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Interactions between bilayer membranes and latex
Colloids and Surfaces A: Physiochemical and Engineering Aspects 153 Ž1999. 355]361
Interactions between bilayer membranes and latex Ana Maria Carmona-RibeiroU , Marina de Moraes Lessa Departamento de Bioquımica, Instituto de Quımica, Uni¨ ersidade de Sao ´ ´ ˜ Paulo, CP 26077, 05599-970 Sao ˜ Paulo SP, Brazil
Abstract The interaction between sulfate polystyrene latex and bilayer vesicles is described from the point of view of membrane-induced latex deflocculation and theoretical colloid stability for bilayer-covered particles from a DLVO model without free parameters. Unilamellar cationic or anionic vesicles were prepared from dioctadecyldimethylammonium bromide ŽDODAB. or asolecithin ŽASO., respectively, and allowed to interact with sulfate polystyrene microspheres of several sizes. Mean zeta-average diameters for latex particles in the interacting mixtures decreases as a function of interaction time between sulfate polystyrene particles and asolecithin vesicles attaining a final size compatible with a negatively charged asolecithin monolayer deposited onto the anionic latex surface. In contrast to the ASO behaviour, the cationic DODAB was previously reported to deposit as a bilayer onto the sulfate latex. The occurrence of an ordered amphiphile assembly deposited onto the latex is highly dependent on the proportion of total surface areas for vesicles and particles. From zeta-potential measurements and a DLVO model, theoretical colloid stabilities are calculated as a function of monovalent salt concentration or particle size and found to be much higher than experimentally measured stabilities. DLVO expectations are not fulfilled in spite of the presumably ideal nature of the bilayer-covered polystyrene microsphere as a colloid. Q 1999 Elsevier Science B.V. All rights reserved. Keywords: Interactions; Bilayer-vesicles; Latex; Colloid stability; Bilayer-covered particles
1. Introduction The self-assembly of bilayer-forming amphiphiles on latex is becoming increasingly impor-
U
Corresponding author. Fax: q55 11 8155579. E-mail address: [email protected] ŽA.M. CarmonaRibeiro.
tant in several areas of research that range from understanding the basic interactive process between a simple bilayer-membrane and a solid surface w1]6x to designing immunological kits and biosensors for amplification of biomolecular recognition w7,8x. A corollary of these developments is the preparation of the ‘ideal colloid’ Žhomodisperse polystyrene microspheres covered with an evenly charged bilayer membrane. useful
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A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
for evaluation of current theories that predict colloid stability w9]12x. Bilayer-forming amphiphiles assemble on latex depending on amphiphile type and concentration and on functional groups on latex w1]3x. Neutral phospholipids have a specially high affinity for amidine polystyrene microspheres basically depositing as a phospholipid monolayer with the polar heads uppermost w2,6]8x whereas the electrostatic attraction between cationic vesicles and oppositely charged microspheres may lead to deposition on latex of the bilayer membrane as a whole w1,3]5x. Asolecithin is a mixture of soybean phospholipids that produces anionic bilayer vesicles w13x. In this work, we further advance our understanding of the interaction between bilayer vesicles and latex by describing latex deflocculation induced by vesicles due to deposition of an adsorbed anionic monolayer of asolecithin lipids on latex with their polarheads uppermost. Furthermore, the bilayer-covered assembly for dioctadecyldimethylammonium bromide ŽDODAB. bilayers on sulfate latex is used to evaluate the suitability of the DLVO theory for explaining the colloid stability of bilayer-covered polystyrene microspheres. DLVO calculations show that experimental colloidal stability w4x is much lower than the theoretical colloid stability calculated from zeta-potentials and DLVO theory over a range of particle sizes and monovalent salt concentrations. The theory definitely does not explain the low experimental stabilities of the bilayer-covered latex. 2. Materials and methods ASO from soybean Žtype IV-S. and DODAB of the highest purity available were from Sigma. Phospholipid concentration was determined by inorganic phosphorus ŽPi . analysis w14x. DODAB concentration was determined by solubilization of a dye-amphiphile complex in non-ionic micelles w15x or by microtitration w16x. Charged polystyrene microspheres described as ultraclean by the supplier were obtained from Interfacial Dynamics Co. ŽPortland, OR. and used as supplied. Properties of the microspheres are given in Table 1. Mean diameters were obtained
by the supplier using an electron microscope. All other reagents were analytical grade and used without further purification. Water was MILLI-Q quality. ASO or DODAB small vesicles were prepared in pure water by sonication with a tip as previously described in references w17x and w18x, respectively. Polystyrene microspheres were always diluted in pure water. Interaction between vesicles and microspheres was induced by adding the vesicles to the polystyrene. The final number density of microspheres Ž Np . was different for each particle size and is indicated for each kind of experiment. Mixtures of DODABrmicrospheres were thermostated at 258C for 1 h before measurements were carried out. Total surface area on polystyrene Ž A p . was calculated from the particle number density and the mean particle radius given by the supplier. Total surface area on DODAB vesicles Ž A v . was calculated from the area per monomer for DODAB at an air-water interface, i.e. 0.57 nm2 w19x, the amphiphile concentration in the vesiclerparticle mixture, the final volume of the mixture, and the traditional closed bilayer model for the vesicle, which assumes that there is one monomer in the outer monolayer per each monomer in the inner monolayer of the bilayer. Particle or vesicle sizes were measured using a Malvern 4700c PCS apparatus employing a Coherent Innova 90 laser. The size quoted is the mean harmonic z-average diameter Ž Dz . of at least 15 independent measurements at 258C. Zeta-potentials were determined at 258C using a Zetasizer IIc Particle Electrophoresis and Submicron Particle Analyser. The light source was a 5 mW He]Ne laser. Experimental error was "0.5]" 1.0 mV. Previously to determining zetapotential as a function of NaCl concentration for DODAB-covered particles, zeta-potential for the covered particles in water were measured as a control. The values obtained were those previously reported in ref. w4x. The proportion between total surface areas for vesicles and particles used for bilayer-covering the latex was equal to 1.0 w4x, independently of particle size. Zeta-potentials as a function of NaCl concentration were determined 1 h after adding NaCl to the bilayer-
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
covered particles to final NaCl concentrations varying from 10y7 up to 10y2 M NaCl. The total potential energy Ž V . of interaction as a function of the separation distance Ž u. was calculated from the bilayer-covered particle radius, the Stern potential Žextrapolated from the zeta-potential against NaCl concentration curve that was determined experimentally for the bilayer-covered microspheres as described above., NaCl concentration and Hamaker constants for polystyrene, water and a hydrocarbon layer taken as 7.8= 10y2 0 , 3.7= 10y2 0 and 5.8= 10y2 0 J, respectively w20,21x. A DLVO model for vesicles w9,10x, without free parameters, was used to calculate colloid stabilities for bilayer-covered microspheres. The Ž VrkT, u. curves were tabulated and plotted with k units in metres, zeta-potentials in volts and V units as joules. The dimensionless dielectric constant of water at 258C was taken as 78.54. The stability ratio ŽW . was obtained by numerical integration of eq. Ž8. in ref. w9x. In all computations of V and W only aqueous solutions of 1:1 electrolyte at 258C were considered. The experimental stability values ŽWe . were calculated from data for bilayer-covered particles w4,5x using the DLVO algorithm w9x adapted for polystyrene microspheres covered with a hydrocarbon layer of one bilayer thickness Ž5 nm. interacting across water. 3. Results and discussion 3.1. The interaction between bilayer ¨ esicles and polystyrene microspheres as a function of bilayer and latex type Previous work on the interaction vesiclerlatex has shown that the interaction allows many possibilities. Major features of the interaction are described in Fig. 1. Electrostatic andror van der Waals attraction andror hydrophobic attraction leads to vesicle aggregation to the latex Žstep 1.. These interaction forces may disrupt the vesicle bilayer and promote bilayer adsorption onto the microsphere Žstep 2. andror promote further microsphere attachment Ž29.. The adsorbed bilayer may attract another microsphere Žstep 3.. If neutral lipids in the bilayer vesicle are in the more
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Fig. 1. The interaction between one bilayer vesicle and two polystyrene microspheres considering vesicles and microspheres of similar sizes. Electrostatic andror van der Waals andror hydrophobic attraction leads to aggregation of a vesicle and a polystyrene microsphere Ž1.. These same interaction forces may disrupt the vesicle bilayer and promote bilayer adsorption onto the microsphere Ž2. andror further aggregation with the other microsphere Ž29.. The adsorbed bilayer may attract the second microsphere Ž3.. The hydrophobic interaction between the polystyrene surface and the hydrocarbon chains in the bilayer may completely destroy the bilayer structure flip-flopping the hydrocarbon chains onto the polystyrene surface and generating a monolayer coverage on each microsphere Ž4..
fluid liquid-crystalline state, the hydrophobic attraction between latex and hydrocarbon chains in the lipid bilayer completely destroys the bilayer structure. The hydrocarbon chains on the polystyrene surface generate a monolayer coverage with polarheads uppermost Žstep 4.. If charged lipids in the bilayer vesicle are oppositely charged relative to the latex, at suitable ratios for total surface areas for vesicles and latex, a stable dispersion composed of bilayer-covered latex may
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A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
result. Experimental data available from the literature w1]8x validate the picture shown as Fig. 1. 3.2. The deflocculating effect of bilayer-¨ esicles on aggregated polystyrene microspheres Bilayer vesicles may cause latex deflocculation ŽFig. 2.. The effect of addition of anionic vesicles to anionic sulfate polystyrene Ž0.303 mm mean diameter obtained from electron microscopy. is a decrease of mean zeta-average diameter of the particles as a function of time ŽFig. 2.. Negatively charged asolecithin vesicles do interact with sulfate polystyrene particles of the same charge sign to yield a final particle size after 72 h of interaction time equal to 0.308 mm. This figure is consistent with deposition of a 2.5 nm asolecithin layer on latex, i.e. deposition of an asolecithin monolayer with polarheads uppermost. This result suggests that the hydrophobic attraction overwhelms the electrostatic repulsion between vesicle and particle latex and stabilizes the asolecithin monolayer assembly on the latex particle. At a ratio for total surface areas for vesicles and particles equal to 0.5, a similar though more rapid stabilizing effect of neutral lipids on amidine latex at monolayer coverage was previously described w1,2x. 3.3. The effects of NaCl concentration and particle radius on zeta-potentials and colloid stability of bilayer-co¨ ered polystyrene microspheres The zeta-potential monotonically decreases as a function of the logarithm of NaCl concentration for the five covered-particle sizes tested Žsee Table 1, particles I, IV, V, VI, VIII., though only results for latex V and VIII are shown in Fig. 3. Zetapotential dependence on ionic strength over a range of NaCl concentrations was obtained over a range of concentrations where flocculation is absent Ž10y7 ]10y2 M NaCl.. This allowed extrapolation of surface potentials over the region of higher NaCl concentrations where flocculation does occur w4x. From the estimated surface potential, the potential energy of interaction as a function of the separation distance between bilayercovered particles was calculated using the DLVO model ŽFig. 5..
Fig. 2. Mean zeta-average diameter for latex XIII as a function of interaction time with asolecithin small vesicles Ž112.1 mean zeta-average diameter and 0.206 of polydispersity index.. Final number densities for vesicles, Nv , is 7.9 = 10 11 vesiclesrml and for particles, Np , is 3.1= 10 10 particlesrml so that NvrNp is 22.6 whereas the ratio for the total surface areas is A vrA p s 3.7.
The effect of size on the zeta-potential of bilayer-covered particles is shown in Fig. 4. In water, the zeta-potential displays a weak dependence on size ŽFig. 4.. However, in NaCl solution, there is a sigmoidal increase of the zeta-potential as a function of particle radius at each of the three different NaCl concentrations tested. Larger bilayer-covered particles present higher surface potentials ŽFig. 4. but are, surprisingly, less stable in a colloidal sense than are the smaller ones Žref. w4x and Fig. 6.. At this stage, the importance of aggregation at the secondary minimum as a factor Table 1 Properties of sulfate polystyrene microspheres in water at 258C Latex
Mean diameter Žnm.
Area per charge group Žnm2 .
Specific surface area Žcm2 gy1 .
I IV V VI VIII XIII
76 137 249 301 412 303
7.1 20.1 7.7 9.5 4.3 1.1
748 316 415 124 228 401 188 944 138 039 187 696
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
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Fig. 4. Zeta-potential as a function of the bilayer-coered particle radius at four different NaCl concentrations.
Fig. 3. Zeta-potential for bilayer-covered latex V ŽA. and VIII ŽB. as a function of the logarithm of the monovalent salt concentration. Zeta-potentials at NaCl concentrations larger than 0.01 M were obtained by extrapolation allowing DLVO calculations of theoretical curves in Fig. 4.
that destabilizes the colloidal dispersion has to be evaluated. Table 2 presents depths of secondary minima Žtaken from DLVO calculations such as those in Fig. 5. and critical coagulation concentrations Žtaken from ref. w4x. as a function of size for covered particles. The depth of the secondary minimum indeed increases as a function of the covered-particle radius. Thus, a plausible hypothesis for explaining the low colloidal stability of the larger covered-particles would be aggregation at the secondary minimum. In fact, by taking the secondary minimum into account, Marmur
w22x calculated a theoretical dependence of colloid stability on particle size at a given NaCl concentration that displays a maximum as a function of particle size in very good qualitative agreement with the dependence of colloid stability on particle size previously measured for the bilayercovered particles w4x. Combining flocculation in the primary and secondary minimum in a kinetical model, Marmur has found a critical dependence of stability on the kinetical energy of the interacting particles. If this energy is smaller than the depth of the secondary minimum, aggregation in the secondary minimum is expected to occur. The model is rather sensitive to the existence of the secondary minimum. For example, a mini-
Table 2 Depth of the secondary minimum Ž f min rkT . and critical coagulation concentration ŽCCC. as a function of size for the DODAB bilayer-covered sulfate latex. DLVO calculations were performed for the covered particles at the CCC Radius for the covered particle Žnm.
CCC ŽM.
fmin rkT
43.8 73.5 130.0 156.0 211.0
0.12 0.26 0.28 0.12 0.06
1.7 2.5 4.6 4.5 6.0
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Fig. 5. Interaction energy as a function of interparticle separation distance for bilayer-covered latex. For latex V, the effect of increasing NaCl concentration on the curves is in ŽA.. At 0.12 M NaCl, the effect of size on the interaction energy profile is in ŽB..
mum of 0.5 kT has a small effect on W equal to 30 but a minimum of 0.75 kT decreases W to 25. In Table 2, a considerable reduction in colloid stability is expected as a result of the calculated depths of the secondary minimum, all well above 0.75 kT and therefore, expected to decrease W considerably. We are presently working on calculations using the Marmur model in order to establish quantitatively the role of the secondary minimum on theoretical and experimental stability ratios for bilayer-covered latex. 4. Conclusions A DLVO model without free parameters does not account for the experimental colloid stability of a presumably ideal colloid such as latex cov-
Fig. 6. Theoretical ŽB. and experimental Žv. stabilitites ŽW . of bilayer-covered sulfate latex particles as a function of the logarithm of the NaCl concentration. Data for the experimental stabilities W were taken from ref. w4x. Theoretical W values were computed from a DLVO model for the bilayer-covered particles as described in Section 2.
ered with one DODAB bilayer. The experimental stability measured is much smaller than the theoretical stability ratios. Two possibilities are left for further investigation: either aggregation at a secondary minimum accounts for the much smaller experimental colloid stability or there is an additional attractive force acting between the
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
bilayer-covered particles not considered in the framework of the DLVO theory. Zeta-potential increases whereas experimentally measured colloidal stability displays a maximum as a function of particle size. Therefore, over the range of larger particle sizes, upon increasing zeta-potential, there is a decrease in colloid stability. A possible explanation, justified from calculations of depths for the secondary minimum over a range of sizes, is aggregation at the secondary minimum. Further calculations using the Marmur model will shed new light on this issue. Acknowledgements Grants 93r2288-6; 96r0704-0 from FAPESP and 510022r93-6; 520186r96-6 from CNPq are gratefully acknowledged. References w1x A.M. Carmona-Ribeiro, B.R. Midmore, Langmuir 8 Ž1992. 801. w2x A.M. Carmona-Ribeiro, T.M. Herrington, J. Colloid Interface Sci. 156 Ž1993. 19. w3x L.R. Tsuruta, M.M. Lessa, A.M. Carmona-Ribeiro, Langmuir 11 Ž1995. 2938.
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w4x L.R. Tsuruta, M.M. Lessa, A.M. Carmona-Ribeiro, J. Colloid Interface Sci. 175 Ž1995. 470. w5x L.R. Tsuruta, A.M. Carmona-Ribeiro, J. Phys. Chem. 100 Ž1996. 7130. w6x S.M. Sicchierolli, A.M. Carmona-Ribeiro, Colloids Surf. B Biointerfaces 5 Ž1995. 57. w7x M.M. Lessa, A.M. Carmona-Ribeiro, J. Colloid Interface Sci. 182 Ž1996. 166. w8x S.M. Sicchierolli, A.M. Carmona-Ribeiro, J. Phys. Chem. 100 Ž1996. 16771. w9x A.M. Carmona-Ribeiro, J. Phys. Chem. 93 Ž1989. 2630. w10x A.M. Carmona-Ribeiro, J. Phys. Chem. 96 Ž1992. 9555. w11x A.M. Carmona-Ribeiro, J. Phys. Chem. 97 Ž1993. 4247. w12x A.M. Carmona-Ribeiro, J. Phys. Chem. 97 Ž1993. 11843. w13x Y. Kagawa, E. Racker, J. Biol. Chem. 241 Ž1966. 2467. w14x G. Houser, S. Fleischer, A. Yamamoto, Lipids 5 Ž1970. 594. w15x M. Stelmo, H. Chaimovich, I.M. Cuccovia, J. Colloid Interface Sci. 117 Ž1987. 200. w16x O. Schales, S.S. Schales, J. Biol. Chem. 140 Ž1941. 879. w17x C.H. Huang, Biochemistry 8 Ž1969. 344. w18x C.D. Tran, P.L. Klahn, A. Romero, J.H. Fendler, J. Am. Chem. Soc. 100 Ž1978. 1622. w19x P.M. Claesson, A.M. Carmona-Ribeiro, K. Kurihara, J. Phys. Chem. 93 Ž1989. 917. w20x R.H. Ottewill, T. Walker, J. Chem. Soc. Faraday Trans. I 70 Ž1974. 917. w21x B. Vincent, J. Colloid Interface Sci. 175 Ž1973. 270. w22x A. Marmur, J. Colloid Interface Sci. 72 Ž1979. 41.
Interactions between bilayer membranes and latex Ana Maria Carmona-RibeiroU , Marina de Moraes Lessa Departamento de Bioquımica, Instituto de Quımica, Uni¨ ersidade de Sao ´ ´ ˜ Paulo, CP 26077, 05599-970 Sao ˜ Paulo SP, Brazil
Abstract The interaction between sulfate polystyrene latex and bilayer vesicles is described from the point of view of membrane-induced latex deflocculation and theoretical colloid stability for bilayer-covered particles from a DLVO model without free parameters. Unilamellar cationic or anionic vesicles were prepared from dioctadecyldimethylammonium bromide ŽDODAB. or asolecithin ŽASO., respectively, and allowed to interact with sulfate polystyrene microspheres of several sizes. Mean zeta-average diameters for latex particles in the interacting mixtures decreases as a function of interaction time between sulfate polystyrene particles and asolecithin vesicles attaining a final size compatible with a negatively charged asolecithin monolayer deposited onto the anionic latex surface. In contrast to the ASO behaviour, the cationic DODAB was previously reported to deposit as a bilayer onto the sulfate latex. The occurrence of an ordered amphiphile assembly deposited onto the latex is highly dependent on the proportion of total surface areas for vesicles and particles. From zeta-potential measurements and a DLVO model, theoretical colloid stabilities are calculated as a function of monovalent salt concentration or particle size and found to be much higher than experimentally measured stabilities. DLVO expectations are not fulfilled in spite of the presumably ideal nature of the bilayer-covered polystyrene microsphere as a colloid. Q 1999 Elsevier Science B.V. All rights reserved. Keywords: Interactions; Bilayer-vesicles; Latex; Colloid stability; Bilayer-covered particles
1. Introduction The self-assembly of bilayer-forming amphiphiles on latex is becoming increasingly impor-
U
Corresponding author. Fax: q55 11 8155579. E-mail address: [email protected] ŽA.M. CarmonaRibeiro.
tant in several areas of research that range from understanding the basic interactive process between a simple bilayer-membrane and a solid surface w1]6x to designing immunological kits and biosensors for amplification of biomolecular recognition w7,8x. A corollary of these developments is the preparation of the ‘ideal colloid’ Žhomodisperse polystyrene microspheres covered with an evenly charged bilayer membrane. useful
0927-7757r99r$ - see front matter Q 1999 Elsevier Science B.V. All rights reserved. PII: S 0 9 2 7 - 7 7 5 7 Ž 9 8 . 0 0 5 3 2 - 9
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A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
for evaluation of current theories that predict colloid stability w9]12x. Bilayer-forming amphiphiles assemble on latex depending on amphiphile type and concentration and on functional groups on latex w1]3x. Neutral phospholipids have a specially high affinity for amidine polystyrene microspheres basically depositing as a phospholipid monolayer with the polar heads uppermost w2,6]8x whereas the electrostatic attraction between cationic vesicles and oppositely charged microspheres may lead to deposition on latex of the bilayer membrane as a whole w1,3]5x. Asolecithin is a mixture of soybean phospholipids that produces anionic bilayer vesicles w13x. In this work, we further advance our understanding of the interaction between bilayer vesicles and latex by describing latex deflocculation induced by vesicles due to deposition of an adsorbed anionic monolayer of asolecithin lipids on latex with their polarheads uppermost. Furthermore, the bilayer-covered assembly for dioctadecyldimethylammonium bromide ŽDODAB. bilayers on sulfate latex is used to evaluate the suitability of the DLVO theory for explaining the colloid stability of bilayer-covered polystyrene microspheres. DLVO calculations show that experimental colloidal stability w4x is much lower than the theoretical colloid stability calculated from zeta-potentials and DLVO theory over a range of particle sizes and monovalent salt concentrations. The theory definitely does not explain the low experimental stabilities of the bilayer-covered latex. 2. Materials and methods ASO from soybean Žtype IV-S. and DODAB of the highest purity available were from Sigma. Phospholipid concentration was determined by inorganic phosphorus ŽPi . analysis w14x. DODAB concentration was determined by solubilization of a dye-amphiphile complex in non-ionic micelles w15x or by microtitration w16x. Charged polystyrene microspheres described as ultraclean by the supplier were obtained from Interfacial Dynamics Co. ŽPortland, OR. and used as supplied. Properties of the microspheres are given in Table 1. Mean diameters were obtained
by the supplier using an electron microscope. All other reagents were analytical grade and used without further purification. Water was MILLI-Q quality. ASO or DODAB small vesicles were prepared in pure water by sonication with a tip as previously described in references w17x and w18x, respectively. Polystyrene microspheres were always diluted in pure water. Interaction between vesicles and microspheres was induced by adding the vesicles to the polystyrene. The final number density of microspheres Ž Np . was different for each particle size and is indicated for each kind of experiment. Mixtures of DODABrmicrospheres were thermostated at 258C for 1 h before measurements were carried out. Total surface area on polystyrene Ž A p . was calculated from the particle number density and the mean particle radius given by the supplier. Total surface area on DODAB vesicles Ž A v . was calculated from the area per monomer for DODAB at an air-water interface, i.e. 0.57 nm2 w19x, the amphiphile concentration in the vesiclerparticle mixture, the final volume of the mixture, and the traditional closed bilayer model for the vesicle, which assumes that there is one monomer in the outer monolayer per each monomer in the inner monolayer of the bilayer. Particle or vesicle sizes were measured using a Malvern 4700c PCS apparatus employing a Coherent Innova 90 laser. The size quoted is the mean harmonic z-average diameter Ž Dz . of at least 15 independent measurements at 258C. Zeta-potentials were determined at 258C using a Zetasizer IIc Particle Electrophoresis and Submicron Particle Analyser. The light source was a 5 mW He]Ne laser. Experimental error was "0.5]" 1.0 mV. Previously to determining zetapotential as a function of NaCl concentration for DODAB-covered particles, zeta-potential for the covered particles in water were measured as a control. The values obtained were those previously reported in ref. w4x. The proportion between total surface areas for vesicles and particles used for bilayer-covering the latex was equal to 1.0 w4x, independently of particle size. Zeta-potentials as a function of NaCl concentration were determined 1 h after adding NaCl to the bilayer-
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
covered particles to final NaCl concentrations varying from 10y7 up to 10y2 M NaCl. The total potential energy Ž V . of interaction as a function of the separation distance Ž u. was calculated from the bilayer-covered particle radius, the Stern potential Žextrapolated from the zeta-potential against NaCl concentration curve that was determined experimentally for the bilayer-covered microspheres as described above., NaCl concentration and Hamaker constants for polystyrene, water and a hydrocarbon layer taken as 7.8= 10y2 0 , 3.7= 10y2 0 and 5.8= 10y2 0 J, respectively w20,21x. A DLVO model for vesicles w9,10x, without free parameters, was used to calculate colloid stabilities for bilayer-covered microspheres. The Ž VrkT, u. curves were tabulated and plotted with k units in metres, zeta-potentials in volts and V units as joules. The dimensionless dielectric constant of water at 258C was taken as 78.54. The stability ratio ŽW . was obtained by numerical integration of eq. Ž8. in ref. w9x. In all computations of V and W only aqueous solutions of 1:1 electrolyte at 258C were considered. The experimental stability values ŽWe . were calculated from data for bilayer-covered particles w4,5x using the DLVO algorithm w9x adapted for polystyrene microspheres covered with a hydrocarbon layer of one bilayer thickness Ž5 nm. interacting across water. 3. Results and discussion 3.1. The interaction between bilayer ¨ esicles and polystyrene microspheres as a function of bilayer and latex type Previous work on the interaction vesiclerlatex has shown that the interaction allows many possibilities. Major features of the interaction are described in Fig. 1. Electrostatic andror van der Waals attraction andror hydrophobic attraction leads to vesicle aggregation to the latex Žstep 1.. These interaction forces may disrupt the vesicle bilayer and promote bilayer adsorption onto the microsphere Žstep 2. andror promote further microsphere attachment Ž29.. The adsorbed bilayer may attract another microsphere Žstep 3.. If neutral lipids in the bilayer vesicle are in the more
357
Fig. 1. The interaction between one bilayer vesicle and two polystyrene microspheres considering vesicles and microspheres of similar sizes. Electrostatic andror van der Waals andror hydrophobic attraction leads to aggregation of a vesicle and a polystyrene microsphere Ž1.. These same interaction forces may disrupt the vesicle bilayer and promote bilayer adsorption onto the microsphere Ž2. andror further aggregation with the other microsphere Ž29.. The adsorbed bilayer may attract the second microsphere Ž3.. The hydrophobic interaction between the polystyrene surface and the hydrocarbon chains in the bilayer may completely destroy the bilayer structure flip-flopping the hydrocarbon chains onto the polystyrene surface and generating a monolayer coverage on each microsphere Ž4..
fluid liquid-crystalline state, the hydrophobic attraction between latex and hydrocarbon chains in the lipid bilayer completely destroys the bilayer structure. The hydrocarbon chains on the polystyrene surface generate a monolayer coverage with polarheads uppermost Žstep 4.. If charged lipids in the bilayer vesicle are oppositely charged relative to the latex, at suitable ratios for total surface areas for vesicles and latex, a stable dispersion composed of bilayer-covered latex may
358
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
result. Experimental data available from the literature w1]8x validate the picture shown as Fig. 1. 3.2. The deflocculating effect of bilayer-¨ esicles on aggregated polystyrene microspheres Bilayer vesicles may cause latex deflocculation ŽFig. 2.. The effect of addition of anionic vesicles to anionic sulfate polystyrene Ž0.303 mm mean diameter obtained from electron microscopy. is a decrease of mean zeta-average diameter of the particles as a function of time ŽFig. 2.. Negatively charged asolecithin vesicles do interact with sulfate polystyrene particles of the same charge sign to yield a final particle size after 72 h of interaction time equal to 0.308 mm. This figure is consistent with deposition of a 2.5 nm asolecithin layer on latex, i.e. deposition of an asolecithin monolayer with polarheads uppermost. This result suggests that the hydrophobic attraction overwhelms the electrostatic repulsion between vesicle and particle latex and stabilizes the asolecithin monolayer assembly on the latex particle. At a ratio for total surface areas for vesicles and particles equal to 0.5, a similar though more rapid stabilizing effect of neutral lipids on amidine latex at monolayer coverage was previously described w1,2x. 3.3. The effects of NaCl concentration and particle radius on zeta-potentials and colloid stability of bilayer-co¨ ered polystyrene microspheres The zeta-potential monotonically decreases as a function of the logarithm of NaCl concentration for the five covered-particle sizes tested Žsee Table 1, particles I, IV, V, VI, VIII., though only results for latex V and VIII are shown in Fig. 3. Zetapotential dependence on ionic strength over a range of NaCl concentrations was obtained over a range of concentrations where flocculation is absent Ž10y7 ]10y2 M NaCl.. This allowed extrapolation of surface potentials over the region of higher NaCl concentrations where flocculation does occur w4x. From the estimated surface potential, the potential energy of interaction as a function of the separation distance between bilayercovered particles was calculated using the DLVO model ŽFig. 5..
Fig. 2. Mean zeta-average diameter for latex XIII as a function of interaction time with asolecithin small vesicles Ž112.1 mean zeta-average diameter and 0.206 of polydispersity index.. Final number densities for vesicles, Nv , is 7.9 = 10 11 vesiclesrml and for particles, Np , is 3.1= 10 10 particlesrml so that NvrNp is 22.6 whereas the ratio for the total surface areas is A vrA p s 3.7.
The effect of size on the zeta-potential of bilayer-covered particles is shown in Fig. 4. In water, the zeta-potential displays a weak dependence on size ŽFig. 4.. However, in NaCl solution, there is a sigmoidal increase of the zeta-potential as a function of particle radius at each of the three different NaCl concentrations tested. Larger bilayer-covered particles present higher surface potentials ŽFig. 4. but are, surprisingly, less stable in a colloidal sense than are the smaller ones Žref. w4x and Fig. 6.. At this stage, the importance of aggregation at the secondary minimum as a factor Table 1 Properties of sulfate polystyrene microspheres in water at 258C Latex
Mean diameter Žnm.
Area per charge group Žnm2 .
Specific surface area Žcm2 gy1 .
I IV V VI VIII XIII
76 137 249 301 412 303
7.1 20.1 7.7 9.5 4.3 1.1
748 316 415 124 228 401 188 944 138 039 187 696
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
359
Fig. 4. Zeta-potential as a function of the bilayer-coered particle radius at four different NaCl concentrations.
Fig. 3. Zeta-potential for bilayer-covered latex V ŽA. and VIII ŽB. as a function of the logarithm of the monovalent salt concentration. Zeta-potentials at NaCl concentrations larger than 0.01 M were obtained by extrapolation allowing DLVO calculations of theoretical curves in Fig. 4.
that destabilizes the colloidal dispersion has to be evaluated. Table 2 presents depths of secondary minima Žtaken from DLVO calculations such as those in Fig. 5. and critical coagulation concentrations Žtaken from ref. w4x. as a function of size for covered particles. The depth of the secondary minimum indeed increases as a function of the covered-particle radius. Thus, a plausible hypothesis for explaining the low colloidal stability of the larger covered-particles would be aggregation at the secondary minimum. In fact, by taking the secondary minimum into account, Marmur
w22x calculated a theoretical dependence of colloid stability on particle size at a given NaCl concentration that displays a maximum as a function of particle size in very good qualitative agreement with the dependence of colloid stability on particle size previously measured for the bilayercovered particles w4x. Combining flocculation in the primary and secondary minimum in a kinetical model, Marmur has found a critical dependence of stability on the kinetical energy of the interacting particles. If this energy is smaller than the depth of the secondary minimum, aggregation in the secondary minimum is expected to occur. The model is rather sensitive to the existence of the secondary minimum. For example, a mini-
Table 2 Depth of the secondary minimum Ž f min rkT . and critical coagulation concentration ŽCCC. as a function of size for the DODAB bilayer-covered sulfate latex. DLVO calculations were performed for the covered particles at the CCC Radius for the covered particle Žnm.
CCC ŽM.
fmin rkT
43.8 73.5 130.0 156.0 211.0
0.12 0.26 0.28 0.12 0.06
1.7 2.5 4.6 4.5 6.0
360
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
Fig. 5. Interaction energy as a function of interparticle separation distance for bilayer-covered latex. For latex V, the effect of increasing NaCl concentration on the curves is in ŽA.. At 0.12 M NaCl, the effect of size on the interaction energy profile is in ŽB..
mum of 0.5 kT has a small effect on W equal to 30 but a minimum of 0.75 kT decreases W to 25. In Table 2, a considerable reduction in colloid stability is expected as a result of the calculated depths of the secondary minimum, all well above 0.75 kT and therefore, expected to decrease W considerably. We are presently working on calculations using the Marmur model in order to establish quantitatively the role of the secondary minimum on theoretical and experimental stability ratios for bilayer-covered latex. 4. Conclusions A DLVO model without free parameters does not account for the experimental colloid stability of a presumably ideal colloid such as latex cov-
Fig. 6. Theoretical ŽB. and experimental Žv. stabilitites ŽW . of bilayer-covered sulfate latex particles as a function of the logarithm of the NaCl concentration. Data for the experimental stabilities W were taken from ref. w4x. Theoretical W values were computed from a DLVO model for the bilayer-covered particles as described in Section 2.
ered with one DODAB bilayer. The experimental stability measured is much smaller than the theoretical stability ratios. Two possibilities are left for further investigation: either aggregation at a secondary minimum accounts for the much smaller experimental colloid stability or there is an additional attractive force acting between the
A.M. Carmona-Ribeiro, M. de Moraes Lessa r Colloids Surfaces A: Physiochem. Eng. Aspects 153 (1999) 355]361
bilayer-covered particles not considered in the framework of the DLVO theory. Zeta-potential increases whereas experimentally measured colloidal stability displays a maximum as a function of particle size. Therefore, over the range of larger particle sizes, upon increasing zeta-potential, there is a decrease in colloid stability. A possible explanation, justified from calculations of depths for the secondary minimum over a range of sizes, is aggregation at the secondary minimum. Further calculations using the Marmur model will shed new light on this issue. Acknowledgements Grants 93r2288-6; 96r0704-0 from FAPESP and 510022r93-6; 520186r96-6 from CNPq are gratefully acknowledged. References w1x A.M. Carmona-Ribeiro, B.R. Midmore, Langmuir 8 Ž1992. 801. w2x A.M. Carmona-Ribeiro, T.M. Herrington, J. Colloid Interface Sci. 156 Ž1993. 19. w3x L.R. Tsuruta, M.M. Lessa, A.M. Carmona-Ribeiro, Langmuir 11 Ž1995. 2938.
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w4x L.R. Tsuruta, M.M. Lessa, A.M. Carmona-Ribeiro, J. Colloid Interface Sci. 175 Ž1995. 470. w5x L.R. Tsuruta, A.M. Carmona-Ribeiro, J. Phys. Chem. 100 Ž1996. 7130. w6x S.M. Sicchierolli, A.M. Carmona-Ribeiro, Colloids Surf. B Biointerfaces 5 Ž1995. 57. w7x M.M. Lessa, A.M. Carmona-Ribeiro, J. Colloid Interface Sci. 182 Ž1996. 166. w8x S.M. Sicchierolli, A.M. Carmona-Ribeiro, J. Phys. Chem. 100 Ž1996. 16771. w9x A.M. Carmona-Ribeiro, J. Phys. Chem. 93 Ž1989. 2630. w10x A.M. Carmona-Ribeiro, J. Phys. Chem. 96 Ž1992. 9555. w11x A.M. Carmona-Ribeiro, J. Phys. Chem. 97 Ž1993. 4247. w12x A.M. Carmona-Ribeiro, J. Phys. Chem. 97 Ž1993. 11843. w13x Y. Kagawa, E. Racker, J. Biol. Chem. 241 Ž1966. 2467. w14x G. Houser, S. Fleischer, A. Yamamoto, Lipids 5 Ž1970. 594. w15x M. Stelmo, H. Chaimovich, I.M. Cuccovia, J. Colloid Interface Sci. 117 Ž1987. 200. w16x O. Schales, S.S. Schales, J. Biol. Chem. 140 Ž1941. 879. w17x C.H. Huang, Biochemistry 8 Ž1969. 344. w18x C.D. Tran, P.L. Klahn, A. Romero, J.H. Fendler, J. Am. Chem. Soc. 100 Ž1978. 1622. w19x P.M. Claesson, A.M. Carmona-Ribeiro, K. Kurihara, J. Phys. Chem. 93 Ž1989. 917. w20x R.H. Ottewill, T. Walker, J. Chem. Soc. Faraday Trans. I 70 Ž1974. 917. w21x B. Vincent, J. Colloid Interface Sci. 175 Ž1973. 270. w22x A. Marmur, J. Colloid Interface Sci. 72 Ž1979. 41.