those for DODAAc and DODABr, respectively. Acetate is the largest, more hydrated, and less tightly bound counterion, followed by chloride and bromide. CCC values follow this order both for vesicles and bilayer-covered microspheres (Table 2). Increasing the size for vesicles or covered particles also increases CCC (Table 2).
Tsuruta and Carmona-Ribeiro The stabilizing role of D-glucose suggested for DODAAc LV, which was actually due to the low pH that accompanies dissolution of large amounts of D-glucose in water, was not found for DODACl SV. CCC for DODACl SV prepared in 0.43 M D-glucose 0.43 M from ref 2 is 0.122 M NaCl,2 whereas from Table 2 and for the same vesicle preparation in water it is 0.091 M NaCl. CCC for DODACl LV in 0.6 M D-glucose is 0.117 M NaCl (Figure 1D; Table 2). This value is smaller than the 0.141 M NaCl previously obtained for the same vesicle preparation under completely analogous experimental conditions but using DODACl that had a certain proportion of unequal chain lengths.2 This might have favored interdigitation.21 The present results were obtained with amphiphiles that are highly pure and composed of 99.9% C18 double chains. Therefore, if interdigitation occurs, it is not due to unequal chain lengths. Interdigitation increases molecular packing inside the bilayer21 what could possibly increase colloid stability. In fact, the highest colloid stability in this study is that obtained for DODAAc LV (Figure 2), a system where evidence of full interdigitation has been reported.8,9,12 The effect of size on W is depicted from the comparison between Figure 1A,B for the supported particles. W is significantly smaller for the larger supported bilayers. For vesicles, a similar tendency is observed (compare Figure 1C with 1D), W being slightly larger for the smaller vesicles. As it should be, there is aggregation at a secondary minimum for both LV and large covered particles.5,19,20 Good evidence for this is given in the next subsection from experiments of reversibility of salt-induced aggregation for small or large DODABr covered microspheres upon salt removal. 2. Reversibility of Aggregation for Small and Large Bilayer-Covered Microspheres. At a given NaCl concentration, colloidal stability of bilayer-covered latexes attains a maximum at 40-50 nm covered particle radius and then decreases as a function of size.5 The explanation for this result is to be found in theoretical models for the colloid stability that take into account aggregation at the secondary minimum.10 As monovalent salt concentration and size increase, the depth of the secondary minimum increases19 as does its importance as a destabilizing factor. To definitely demonstrate whether aggregation for the large bilayer-covered latexes in Figure 1B could be accounting for the observed low colloid stability and also to explain the previously described decrease in stability upon an increase in particle size,5 experiments to determine reversibility of aggregation upon salt removal are shown in Figure 3. While for the large bilayer-covered particles the reversibility is close to 1 up to 0.1 M NaCl, for the small bilayercovered latexes it was rather low and close to zero. Therefore, at least up to 0.1 M NaCl, aggregation is reversible upon salt removal and can be associated with adhesion of bilayer-covered microspheres at a secondary minimum. 3. Hydrophobic Attraction between Interdigitated DODAAc Vesicles and Latexes. When DODACl or DODABr with 100% of double C18 are dispersed in aqueous solution at very low ionic strength, vesicles composed of traditional bilayers are obtained.8,9 On the other hand, interdigitated and closed monolayers of DODAAc were shown to be composed of DODAAc dispersions prepared at low ionic strength.9 Because the vesicle structure is so different in the case of DODAAc, it is difficult to predict the type of coverage on oppositely charged latexes. Full interdigitation could possibly be relieved by formation of ion pairs at the sulfate latex interface so that a more traditional bilayer structure would result as coverage. Figure 4A shows the adsorption isotherm of DODAAc from SV onto sulfate polystyrene latex with 76 nm of mean diameter
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J. Phys. Chem., Vol. 100, No. 17, 1996 7133
TABLE 2: Critical Coagulation Concentration (CCC) and Slope of Stability Curves for Bilayer-Covered Polystyrene Microspheres and Small (SV) and Large Vesicles (LV) vesicle type
amphiphile concentration (mM)
particle diameter (nm)
Np (part/mL)
Av/Ap
DODAAc SV DODACl SV DODABr SV DODAAc SV DODACl SV DODABr SV DODAAc SV DODACl SV DODABr SV DODACl LV DODABr LV
0.240 0.315 0.197 0.006 0.136 0.011 0.200 0.200 0.200 0.200 0.200
76 76 76 249 249 249
1.31 × 1012 1.31 × 1012 1.31 × 1012 0.7 × 1010 0.7 × 1010 0.7 × 1010
4.0 4.4 2.8 2.4 47.1 3.9
CCC (M)
d log W/d log C
0.288 0.200 0.078 0.331 0.219 0.162 0.091 0.091 0.063 0.117 0.051
-2.25 -2.69 -3.18 -1.25 -1.13 -1.06 -1.1 -1.1 -2.4 -2.1 -1.2
TABLE 3: Reversibility of NaCl-Induced Aggregation for DODAB-Covered Polystyrene Microspheresa particle diameter (nm)
CNaCl (mM)
A1
A2
76
0 23 33 65 84 95 110 140 165 0 33 48 65 95 165 190 270 310
1.159 1.286 1.302 1.443 1.857 1.839 1.872 2.034 2.062 0.724 0.807 0.853 0.853 0.906 0.946 0.942 0.969 0.988
1.173 1.280 1.293 1.465 1.801 1.808 1.884 1.998 2.000 0.676 0.629 0.698 0.673 0.722 0.815 0.816 0.853 0.869
249
R 0.17 0.17 0 0.04 0.03 0.01 0.05 0.04 1.00 0.82 1.00 0.73 0.33 0.31 0.23 0.22
a Final particle number densities are 1.31 × 1012 and 7.0 × 109 particles/mL, for small and large covered particles, respectively. Final DODAB concentration are 0.197 and 0.0113 mM with Av/Ap ) 2.8 and 3.9, for small and large covered particles, respectively. Notice that controls A1c and A2c are A1 and A2, respectively, at CNaCl ) 0.
in pure water or in a 1 mM NaAc water solution. Both isotherms are of high affinity and present an adsorption maximum tipical of competitive adsorption. The adsorption maximum at 1 mM NaAc is higher than that in water. Adsorption attains a plateau value at high DODAAc concentration in the supernatant: 40 × 1017 DODAAc molecules/m2 of polystyrene for both cases (Figure 4A). As the DODAAc concentration in the supernatant increases, interdigitated and free DODAAc vesicles start to compete with covered latexes for adhering DODAAc vesicles attached to the latex. At 1 mM NaAc, the polarity of the intervening medium is higher than in water. The ionic strength being higher, electrostatic attraction between vesicles and particles would be smaller, and hydrophobic attraction between vesicles and latex would be larger than in pure water. If the hydrophobic attraction between vesicles and particles were absent and the adsorption process at its early stages and at low amphiphile concentration were purely electrostatically controlled, adsorption would be smaller at 1 mM NaAc because of the screening effect. Thus, the higher adsorption maximum at 1 mM NaAc indicates the dominance of the hydrophobic over the electrostatic attraction for the interaction between DODAAc SV and sulfate latex. Possibly, at the adsorption maximum, DODAAc coverage on the latex consists of one interdigitated monolayer that is attaching interdigitated vesicles. Further increasing DODAAc concentration in the supernatant would lead to a decrease in the number of attached interdigitated vesicles on the covered particles that would become attached to free vesicles. Vesicles
Figure 2. Turbidity at 400 nm as a function of time for DODAAc LV prepared in D-glucose 0.6 M (A) or in 0.6 M D-glucose and 1 mM NaAc (B) after NaAc addition to final concentrations in the cuvette equal to 0.02, 0.05, 0.08, 0.11, and 0.155 M (from bottom to top). The final DODAAc concentration is 0.2 mM.
Figure 3. Reversibility (R) of NaCl-induced aggregation between bilayer-covered microspheres as a function of NaCl concentration. DODABr SV were used to obtain coverage on the microspheres. Microspheres were previously covered with DODABr. Experimental conditions are given in Table 3.
excess at the covered microsphere surface would be removed, leaving ultimately a bilayer coverage on the latex. In fact, the curves in Figure 4A tend ultimately to a plateau value for adsorption that is consistent with bilayer coverage at the highest DODAAc concentrations in the supernatant. Adsorption at bilayer coverage should be (36-40) × 1017 DODAAc molecules/
7134 J. Phys. Chem., Vol. 100, No. 17, 1996
Tsuruta and Carmona-Ribeiro vesicle surface but cannot bind at the inner vesicle surface because salt does not penetrate into the vesicle interior. With acetate as the counterion at pH 5.1-5.3, specific acetate binding at the inner vesicle surface would be possible due to permeation of the neutral acetic acid through the vesicle membrane. Thereby, DODAAc large vesicles achieve a very high colloid stability at pH 5.1-5.3. This stability is the highest hitherto observed for DODA vesicles and is even higher than colloid stability for DODA-covered polystyrene microspheres. As counterion size and hydration increase (from bromide to acetate) so does colloid stability of vesicles and covered microspheres. The effect of increasing size is to decrease colloid stability due to reversible aggregation at a secondary minimum. DODAAc adsorption from vesicles onto oppositely charged latexes takes place via a competitive mechanism between vesicle/vesicle and vesicle/monolayer-covered latex that ends up with bilayer coverage at the highest free amphiphile concentrations in the supernatant. Perspectives
Figure 4. DODAAC adsorption from DODAAc SV onto sulfate polystyrene microspheres with 76 nm of mean diameter at 25 °C, in water or in 1 mM NaAc. Particle number density is 1012 particles/cm3. In A, adsorption as a function of the free DODAAc concentration remaining in the supernatant. In B, adsorption as a function of the proportion between total surface areas for vesicles and particles, Av/ Ap.
m2 of polystyrene, i.e., 0.56-0.50 nm2/two DODAAc molecules adsorbed on the latex. These figures are consistent with 40 × 1017 DODAAc molecules/m2 of polystyrene from the isotherms at the highest DODAAc concentrations in the supernatant (Figure 4A). If free DODAAc vesicles in pure water remove the excess of vesicles attached to the latex, adhesion between vesicles must be occurring in pure water despite the repulsive electrostatic and hydration interactions between them. To gather some evidence for this, we used photon correlation spectroscopy to determine the vesicle size distribution for DODAAc LV in pure water. Two peaks for the intensity of light scattered were detected: one at 108.7 nm of mean Dz that possibly corresponds to separate vesicles, and the other at 257.7 nm that is close to double the former (not shown). Intensities of light scattered for each peak amounted to about 50% of the total intensity scattered. Also video-enhanced microscopy for shorter chained derivatives with acetate as counterion showed aggregated vesicles in pure water.22 Therefore, DODAAc vesicle adhesion in pure water seems to be a real phenomenon caused by the interdigitated nature of these vesicles. One of the reviewers pointed out to the speculative nature of an interpretation based on interdigitation as the structure for the DODAAc bilayer vesicle. In fact, we are presently employing spectroscopic methods to offer stronger and more direct evidences for the interdigitation hypothesis and to look at counterion effects on the bilayer structure. Conclusions DODABr, DODACl, and DODAAc bilayer-covered microspheres are more stable than vesicles of similar size under identical medium composition. Vesicle instability can be ascribed to the asymmetry of charge distribution that occurs when salt is added to the outside. Counterions bind at the outer
The polystyrene particles are colloidal in nature, and their Hamaker constants can be estimated with some assurance. Estimates of surface potential for bilayers and covered particles with DODA acetate, chloride, and bromide are presently being obtained in our laboratory over a range of particle sizes. In the near future we expect to combine the information obtained from W with the DLVO theory. This will shed new light on the intricate relationship between particle size, surface potential, and colloid stability in a possibly ideal colloidal system. Acknowledgment. Grants 93/2288-6 and 510022/93-6 from FAPESP and CNPq, respectively, are gratefully acknowledged. L.R.T. thanks FAPESP for an undergraduate fellowship (94/ 1405-1). References and Notes (1) Carmona-Ribeiro, A. M. Chem. Soc. ReV. 1992, 21, 207. (2) Carmona-Ribeiro, A. M.; Yoshida, L. S.; Chaimovich, H. J. Phys. Chem. 1985, 89, 2928. (3) Carmona-Ribeiro, A. M. J. Phys. Chem. 1989, 93, 2630. (4) Carmona-Ribeiro, A. M.; Midmore, B. R. Langmuir 1992, 8, 801. (5) Tsuruta, L. R.; Lessa, M. M.; Carmona-Ribeiro, A. M. J. Colloid Interface Sci., in press. (6) Ta´pias, G. N.; Sicchierolli, S. M.; Mamizuka, E. M.; CarmonaRibeiro, A. M. Langmuir 1994, 10, 3461. (7) Sicchierolli, S. M.; Mamizuka, E. M.; Carmona-Ribeiro, A. M. Langmuir 1995, 11, 2991. (8) Tsuruta, L. R.; Lessa, M. M.; Carmona-Ribeiro, A. M. Langmuir 1995, 11, 2938. (9) Nascimento, D. B.; Lessa, M. M.; Carmona-Ribeiro, A. M., unpublished results. (10) Marmur, A. J. Colloid Interface Sci. 1979, 72, 41. (11) Pashley, R. M.; MacGuiggan, P.; Ninham, B. R.; Brady, J.; Evans, D. F. J. Phys. Chem. 1986, 90, 1637. (12) Tsao, Y.; Evans, D. F.; Rand, R. P.; Parsegian, V. A. Langmuir 1993, 9, 233. (13) Carmona-Ribeiro, A. M.; Chaimovich, H. Biochim. Biophys. Acta 1983, 733, 172. (14) Fendler, J. H. Acc. Chem. Res. 1980, 13, 7. (15) Stelmo, M.; Chaimovich, H.; Cuccovia, I. M. J. Colloid Interface Sci. 1987, 117, 200. (16) Schales, O.; Schales, S. S. J. Biol. Chem. 1941, 140, 879. (17) Reerink, H.; Overbeek, J. Th. G. Discuss. Faraday Soc. 1954, 18, 95. (18) Marra, J. J. Phys. Chem. 1986, 90, 2145. (19) Carmona-Ribeiro, A. M. J. Phys. Chem. 1993, 97, 11843. (20) Carmona-Ribeiro, A. M. J. Phys. Chem. 1993, 97, 4247. (21) Huang, C. In Liposome Letters; Bangham, A. D., Ed.; Academic Press: London, 1983; pp 37-47. (22) Brady, J. E.; Evans, D. F.; Kachar, B.; Ninham, B. W. J. Am. Chem. Soc. 1984, 106, 4279.
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