Shear-Induced Phase Separation in Polyelectrolyte/Mixed Micelle Coacervates

Transcription

1 pubs.acs.org/langmuir 2009 American Chemical Society Shear-Induced Phase Separation in Polyelectrolyte/Mixed Micelle Coacervates Matthew W. Liberatore,* Nicholas B. Wyatt, and MiKayla Henry Department of Chemical Engineering, Colorado School of Mines, Golden, Colorado Paul L. Dubin and Elaine Foun Department of Chemistry, University of Massachusetts, Amherst, Massachusetts Received June 1, Revised Manuscript Received September 25, 2009 A quantitative study of the shear-induced phase separation of a polycation/anionic-nonionic micelle coacervate is presented. Simultaneous rheology and small-angle light scattering (SALS) measurements allow the elucidation of micrometer-scale phase separation under flow in three coacervate solutions. Below 18 C, all three of the coacervate solutions are optically clear Newtonian fluids across the entire shear rate range investigated. Once a critical temperature range and/or shear rate is achieved, phase separation is observed in the small-angle light scattering images and the fluid exhibits shear thinning. Two definitive SALS patterns demonstrate the appearance of circular droplets at low shear rates near the critical temperature and ellipsoidal droplets at higher temperatures and shear rates. The shear-induced droplets range in size from 1 to4μm. The ellipsoidal droplets have aspect ratios as high as 4. A conceptual picture in which shear flow extends the polyelectrolyte chains of the clear coacervate liquid phase is proposed. The extended chains create interpolyelectrolyte-micelle interactions and promote expulsion of small ions from the complex, resulting in the formation of micrometer-scale phase-separated droplets. Introduction When two oppositely charged macromolecules [such as polyelectrolytes (PEs)] are mixed, spontaneous liquid-liquid phase separation can occur with the formation of a dense macroion-rich phase in equilibrium with a dilute macroion-poor phase. This process, called complex coacervation, converts a metastable suspension of coacervate droplets to separate coacervate and dilute supernatant phases upon standing or centrifugation. 1 Coacervates composed of polyelectrolytes and oppositely charged colloidal particles can be found in shampoos where the nanoparticles are micelles or in food formulations where the nanoparticles are proteins. Such coacervates form a locally segregated environment that can allow selective absorption of apolar molecules from the surrounding medium, e.g., extracting aliphatic compounds from solutions. 2,3 Coacervates can also be used to deliver drugs, nutraceuticals, or topically active ingredients. 4-6 Complex coacervation can be viewed as an entropically favorable ion-exchange process, which occurs when the oppositely charged macroions release some of their initially bound counterions upon forming a complex. When the concomitant loss of hydration is smaller than that leading to precipitation, liquid-liquid phase separation will occur. Since soluble complexes, or aggregates thereof, are the precursors of coacervation, their mutual repulsion must be *To whom correspondence should be addressed. mliberat@ mines.edu. (1) Stuart, M. A. C. Colloid Polym. Sci. 2008, 286(8-9), (2) Sudbeck, E. A.; Dubin, P. L.; Curran, M. E.; Skelton, J. J. Colloid Interface Sci. 1991, 142(2), (3) Luque, N.; Rubio, S.; Perez-Bendito, D. Anal. Chim. Acta 2007, 584(1), (4) Thimma, R. T.; Tammishetti, S. J. Microencapsulation 2003, 20(2), (5) Zhang, L.; Liu, Y. Z.; Wu, Z. C.; Chen, H. X. Drug Dev. Ind. Pharm (3), (6) Gander, B.; Blanco-Prieto, M. J.; Thomasin, C.; Wandrey, C.; Hunkeler, D. Encycl. Pharm. Technol. 2006, 1 5. weakened for this transition to take place. Liquid-liquid phase separation occurs only when the charge fraction f (the ratio of the number of macroion charges of a given sign to the total macroion charge) is close to 0.5. It is important to identify f as the charge ratio in the complex (microstoichiometry) and distinguish it from that of the entire system, f bulk (bulk or macrostoichiometry, also known as the mixing ratio) and, further, to recognize that f itself could present some polydispersity. Phase separation in the region of bulk charge stoichiometry has been reported for mixtures of cationic chitosan with synthetic polyanions, 7 histones with DNA or with poly(styrene sulfonate), 8 and lysozyme with poly(styrene sulfonate). 9 Coacervation can take place in the vicinity of an f bulk of 0.5 because of system polydispersity or because of disproportionation or polarization. 10 However, in these cases, the residual charge of the coacervating species will result in structure formation at submicrometer length scales, the morphologies of which are currently being determined. 9,11-16 (7) Mincheva, R.; Manolova, N.; Paneva, D.; Rashkov, I. Eur. Polym. J. 2006, 42(4), (8) Raspaud, E.; Chaperon, I.; Leforestier, A.; Livolant, F. Biophys. J. 1999, 77 (3), (9) Gummel, J.; Boue, F.; Clemens, D.; Cousin, F. Soft Matter 2008, 4(8), (10) Zhang, R.; Shklovskii, B. T. Physica A 2005, 352(1), (11) Wang, X. Y.; Lee, J. Y.; Wang, Y. W.; Huang, Q. R. Biomacromolecules 2007, 8(3), (12) Chodankar, S.; Aswal, V. K.; Kohlbrecher, J.; Vavrin, R.; Wagh, A. G. Phys. Rev. E: Stat. Phys., Plasmas, Fluids, Relat. Interdiscip. Top. 2008, 78, 3. (13) Singh, S. S.; Aswal, V. K.; Bohidar, H. B. Int. J. Biol. Macromol. 2007, 41 (3), (14) Kayitmazer, A. B.; Strand, S. P.; Tribet, C.; Jaeger, W.; Dubin, P. L. Biomacromolecules 2007, 8(11), (15) Kayitmazer, A. B.; Bohidar, H. B.; Mattison, K. W.; Bose, A.; Sarkar, J.; Hashidzume, A.; Russo, P. S.; Jaeger, W.; Dubin, P. L. Soft Matter 2007, 3(8), (16) Menjoge, A. R.; Kayitmazer, A. B.; Dubin, P. L.; Jaeger, W.; Vasenkov, S. J. Phys. Chem. B 2008, 112(16), DOI: /la903260r Published on Web 10/23/2009

2 Liberatore et al. Coacervates formed from polyelectrolytes and proteins of opposite charge have been studied by static and dynamic light scattering (DLS), 17 small-angle neutron scattering, fluorescence recovery after photobleaching, 15 rheology, 17,18 total internal reflectance microscopy, cryo-tem, 15 and pulsed-field gradient NMR. 16 The results of these studies show that these optically clear, viscous fluids have complex internal structures that are heterogeneous on many length scales. While biological and biotechnological objectives motivate studies with proteins, their idiosyncratic charge anisotropies complicate elucidation of electrostatic effects. In this regard, PE/micelle coacervates represent a simplification, even though micelle lability is a consideration. In particular, mixtures of polyelectrolytes with ionic-nonionic mixed micelles are good model systems because the micelle surface charge density can be modulated by the mole fraction of ionic surfactant, especially when the surfactant concentrations are much higher than the mixed surfactant critical micelle concentration. Dubin et al. performed extensive studies on a system comprised of poly(dimethyldiallylammonium chloride) (PDADMAC), together with mixed micelles of the anionic surfactant sodium dodecyl sulfate (SDS) and the nonionic surfactant Triton X-100 (TX100), using narrow molecular weight distribution samples of the nonhydrophobic polycation. The authors showed that the micelle surface charge density (σ) and surface potential (j) varied directly with the mole fraction of the anionic surfactant ( Y ). 19 Consequently, gradual addition of SDS to a mixture of polycation and nonionic micelles resulted in progressive changes in σ and j, leading to transitions from noninteracting solutions to soluble complexes at Y c, and then to liquid-liquid phase separation (coacervation) at Y φ. While Y φ appears to be a true liquid-liquid phase transition, becoming infinitely sharp when system polydispersity is eliminated, 20 Y c may be more appropriately described as a secondorder phase transition. 21 These transitions can be identified by dynamic light scattering, electrophoretic mobility, or precise turbidimetry. Turbidimetry was used to obtain phase behavior as a function of micelle surface charge density, PE molecular weight, PE:surfactant stoichiometry, and ionic strength. These measurements led to maps that identify the conditions corresponding to soluble complex formation, coacervation, and precipitation. 22 In contrast to PE/protein systems, the PE/micelle solutions undergo temperature-induced coacervation, which also appears to be a true liquid-liquid phase separation when system polydispersity is minimized. 23 The critical temperature of the liquid-liquid phase transition (T φ ) decreases with an increasing PE molecular weight, and T φ is highly sensitive to the other key variables, including ionic strength, micelle surface charge density, and PE:surfactant stoichiometry. 23 The effects of the micelle surface charge density and PE:surfactant stoichiometry are closely correlated with conditions at which the effective Article Figure 1. Turbidity as a function of temperatures for a coacervate solution. The transition from clear to turbid allows the determination of critical temperature T j 0. Reproduced from ref 26. Copyright American Chemical Society. (electrophoretic) charge of soluble complexes is close to zero, 20 which has also been observed for protein/pe systems. 24,25 Low-speed centrifugation of the metastable droplet suspensions formed when T > T φ yields optically clear, viscous fluids (typically 5-10 wt % surfactant and 1-2 wt % PE), which display some interesting phenomena. DLS shows abundant species with a diffusivity that is only 7 times smaller than that of micelles in dilute solution (despite a coacervate viscosity that is 100 times larger) and are therefore described as free micelles. 26 The equilibrium nature of the dense coacervate fluids thus formed is well-established by multiple studies, which demonstrate that their properties are reversible and independent of both the time and route of preparation, as long as irreversible liquid-solid separation is avoided. 27 When this coacervate is heated, a second phase separation temperature (denoted as T φ 0 ) is observed turbidimetrically as shown in Figure 1 from ref 26. As pointed out in ref 26, the corresponding transition is essentially reversible, only showing a weak hysteresis. Of particular interest is the appearance of a second phase with shear or elongational flows at temperatures slightly below T φ This flow-induced phase separation was observed when samples were loaded into confined geometries as shown in Figure 2 of ref 26. A substantial body of literature describes shear-induced phase separation (SIPS) 28 for wormlike micelles and selected highmolecular weight (MW) polymers, 28,35,36 but this is the first observation of SIPS for a polymer-micelle complex. Since neither the micelle nor the PE alone exhibits such behavior, the implication is that complexation with PE can cause ellipsoidal micelles to behave like other macromolecular systems exhibiting SIPS. One interesting consequence of coacervation is the possibility of achieving very high viscosities at relatively low surfactant concentrations, replacing surfactants with lower concentrations of less expensive polymers. (17) Bohidar, H.; Dubin, P. L.; Majhi, P. R.; Tribet, C.; Jaeger, W. Biomacromolecules 2005, 6(3), (18) Mohanty, B.; Bohidar, H. B. Int. J. Biol. Macromol. 2005, 36(1-2), (19) Dubin, P. L.; The, S. S.; McQuigg, D. W.; Chew, C. H.; Gan, L. M. Langmuir 1989, 5(1), (20) Wang, Y. L.; Kimura, K.; Huang, Q. R.; Dubin, P. L.; Jaeger, W. Macromolecules 1999, 32(21), (21) McQuigg, D. W.; Kaplan, J. I.; Dubin, P. L. J. Phys. Chem. 1992, 96(4), (22) Wang, Y. L.; Kimura, K.; Dubin, P. L.; Jaeger, W. Macromolecules 2000, 33(9), (23) Kumar, A.; Dubin, P. L.; Hernon, M. J.; Li, Y. J.; Jaeger, W. J. Phys. Chem. B 2007, 111(29), (24) Xia, J. L.; Dubin, P. L.; Kim, Y.; Muhoberac, B. B.; Klimkowski, V. J. J. Phys. Chem. 1993, 97(17), (25) Gupta, A.; Reena; Bohidar, H. B. J. Chem. Phys. 2006, 125, 5. (26) Dubin, P. L.; Li, Y. J.; Jaeger, W. Langmuir 2008, 24(9), (27) Kaibara, K.; Okazaki, T.; Bohidar, H. B.; Dubin, P. L. Biomacromolecules 2000, 1(1), (28) Larson, R. G. Rheol. Acta 1992, 31(6), (29) Fischer, P.; Wheeler, E. K.; Fuller, G. G. Rheol. Acta 2002, 41(1-2), (30) Oda, R.; Panizza, P.; Schmutz, M.; Lequeux, F. Langmuir 1997, 13(24), (31) Narayanan, J.; Manohar, C.; Kern, F.; Lequeux, F.; Candau, S. J. Langmuir 1997, 13(20), (32) Liu, C. H.; Pine, D. J. Phys. Rev. Lett. 1996, 77(10), (33) Rehage, H.; Hoffmann, H.; Wunderlich, I. Phys. Chem. Chem. Phys. 1986, 90(11), (34) Hu, Y. T.; Boltenhagen, P.; Pine, D. J. J. Rheol. 1998, 42(5), (35) Migler, K.; Liu, C. H.; Pine, D. J. Macromolecules 1996, 29(5), (36) Rangelnafaile, C.; Metzner, A. B.; Wissbrun, K. F. Macromolecules 1984, 17(6), DOI: /la903260r 13377

3 Article Liberatore et al. Table 1. Preparation Conditions and Properties of the Coacervate Solutions a sample Y b C P b (g/l) f - c MW n b (kda) T j ( C) d T j 0 ( C) d A B C ( 2 a All coacervates prepared in 0.40 M NaCl. b Solution from which coacervate is obtained. Y is the mole fraction of anionic surfactant, and C P and MW are the polymer concentration and number-average molecular weight, respectively. c The number of SDS charges divided by the sum of SDS and polycation charges. d Property of coacervate (see the text). T j and T 0 j values are all (1 C, except as shown. Figure 2. Photo of coacervate exhibiting flow-induced phase separation in a confined sample geometry. Reproduced from ref 26. Copyright American Chemical Society. In this work, we characterize the thermal and shear-induced phase transitions in this polyelectrolyte/micelle system by smallangle light scattering (SALS) to determine the onset of phase separation as a function of the macromolecular solute concentration, shear rate, and temperature. While previous SANS and cryo-tem indicated the formation of dense phases (i.e., micellerich phases) at length scales of several hundred nanometers under quiescent conditions, this work probes the formation of the dense phase under shear at larger length scales. In addition, the size and shape of the phase-separated droplets are measured and a conceptual picture of the transient structures is proposed. This work is representative of ongoing efforts to provide a molecular basis for understanding the macroscopic properties of intermacroionic coacervates. Experimental Methods Materials. Poly(diallyldimethylammonium chloride) (PDA- DMAC) was prepared by free radical aqueous polymerization of diallylmethylammonium chloride. 37 The weight- and numberaverage molecular masses of the purified lyophilized polymer were and Da, determined by light scattering and membrane osmometry, respectively. 37 Triton X-100 (TX100), a nonionic surfactant, was purchased from Fluka, and sodium dodecyl sulfate (SDS), an anionic surfactant, with a purity of >99% and NaCl were purchased from Fisher. All were used without further purification. Milli-Q water was used in all samples. Coacervate Preparation. PDADMAC/TX100 solutions (containing either 3 g/l PDADMAC with 20 mm TX100 or 2 g/l PDADMAC with 10 mm TX100) and 60 mm SDS solutions were prepared separately in 0.4 M NaCl. The samples were brought to coacervation via addition of SDS to the mixed PDADMAC/TX100 solution to produce the desired mole fraction of SDS defined as Y=[SDS]/([SDS] + [TX100]). Coacervates can also be prepared by the addition, at fixed Y and ionic strength, of mixed micelles to PDADMAC, or PDADMAC to micelles, 38 although systematic comparisons of the coacervates so formed are incomplete. The equilibrium nature of this system is demonstrated by the reversibility under conditions corresponding to soluble complexation or coacervation, although precipitation (e.g., induced by addition of SDS to PDADMAC in the absence (37) Dautzenberg, H.; Gornitz, E.; Jaeger, W. Macromol. Chem. Phys. 1998, 199 (8), (38) Dubin, P. L.; Rigsbee, D. R.; Gan, L. M.; Fallon, M. A. Macromolecules 1988, 21(8), of nonionic surfactant) is effectively irreversible. As noted above, coacervation could also be attained by increasing the temperature above a critical value T j which depends on Y, polymer concentration (C P ) and molecular weight, and ionic strength. 23 The value of T j for the formation of coacervate from a homogeneous polycation/mixed micelle solution is typically 5-10 C below the value of T 0 j, the temperature at which the coacervate itself undergoes additional phase transition (see Figure 1). Thus, values of Y and C P were chosen to produce different values of T j and hence T 0 j.theturbid sample was then centrifuged for 1 h at 3500 rpm to produce an optically clear dilute (upper) phase and a dense (lower) phase ( coacervate ). Preparation conditions and values of T j and T 0 j are given in Table 1. The progressive increase in T 0 j from sample A to sample B to sample C indicates that these coacervate samples (at, e.g., 20 C) correspond to further progress toward coacervate phase separation, which is confirmed by results reported below. We note that values of T 0 j (determined by turbidimetry) in Table 1 are typically 2 C higher than the onsets of phase separation as determined by SALS as discussed below. The differences in the values of T 0 j are probably a result of the heterogeneity of the coacervate, arising from the heterogeneity of TX100 and the heterogeneity among the soluble complexes from which coacervate forms. Because of the heterogeneity of several components of these samples, relative contributions of components may differ for turbidity versus small-angle scattering. The effect of system heterogeneity, arising to a large extent from the chemical heterogeneity of TX100 (vis-a-vis, for example, C 12 E 23 8 ), is to broaden all observable transitions, making it difficult to establish their order. Rheology and Small-Angle Light Scattering. Rheology and SALS data were collected simultaneously using an AR-G2 rheometer (TA Instruments, New Castle, DE) with the commercially available SALS attachment. A transparent parallel plate configuration (50 mm diameter) with a 1 mm gap was used for all tests. SALS images were recorded for at least four different shear rates (from 0.1 to 32 s -1 )along the flow curves for each of the three different samples (A, B, and C) over a range of temperatures. All images were taken at steady state; i.e., the viscosity and SALS image were not changing with time at a given shear rate. The size scale of scattering objects captured by this SALS setup is μm (q = μm -1 ). To examine the shearinduced phase separations, the SALS images were analyzed using ImageJ with standard protocols for subtracting the background and removing the beam stop from the raw images. 39,40 The locations of the phase boundaries under shear were determined by the normalized mean intensity of the SALS images. The normalized mean intensity is a representation of the turbidity of the solutions from the perspective of small-angle scattering. Similar analysis has been used to determine critical shear rates (39) Kline, S. R. J. Appl. Crystallogr. 2006, 39, (40) TA Instruments. AR Series Small Angle Light-Scattering (SALS) Accessory Manual; TA Instruments: Newark, DE, DOI: /la903260r

4 Liberatore et al. Article Figure 4. Viscosity (either Newtonian μ or η o from the Cross model fit) as a function of temperature for three coacervate solutions: (a) sample A, (b) sample B, and (c) sample C. Figure 3. Viscosity as a function of shear rate at several temperatures for three coacervate solutions: (a) sample A, (b) sample B, and (c) sample C. from SALS images. 41 The normalized mean intensity is defined as the ratio of the average intensity of the image (after removal of the beam stop) to the intensity if all pixels in the image were saturated (i.e., a value of 255 for the 8-bit camera used). A positive value of the normalized mean intensity indicates the presence of micrometer size structures in the flow; however, (41) Saito, S.; Hashimoto, T.; Morfin, I.; Lindner, P.; Boue, F. Macromolecules 2002, 35(2), the magnitude of the normalized mean intensity does not correlate with the size of the micrometer-scale scattering objects. Following the method employed by Walker et al., 42 the correlation length (a c ) and aspect ratio (a r ) of the droplets were determined using Debye-Bueche 43 plots (I -0.5 vs q 2 ). A linear fit to the radially averaged data plotted in the Debye-Bueche format was calculated in each case using the method of least squares (example plot included as Figure S1 of the Supporting Information). The slope and intercept of the linear fit were used to calculate a characteristic length [a c = (slope/intercept) 0.5 ] for the droplets at various shear rates. (42) Walker, L. M.; Kernick, W. A.; Wagner, N. J. Macromolecules 1997, 30(3), (43) Debye, P.; Bueche, A. M. J. Appl. Phys. 1949, 20, DOI: /la903260r 13379

5 Article Liberatore et al. A calibration of the Debye-Bueche characteristic length was performed using a Microbead NIST Traceable Particle Size Standard (Polysciences, Inc., Warrington, PA) of polystyrene microspheres (3 μm in diameter). A correction factor was determined by comparing the calculated a c with the reported diameter of the standard. This correction factor was then applied to our calculations of a c for the coacervate solutions. In addition, the distortion of the droplet shape with increasing shear rate was characterized by the aspect ratio, which was determined by taking the ratio of the q values in the vorticity and flow directions for a given value of the measured intensity. Results and Discussion The rheology of the coacervate solutions shows dramatic changes as a function of temperature and shear rate. The thermodynamically stable coacervates behave as Newtonian fluids at low temperatures (typically below T 0 φ ) across the entire shear rate range studied (Figure 3). Once the samples begin to exhibit shear-induced phase separation (as observed by SALS), their viscosity becomes shear thinning. The Cross rheological model (eq 1) was used to fit the non-newtonian response of the coacervates. 44 The model quantifies the zero shear viscosity, degree of shear thinning, and relaxation time of the fluid. The fits (included as Figure S2 and Table S1 of the Supporting Infromation) exhibit similarities in all three coacervates. First, the shear thinning index is 1 for almost all of the temperatureshear rate combinations exhibiting shear-induced phase separation (as quantified by SALS). Since the shear thinning index is 1, the samples exhibit a stress plateau (i.e., shear stress is independent of shear rate), which is one indicator of possible shear banding. 34,45 In addition, the relaxation time increases with an increase in temperature. Therefore, the onset of shear thinning, which corresponds to the appearance of SIPS, appears at lower shear rates at higher temperatures. η ¼ η 0 -η 1 þð_γτ R Þ m þ η ð1þ The coacervate samples exhibit similarites in the temperature dependence of the viscosity [either μ when Newtonian or η o from the Cross model (Figure 4)]. Before entering the phase separating regime, samples A and C exhibit a simple monotonic decrease in the viscosity as the temperature is increased; however, the viscosity of sample B is nearly independent of temperature below T j. As all of the coacervate solutions approach the phase boundary (by temperature, shear, or the combination of the two), the solutions begin to exhibit shear thinning. All of the samples exhibit an increase in the viscosity (for 5-8 C) once in the shear thinning regime. The increased viscosity is an indication of the phase separation occurring; i.e., the transition from the nanoscale domain to micrometer-scale structures would increase the viscosity. The shear thinning regime begins 1-5 C below the critical temperature, T j 0. Finally, the viscosity decreases for the highest temperature measured for all samples. The turbidity (and thus the phase boundaries) of the solutions from the perspective of small-angle scattering is quantified using the normalized mean intensity (Figure 5). The beam stop does not permit collection of the light exiting the sample at a scattering angle of 0 ; thus, direct comparison with the previous turbidity measurements 40 used to locate the phase boundaries is not possible. The lowest temperature with a non-zero normalized (44) Cross, M. M. J. Colloid Sci. 1965, 20, (45) Hu, Y. T.; Lips, A. J. Rheol. 2005, 49(5), Figure 5. Normalized mean intensity of the SALS images as a function of temperature at several shear rates for samples A (a), B (b), and C (c). Dashed lines are to guide the eye. Gray shading indicates the region in which the samples exhibit SIPS. The vertical line indicates the location of T j 0. mean intensity indicates the location of the phase boundary at that representative shear rate (indicated by the gray regions in Figure 5). For example, sample A exhibits shear-induced phase separation at 20 C for all four shear rates, which is 2 CbelowT j 0 (Figure 5a). Sample B also exhibits SIPS below T j 0 (at 22 Cand 0.1 s -1 in Figure 5b). The first appearance of SIPS for sample C is at 27 C for all measured shear rates (Figure 5c), confirming that SIPS in all three samples is measured below T j 0. The anomalous appearance of Figure 5c might suggest heterogeneity of transitions, and temperature-induced transitions in quiescent samples do become markedly sharper when system polydispersity is reduced via replacement of the chemically polydisperse nonionic surfactant TX-100 with C 12 E 8, a monodisperse analogue. 23 Overall, the nonmonotonic nature of the normalized mean intensity as a function of temperature does not directly correlate with the size of DOI: /la903260r

6 Liberatore et al. Figure 6. Viscosity as a function of temperature for sample A at a shear rate of 10 s -1 showing the transition from a one-phase state to a phase-separated state. Arrows correspond to the locations of reported SALS images. the phase-separated domains. The definitive information from determination of the normalized mean intensity is the location of the transition from a single-phase fluid to a phase-separated solution. The shear-induced phase boundary may also be characterized rheologically using a temperature sweep. For example, sample A was heated at a rate of 1 C/min while being held at a constant shear rate of 10 s -1 (Figure 6). The evolution of the viscosity of the coacervate is continuous with three distinct regions (i.e., one phase at low temperatures, transition to the second phase near T j 0, and a two-phase region at higher temperatures). At low temperatures, the viscosity decreases slowly with an increase in temperature until 22 C (an average change in viscosity of Pa s C -1 ). Between 22 and 25 C, the viscosity drops much more dramatically (-0.12 Pa s C -1 ). The clear coacervate solution at temperatures below 20 C undergoes a transition to a completely phase-separated state at 25 C and 10 s -1.Above 25 C, the viscosity changes very slowly with temperature ( Pa s C -1 ). The appearance of phase-separated structures via SALS (see the SALS image at 22 CinFigure6)precedes the period of strongly decreasing viscosity. Samples B and C also exhibit shear-induced phase separation before the large decrease in the viscosity of the solution. Therefore, SALS is a more sensitive measure of SIPS than viscosity measurements alone. The native aggregate structure of these two samples can be directly compared using cryo-tem (Figure 7). Sample B exhibits a distribution of ca. 50 nm aggregates interconnected to form extended clusters. For sample A, these clusters appear to be disconnected and collapsed into more dense objects similar in size. These sizes are identical to those seen for soluble interpolymer aggregates prior to coacervation; the aggregates appear to arise from association of smaller intrapolymer complexes. For sample A, the cryo-tem snapshot is taken at a vitrification temperature well above T j 0. However, the T j 0 of sample B is close to the cryo-tem vitrification temperature of 24 C. The native 50 nm aggregates in the coacervates transform into the micrometer-scale objects under shear, which are characterized by SALS. It is of interest to compare samples A and B vis-a-vis the values of f - in Table 1, inasmuch as sample A, formed from a mixture closer to charge stoichiometry than sample B (f - values of 0.49 and 0.39, respectively), shows more collapsed aggregates (Figure 7) and shows a transition at a lower temperature (Figure 5). The importance of charge stoichiometry has been underlined by several experimental 9 and theoretical 10 works. Article However, the correlation between the bulk or mixing charge stoichiometry, e.g., f -, and the microscopic stoichiometry (the value of f - in the coacervate or the complex aggregates that precede them) is less clear for phase separation in the salty solutions studied here than in salt-free systems that undergo phase separation stoichiometrically. 46,47 For example, a coacervate formed at Y=0.50, at an ionic strength of 0.80 M, was found to contain only 23% of the total polymer and even less (8%) of the total surfactant, preferentially the anionic, i.e., a Y value of 0.61 in the coacervate; 48 with a similar result from a solution in 0.40 M NaCl and a Y of 0.35, yielding a coacervate with a Y of 0.51, and with excess polycation on a charge basis. 49 The tendency toward charge neutralization and retention of counterions are both factors that influence coacervate composition. In addition, comparisons of samples A and C with nearly identical values of f - show that total macromolecular concentrations influence coacervate properties. The pronounced effects of polycation MW at fixed f - noted elsewhere 23 also suggested caution in interpretation of the effects of bulk stoichiometry here. Two-dimensional SALS images provide information about the microstructure of a fluid by inspection. Overall, the coacervates undergo a transition from a homogeneous isotropic one-phase solution to a heterogeneous anisotropic phase-separated system. A set of scattering images for sample A are representative of the three main types of scattering measured for the coacervate system as a function of temperature and shear rate (Figure 8). An almost completely black image is observed at lower temperatures and all shear rates. The coacervate phase separation is a nanoscale phenomenon under low-temperature conditions and thus cannot be detected by SALS. Next, a circular region of high scattering intensity is observed (e.g., T= 22 C and shear rate = 0.1 s -1 in Figure 8). The circular scattering pattern indicates a nearly circular droplet on the micrometer scale. The larger areas of intense scattering in the images at 22 C versus 20 C(atashearrateof0.1s -1 )indicate a smaller droplet size (i.e., images represent length scales in inverse space). The third type of scattering image as seen at 24 C and1s -1 (Figure 8, right column) is ellipsoidal with the long dimension of high-intensity scattering in the vorticity direction. SALS is a more sensitive measurement of shear-induced phase separation than flow rheology as indicated by the appearance of micrometer-scale structures via SALS before the sample begins to shear thin (e.g., sample A at 22 C and1s -1 ). Measured SALS images for samples B and C are included as Supporting Information (Figures S3 and S4 of the Supporting Information). Similar transitions from no scattering to circular and ellipsoidal scattering are observed with increasing temperatures for all three samples. Micron-sized scattering objects are observed at g20 C for sample A (at all of the measured shear rates). The phaseseparated droplets are very close to circular (assumed to be spherical if three-dimensional data were available) at a shear rate of 0.1 s -1 and ellipsoidal at all higher shear rates. Sample B exhibits phase-separated droplets beginning at 24 C (nearly circular 2D scattering image). Ellipsoidal droplets are observed for sample B from 24 Cand1s -1 to 30 C and 32 s -1. SALS from (46) Ahmed, L. S.; Xia, J. L.; Dubin, P. L.; Kokufuta, E. J. Macromol. Sci., Pure Appl. Chem. 1994, A31(1), (47) Tsuboi, A.; Izumi, T.; Hirata, M.; Xia, J. L.; Dubin, P. L.; Kokufuta, E. Langmuir 1996, 12(26), (48) Dubin, P. L.; Oteri, R. J. Colloid Interface Sci. 1983, 95(2), (49) Davis, D. D. Intermacromolecular association of polycations with oppositely charged micelles. M.S. Thesis, Purdue University, West Lafayette, IN, DOI: /la903260r 13381

7 Article Liberatore et al. Figure 7. Cryo-TEM images of (a) sample A and (b) sample B showing a native aggregate size of 50 nm. Figure 8. Small-angle light scattering images as a function of temperature at two shear rates for sample A. SIPS occurs at higher temperatures and shear rates. sample C first appears at a higher temperature, 27 C, than that from samples A and B and exhibits the same progression through circular and ellipsoidal scattering as temperature and shear rate increase. The characteristic size and aspect ratio of the phase-separated droplets were derived from the radially averaged intensity (e.g., using Debye-Bueche plots like Figure S1 of the Supporting Information). The droplet size ranges from 1 to 4 μm, and the aspect ratio of the droplet varies from 1 to 4. The continuous Figure 9. (a) Viscosity as a function of shear rate at 26 C for sample A (top) with inset SALS patterns representing the smooth transition from circular to ellipsoidal droplets with an increase in aspect ratio. (b) Characteristic length (a c ) and aspect ratio of the phase-separated droplets for sample A corresponding to the SALS patterns in panel a. transition from no small-angle scattering to circular and finally elongated scattering patterns is observed with an increasing shear rate at a constant temperature. For example, sample A shows the continuous progression of shapes in the shear-induced structures at a temperature above T φ 0 (Figure 9). At 0.1 s -1 and 26 C, a nearly circular SALS pattern is observed, which corresponds to a droplet with an a c of 1.0 μm and an a r of 1.0. At 0.1 s -1, approximately 1 μm nearly circular droplets appear as the first DOI: /la903260r

8 Liberatore et al. Article Figure 10. Schematic representation of shear- and temperatureinduced phase separation for PDADMAC/TX100-SDS coacervate. Both processes involve loss of counterions arising from an increased number of polyelectrolyte-micelle interactions, but by an intercomplex vs intracomplex mechanism for the former. Reproduced from ref 26. Copyright American Chemical Society. observation of shear-induced phase separation in all three coacervate solutions. The nearly circular droplet becomes elongated at higher shear rates until the aspect ratio of the droplet reaches 3.0 at 25 s -1 for sample A (Figure 9). The increasing aspect ratio with shear rate indicates the growth of the droplets is almost exclusively in the flow direction (and is observed for all three samples). A conceptual picture of SIPS in coacervates can be derived from the experimental observations presented earlier. In general, shear flow transforms the clear coacervate fluid with domain sizes of nm by transforming the polyelectrolyte/micelle complexes into extended chains or necklaces of polyelectrolyte decorated with micelle beads 50 as portrayed in Figure 10. In a manner entirely analogous to shearinduced phase separation of simple polymers, 28 these extended chains create efficient inter(polyelectrolyte/micelle) interactions, presumably at the expense of intra(polyelectrolyte/micelle) interactions, with complementary spacing of bound micelles that can then interact electrostatically with micelle-poor domains of adjacent complexes, a form of polarization. 10 These more efficient interactions promote the expulsion of small ions from the complex, resulting in the formation of micrometer-scale phase-separated droplets. The phase transition temperature T j 0 coincides with the transition from shear-independent (Newtonian) to shear-dependent viscosity. The initial appearance of small-angle scattering precedes the transition in rheology from Newtonian to shear thinning. Further studies are needed to correlate transition temperatures for quiescent phase transitions reported from turbidimetry and small-angle scattering with observations made under shear, a task that may be facilitated by the reduction of system polydispersity. Conclusions The phase behavior under flow of a polycation/mixed micelle coacervate was investigated by rheology and rheo-sals. (50) Lee, L. T.; Cabane, B. Macromolecules 1997, 30(21), Figure 11. Schematic of the relevant domain size of the coacervate solutions with changing temperature and shear rate. Although shear-induced phase separation has been extensively reported for solutions of certain polymers and wormlike micelles, this is the first observation of SIPS for a polymer/micelle system and likely the first quantitative study of SIPS in a complex coacervate. Under shear, the coacervate solutions convert from homogeneous, isotropic, one-phase systems to heterogeneous, anisotropic, two-phase systems. Thus, behavior typically observed for wormlike micelles is attained for small micelles bound to a polyelectrolyte. Figure 11 provides a summary of the shear- and temperature-induced phase transitions observed in this work. Below 18 C, all three of the coacervate solutions are optically clear Newtonian fluids across the entire shear rate range investigated. Once a critical temperature and/or shear rate is achieved, phase separation occurs. Two definitive SALS patterns demonstrate the appearance of circular droplets at low shear rates near the critical temperature and ellipsoidal droplets at higher temperatures and shear rates. The shear-induced droplets range in size from 1 to4 μm. The ellipsoidal droplets have aspect ratios as high as 4. Overall, the shear-induced phase separation has been explored as a function of shear rate and temperature at the steady state. Additional insights will be gained via exploration of the kinetics of the phase separation under flow and the possibility of shear banding. Acknowledgment. Partial support of this work was received from the donors of the Petroleum Research Fund (M.W.L.). Portions of this work were supported by grants from Shiseido Corp. (P.L.D.) and from the donors of the Petroleum Research Fund to A. Dinsmore and P.L.D. We acknowledge assistance from Dr. JoAn Hudson (AMRL, Clemson University, Clemson, SC) with cryo-tem. Supporting Information Available: Description of the material, an example fit of the small-angle light scattering data to the Debye-Bueche model, viscosity curves fit to the Cross model, model fit parameters, and 2D SALS images as a function of shear rate and temperature for samples B and C. This material is available free of charge via the Internet at DOI: /la903260r 13383