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Influence of the Formulation Process in Electrostatic Assembly of Nanoparticles and Macromolecules in Aqueous Solution: The Interaction Pathway Ling Qi,†,§ Je´rome Fresnais,‡ Jean-Franc¸ois Berret,‡ Jean-Christophe Castaing,† Fanny Destremaut,§ Jean-Baptiste Salmon,§ Fabrice Cousin,| and Jean-Paul Chapel*,†,⊥ Complex Assemblies of Soft Matter Laboratory (COMPASS), CNRS UMI3254, Rhodia Center for Research and Technology in Bristol, 350 Georges Patterson BouleVard, Bristol, PennsylVania 19007, Matie`re et Syste`mes Complexes (MSC), UMR 7057 CNRS, UniVersite´ Denis Diderot Paris-VII, Baˆtiment Condorcet, 10 rue Alice Domon et Le´onie Duquet, 75205 Paris, France, Lab of the Future (LOF), UMR 5258 Rhodia, CNRS, UniVersite´ Bordeaux 1, 178 aVenue du Docteur Schweitzer, F-33608 Pessac cedex, France, Laboratoire Le´on Brillouin (LLB), UMR CEA, CNRS 12, CEA Saclay, 91191 Gif-sur-YVette, France, and Centre de Recherche Paul Pascal (CRPP), UPR CNRS, UniVersite´ Bordeaux 1, 33600 Pessac, France ReceiVed: July 16, 2010; ReVised Manuscript ReceiVed: August 24, 2010
The influence of the formulation process/pathway on the generation of electrostatic complexes made from polyelectrolyte-neutral copolymers and oppositely charged nanocolloids is investigated in this work. Under strong driving forces like electrostatic interaction and/or hydrogen bonding, the key factor controlling the polydispersity and the final size of the complexes is the competition between the reaction time of the components and the homogenization time of the mixed solution. The latter depends on the mixing pathway and was investigated in a previous publication by tuning the mixing order and/or speed (Qi, L.; Fresnais, J.; Berret, J.-F.; Castaing, J.-C.; Grillo, I.; Chapel, J.-P. J. Phys. Chem. C 2010, 114 (30), 12870-12877). The former depends on the initial concentration of the individual stock solutions and the strength of the interaction and is investigated here on a system composed of anionic cerium oxide functional nanoparticles (CeO2-PAA) and cationic charged-neutral diblock copolymers (PTEA11K-b-PAM30K) or homopolyelectrolytes (PDADMAC100K). The electrostatic interaction was screened off completely by adding a large amount of salts. Desalting kinetics was then controlled by slowly decreasing the ionic strength from Ib ≈ 0.5 M, the minimum ionic strength to totally prevent the complexation of the two components, to lower values where the electrostatically screened system undergoes an (abrupt) transition between an unassociated and a clustered state. Neutron scattering data evidenced differences in the nanostructure of complexes formed by either dilution or simple mixing. Furthermore, adsorption optical reflectometry experiments showed the impact of these different formulation processes on the wettability and antifouling properties of treated silica and polystyrene model surfaces. Better controlled mixing processes were put forward at the end to improve the productivity and reproducibility of the complexes generation. In particular, a microfluidic chip coupled with dynamic light scattering was used to better control the hydrodynamics of the complexation process. Introduction The electrostatic complexation of macromolecules and nanoparticles2,3 has attracted much attention in the past decade. Combining the advantageous properties of both the organic and inorganic worlds offers a great promise for engineering versatile functional structures with controlled physical and chemical attributes at the nanometer scale. This synergy will certainly trigger the emergence of a wide range of novel materials and processing techniques in various scientific and technological fields such as material science4-8 and biology.9-13 Compared to the abundant work on the mechanisms, structure characterizations, and functionalities, however, not much attention has been paid to the formulation process, which is a key issue for generating functional systems or devices on a large scale. Under a strong interaction as in the case of an electrostatic * To whom correspondence should be addressed. E-mail: chapel@ crpp-bordeaux.cnrs.fr. † Rhodia Center for Research and Technology in Bristol. § UMR 5258 Rhodia, CNRS, Universite´ Bordeaux 1. ‡ Universite´ Denis Diderot Paris-VII. | UMR CEA, CNRS 12, CEA Saclay. ⊥ UPR CNRS, Universite´ Bordeaux 1.
complexation, it often takes a very long time to reach true equilibrium leading to the formation of out-of-equilibrium “frozen” structures, which are not thermodynamically favored. It has been pointed out that the order of addition of inorganic ions and polyelectrolytes affects the final structure of adsorbed polyelectrolyte layers,14-16 i.e., the resulting structure depends not only on the bulk composition but also on whether the polyelectrolyte or the salt was added first. Similarly, the order of addition of two oppositely charged polyelectrolyte solutions17 determines the final net charge of the system, and that deviation from 1:1 stoichiometry in the formed aggregates increases with the ionic strength of the system. The mixing protocol seems to have a18,19 great impact on the size of the aggregates initially formed. These processdependent features have of course important consequences in technological applications. Why is such complexation processdependent? The competition between the “reaction time” (depending on the initial concentration and the nature of the interaction) and the “homogenization time” (ranging from milliseconds20,21 to hours) of the mixed solution is certainly at the origin of the “process hysteresis”.
10.1021/jp106610t 2010 American Chemical Society Published on Web 09/13/2010
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The “homogenization time” depends on the mixing pathway and was investigated in a former publication1 by tuning the mixing order and/or speed on a very similar system composed of cerium oxide nanoparticles and charged-neutral diblock copolymers. The complexes final morphologies (size, shape, polydispersity) were found to depend strongly on the formulation process, while keeping at a smaller scale (clusters) the same nanostructure as shown via light and neutron scattering experiments. The impact of the structures of the complexes were evaluated on some bulk (rheology) and surface (wetting/ antifouling) properties. The results highlighted that a processdependent formulation seen a priori as a drawback can be turned into an advantage: different properties can be developed from different morphologies while keeping the chemistry constant. The “reaction time” depends on the initial concentration of the individual stock solutions and the nature of the interaction and is put under scrutiny here on a system composed of anionic cerium oxide functional nanoparticles (CeO2-PAA) and cationic charged-neutral diblock copolymers or homopolyelectrolytes. The complexation is here purely electrostatically driven and both basic components have a good stability toward high ionic strength (g1 M). These features enabled us to formulate “dormant solutions” in which the interaction between the two building blocks is completely screened off allowing the strength of the electrostatic interaction to be tuned by simple dilution or dialysis. The influence of the process on the nanostructures of the complexes together with their impact on the wettability and antifouling properties of treated model surfaces were studied. Finally, better controlled mixing processes were put forward at the end to improve the productivity and reproducibility of the generation of the complexes. In particular, a microfluidic chip coupled with dynamic light scattering was used to better control the hydrodynamics of the complexation process. Materials and Methods Chemicals. Nanoparticles. The nanoparticles used in this work were cationic cerium oxide nanocrystals, or nanoceria (CeO2). The CeO2 nanoparticles were synthesized by Rhodia chemicals. CeO2 dispersion is naturally stable only at pH 0.3 M) results in an irreversible aggregation of the particles, and destabilization of the sols leading eventually to a macroscopic phase separation. For this system, the destabilization of the sols occurs well below the point of zero charge of the ceria particles (pzc ) 7.9). The nanoceria particles have a ζ-potential ζ ) +30 mV and an estimated structural charge of QCeO2 ) +300 e.22,23 The hydrodynamic radius RH of the CeO2 particles was found by dynamic light scattering to be 4.9 ( 0.6 nm. Furthermore, in order to complex with cationic polyelectrolytes, ceria nanoparticles were coated with poly(acrylic acid) with a molecular weight of 2000 g/mol, noted hereafter as CeO2-PAA2K through a precipitation-redispersion process published previously.24 The structural charge is estimated in this case as QCeO2-PAA2K ) -700 e (at neutral pH). The hydrodynamic diameter RH of anionic CeO2-PAA2K particles (NP) was found by dynamic light scattering to be 6 ( 0.8 nm. Polymers. Both charged-neutral block copolymers and homopolyelectrolytes were used in this work. Poly(trimethylammonium ethylacrylate methylsulfate-b-poly(acrylamide)) was
Qi et al. TABLE 1: Main Characteristics of the Different Mixing Systems Used To Bring the Stock Solutions into Intimate Contact direct simple microfluidics mixing desaltinga micromixer chip mixing time, s interacting vol, mL flow rate, mL · h-1
1 1-10 3600
3600 10-100 NA
2 0.3 600
0.2 0.1 2
a
In the case of desalting, the mixing time is not relevant; the extent on which the electrostatic interaction is fully turned off is mentioned.
used as a cationic-b-neutral block copolymer abbreviated as PTEA11K-b-PAM30K in the paper. The values in subscript are the weight average molecular weight Mw obtained by the synthesis (controlled radical polymerization process-Rhodia MADIX technology25) with a polydispersity index Ip ) Mw/Mn ) 1.6 ( 0.1. Poly(diallyldimethylammonium chloride) abbreviated as PDADMAC100K with an Mw of about 100 000 g/mol was used as a cationic homopolyelectrolyte. It was purchased from Sigma-Aldrich and used without further purification. Formulation Protocol. Before mixing, dilute stock solutions of nanoparticles and polymers were prepared separately at the same concentration (c ) 0.1 wt %). The relative amount of each component was monitored by the volume ratio X, yielding for the final concentrations cNP ) cX/(1 + X) and cP ) c/(1 + X). Two different formulation paths were used: Direct Mixing. Three different mixing methods were used: adding drop by drop with a pipet or pouring or high speed injection (1-10 mL/s with a syringe) of a nanoparticle solution into a polymer solution.1 In all cases, magnetic stirring was started only at the end to redisperse any sediment and facilitate the sampling for further analyses. Micromixer. The micromixer is comprised of a chamber with two entries and one exit. The volume of the mixer is 0.3 mL (diameter 18 mm, height 1.2 mm) with an overall flow rate of 600 mL/h giving an average retention time of 2 s. Two stock solutions containing the individual nanoparticles and polymers are driven by two syringe pumps into the chamber at a given flow rate. Under the agitation of a ministirrer (600 rpm), both solutions are homogenized. The complexation occurs in the chamber. The final coacervates are collected at the exit. This setup minimizes any fluctuation and enables the tuning of the mixing ratio X accurately and continuously by adjusting the flow rate of each feeding solution. The mixed solution at different X can be easily collected and analyzed by light scattering (Table 1). Desalting-Dilution. Sodium chloride (NaCl) or ammonium chloride (NH4Cl) was added into the initial nanoparticle and polymer solutions until the ionic strength reached a value well above the critical ionic strength Ib, where the interaction is completely screened or turned off.26 The two salted solutions at the same ionic strength were then put together into a dialysis bag made of a cellulose membrane (MWCO ) 10 000 Da). The mixed solution was dialyzed against DI water for about 1 h. The water bath was 50 times larger than the dialyzed solution, ensuring a final ionic strength close to that of the bath. Probing Techniques. Light Scattering. Static (SLS) and dynamic (DLS) light scattering measurements are performed on a BI-9000AT Brookhaven spectrometer (with a vertically polarized laser operating at 488 nm). Rayleigh ratios R and hydrodynamic diameters are measured as a function of the concentration c. R is obtained from the scattered intensity I(c):
Assembly of Nanoparticles and Macromolecules
R(q, c) ) Rstd
( )
I(c) - Is n ITOL nTOL
2
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(1)
where Rstd and nTol are the standard Rayleigh ratio (31.6 × 10-6 cm-1 at 488 nm) and refractive index of toluene, and Is and ITol the intensities measured for the solvent and for the toluene in the same scattering configuration. To accurately determine the size of the colloidal species, dynamic light scattering (DLS) was performed with concentration ranging from c ) 0.01 to 1 wt %. In this range, the diffusion coefficient varies according to D(c) ) D0(1 + D2c), where D0 is the self-diffusion coefficient and D2 is a virial coefficient of the series expansion. The sign of the virial coefficient, the type of interactions between the aggregates, either repulsive or attractive can be deduced. From the value of D(c) extrapolated at c ) 0 (noted D0), the hydrodynamic radius of the colloids is calculated according to the Stokes-Einstein relation, DH ) (kBT/3πηD0), where kB is the Boltzmann constant, T is the temperature (T ) 298 K), and ηS (ηS ) 0.89 × 10-3 Pa · s) is the solvent viscosity. The autocorrelation functions of the scattered light are interpreted by using the CONTIN fitting procedure. Neutron Scattering. Small angle neutron scattering (SANS) experiments were performed on the PAXY spectrometer at Laboratoire Leon Brillouin (LLB, Saclay, France). Two configurations are used (D ) 1.35 and 6.70 m, both at λ ) 6 Å), covering a q-range from 5 × 10-3 to 0.2 Å-1. Exposure times of 2 and 1 h for the small and large angle configuration, respectively, are necessary to obtain good statistics. Raw data are radially averaged. Standard corrections for sample volume, neutron beam transmission, empty cell signal subtraction, and detector efficiency have been applied to obtain the scattered intensities with use of standard SANS procedures27 yielding the neutron scattering cross section (expressed in cm-1). The incoherent background arising from the hydrogen atoms was calculated by using test solutions containing a mixture of H2O and D2O. Cryo-TEM. Cryo-transmission electron microscopy (cryoTEM) was performed on hybrid complexes prepared at concentration c ) 0.1 wt %. For the experiments, a drop of the solution was put on a TEM-grid covered by a 100 nm thick polymer perforated membrane. The drop was blotted with filter paper and the grid was quenched rapidly in liquid ethane in order to avoid the crystallization of the aqueous phase. The membrane was then transferred into the vacuum column of a TEM microscope (JEOL 1200 EX operating at 120 kV) maintained at the temperature of liquid nitrogen. The magnification for the cryo-TEM experiments was selected at 40 000×. Optical Reflectometry. The amount of adsorbed complexes issued from different formulation processes onto poly(styrene) (PS)28 surface was monitored by using stagnation point adsorption reflectometry (SPAR). A complete description of this device developed by Wageningen University (Netherlands) can be found in ref 29. Fixed angle reflectometry measures the reflectance at the Brewster angle on the flat substrate. A linearly polarized light beam is reflected by the surface and subsequently split into a parallel and a perpendicular component with use of a polarizing beam splitter. As material adsorbed at the substrate-solution interface, the intensity ratio S between the parallel and perpendicular components of the reflected light is varied. The change in S is related to the adsorbed amount through
Γ(t) )
1 S(t) - S0 As S0
(2)
where S0 is the signal from the bare surface prior to adsorption. According to this model, the sensitivity factor (As), which is the relative change in the output signal S per unit surface, was found to be proportional to dn/dc and very weakly dependent upon the amount of material adsorbed. In practice, it was regarded as a constant. Furthermore, good accuracy and repeatability were obtained when As is larger than 0.005 m2/mg. Hydrophilic silica substrates were modeled by using smooth silicon wafers covered with a layer of 100 nm SiO2 in order to maximize the reflectometer signal. Hydrophobic poly(styrene) (PS) substrate was modeled by a PS thin layer of 100 nm deposited on top of an HMDS (hexamethyldisilizane)-functionalized silicon wafer by spin-coating a toluene solution (2.5 wt %) at 5000 rpm. The final PS layer thickness was around 100 nm to ensure a good sensitivity and presented water contact angles around 88° typical for PS-coated material. The sensitivity factor As was found to range between 0.02 and 0.035 m2/mg for silica surfaces, and between 0.017 and 0.025 m2/mg for PS surfaces. Results and Discussion Manual Formulation. In a previous publication the influence of the mixing pathway on the CeO2/PSS11K-b-PAM30K system interacting through a combination of electrostatic and H-bonding interactions1 was investigated. In the current work, the influence of the interaction itself on the formulation process was put under scrutiny on the CeO2-PAA2K/PTEA11K-b-PAM30K system interacting solely through electrostatic interactions. As preliminary experiments, we first investigated any possible influence of the mixing pathway on the final morphology of the complexes. Different combinations of mixing orders (CeO2PAA2K into PTEA11K-b-PAM30K or PTEA11K-b-PAM30K into CeO2-PAA2K) and homogenization speeds (high speed injection or pouring) were performed and DLS measurements were conducted on the final solutions to estimate the size of the generated structures (Figure 1). DLS results did not show large differences in the final size or size distribution of the complexes issued from the different mixing processes, however. The insensitivity to the mixing pathway for the CeO2-PAA2K/ PTEA11K-b-PAM30K complexes is due to the very low concentration of the stock solutions (e0.1 wt %). Indeed, only electrostatic interactions are present here and the charged block represents only one-third (by weight) of the polymer chains. When the density of “reactive sites” (positive charges) is rather low, the “reaction time scale” is longer and more comparable to ordinary mixing time scales (e.g., pouring one solution into another one); the morphology of the final complexes becomes hence less sensitive to the mixing pathway. At higher concentration (1 wt % and up), however, the mixing stage will indeed influence the formulation features a lot. Instead of tuning the complexation processes via the mixing stage while keeping the interaction constant between the components (see former publication1), we follow here an alternate approach based on the concept of a dormant-reactiVe solution in which both building blocks are not initially interacting with each other in the aqueous solution. This process, inspired by molecular biology, was originally developed for in vitro reconstitutions of chromatin;the DNA/histones macromolecular substance that forms the chromosomes of our cells30,31 and consists of two steps. In the first step the components are
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Figure 1. Hydrodynamic radius (RH) distribution of CeO2-PAA2K/ PTEA11K-b-PAM30K complexes prepared via different formulation routes.
mixed together in a high ionic strength aqueous solution. The salt screens the electrostatic interactions between the individual components preventing them from forming aggregates. In the second step, the salt is removed progressively by dialysis or by dilution effectively triggering the coassembly process. Desalting kinetics results were very recently published on the system of salted (NH4Cl) solutions of CeO2-PAA and PTEAb-PAM.26 Both the particles and the polymers were shown to be stable in the presence of 1 M NH4Cl. When the two salted solutions were mixed together, no association occurred due to the charge screening effect. DI water was then added stepwise at the average flow rate of 0.3 µL/s until the final salt concentration reached 10 mM. The aggregation of the polymer/ nanoparticle system was monitored by static and dynamic light scattering. Figure 2 (left) shows the hydrodynamic diameter as a function of the ionic strength of the solution. An abrupt transition was observed at a critical ionic strength estimated here at Ib ) 0.43 M. As evidenced by cryo-TEM (insets), the electrostatically screened polymer and nanoparticle system underwent an abrupt transition between an unassociated and a cluster state. By fine-tuning the desalting kinetics, the size of
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Figure 3. Scattering intensity and hydrodynamic diameter RH as a function of the “bulk” ionic strength I (and the corresponding overall nanoparticle + polymer concentration c) for the CeO2-PAA2k/ PDADMAC100K (50/50 ) wt/wt) system at c ) 0.1 wt %. Ib ≈ 0.5 M is the critical ionic strength above which the electrostatic interaction between the particles and the polyelectrolytes in the bulk is turned off.
the clusters was varied from 100 nm to over 1 µm. Cryo-TEM analysis showed that with the “desalting” route, the structures formed were much larger (RH ≈ 200 nm vs 30 nm from direct mixing in salt free condition at c ) 0.1 wt %) and rather spherical (vs “frozen” irregular shape). The CeO2-PAA2k/PDADMAC100k system where the chargedneutral block copolymer PTEA11K-b-PAM30K was replaced with a cationic homopolyelectrolyte was then investigated. An abrupt transition was equally observed via light scattering measurements (Figure 3) with a critical salt concentration around Ib ≈ 0.5 M. An ionic strength similar to that of the previous system suggesting likely that the transition triggered by the salt is controlled by the particle size and charge as predicted theoretically by Muthukumar et al.32 and via MC simulations by Stoll et al.33,34 Neutron scattering experiments (Figure 4) were then performed on these differently generated coacervates. In the direct
Figure 2. Ionic strength dependence of the hydrodynamic diameter for a dispersion containing CeO2-PAA2K particles and oppositely charged PTEA11K-b-PAM30K block copolymers (closed symbols). The dispersions were obtained by dilution. With decreasing ionic strength, an abrupt transition was observed at the critical value of Ib ) 0.43 M. The open symbols represent the hydrodynamic diameters for CeO2-PAA2K (circles) and PTEA11K-b-PAM30K (squares), respectively. Inset: Cryo-TEM images of particles and clusters apart from the transition. Upper right: Cryo-TEM analysis of CeO2-PAA2K/PTEA11K-b-PAM30K coacervates prepared via direct mixing (top).
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Figure 4. SANS cross sections of CeO2-PAA2K/PTEA11K-b-PAM30K hybrid complexes formulated in different ways: direct mixing in DI water (open circles) or dialyzed from salted solution (black diamonds). For comparison, the complexes made with CeO2-PAA2K nanoparticles and PDADMAC homopolyelectrolytes prepared by dialysis are also shown (black circles).
Figure 5. Adsorption onto silica of the CeO2-PAA2K/PTEA11K-b-PAM30K system monitored by optical reflectometry and subsequent evaluation of lysozyme adsorption onto the treated surface. Inset: Lysozyme adsorption onto a bare silica surface.
mixing case, frozen core-shell coacervates (RH ≈ 50 nm) were formed under a strong electrostatic interaction as expected,2,28,35 with cores composed of clusters of nanoparticles wrapped by PTEA11K blocks. The structure of densely packed particles in the core was evidenced by the appearance of a scattering peak at q ) 0.075 Å-1, corresponding to an interparticle distance of 8.4 nm, the dimension of CeO2-PAA2K particles.2 It should be noted that direct mixing is here equivalent to a quench in the desalting method as pointed out recently by Fresnais et al.26 In the case of a slow dialysis, much larger (RH ≈ 300 nm) spherical aggregates were built up under a weaker interaction. Because the extended contour length of the copolymer chains (∼25 nm) is shorter than the size of such aggregates, a core-shell structure cannot be obtained. The coacervate is then a mixture of neutral PAM and charged PTEA chains and CeO2 nanoparticles. No discernible nanostructure signature was observed in the case of the dialysis. The disappearance of the scattering correlation peak might be due either to the presence of the neutral part of the diblock throughout the spherical complexes leading to a dilution of the nanoparticles packing or to a lower contrast as suggested by a lower scattered intensity. To assess this hypothesis the block copolymer was replaced by a cationic homopolyelectrolyte. Very large aggregates were generated (which eventually sedimented) giving rise to a very similar yet more pronounced correlation
peak in the scattering intensity, a signature of densely packed nanoparticles wrapped by the cationic homopolyelectrolytes. Surface Modifications. Reflectometry experiments were performed with fresh solutions at 0.1 wt % prepared through different processes (direct mixing and dialysis-desalting) onto silica and polystyrene surfaces. DI water was used to obtain the baseline and a lysozyme protein solution (0.01 g/L) was used to evaluate any subsequent antifouling effect. Figures 5 and 6 show the adsorption of the complexes, DI water rinsing, and protein adsorption stages. It can be seen that on both silica and PS surfaces, the saturated amount of adsorption is higher in the case of dialyzed structures (2.48 mg/m2 vs 1.05 mg/m2 for silica; 1 mg/m2 vs 0.7 mg/m2 for PS). The antifouling efficiency is pretty good in both cases, however. Lysozyme adsorption was negligible on the treated silica surface. On the treated PS surface, the lysozyme adsorption is largely reduced compared to that of bare PS (0.38 mg/m2 vs 1.02 mg/m2). Furthermore it can be totally removed by a gentle water rinsing. Adsorption kinetics similarities and differences were observed on different surfaces. In the case of silica surfaces, both structures adsorb very quickly at the beginning up to Γ ≈ 1.1 mg/m2, with a sharp increase in adsorption (Figure 5). The “direct mixing” adsorption then reaches a plateau while the “dialysis” goes through a step before reaching a new plateau
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Figure 6. Adsorption onto PS of the CeO2-PAA2K/PTEA11K-b-PAM30K system monitored by optical reflectometry and subsequent evaluation of lysozyme adsorption onto the treated surface. Inset: Lysozyme adsorption onto a bare PS surface.
Figure 7. Variation of Rayleigh ratio Rθ as a function of the volume ratio X for CeO2-PAA2k/PTEA11K-b-PAM30K coacervates obtained manually and with the micromixer. Inset left: Schematic view of a simple micromixer. Inset right: Dilution curve of a 1 wt % CeO2-PAA2k via both manual and micromixer mixing.
twice higher (Γ ) 2.48 mg/m2). The “step” is likely due to the polydispersity of the complexes (bimodal distribution as seen by DLS) leading to two different adsorption kinetics. The smaller structures (RH ) 65 nm) made via dialysis and direct mixing have a faster adsorption kinetics than the larger ones (RH ) 205 nm) as expected. This feature was not observed, however, when the adsorption occurred on PS surfaces (Figure 6). It seems that both small and large structures adsorbed much more slowly than in the case of silica surfaces with no visible adsorption steps. Toward Better-Controlled Mixing Processes. The previous results highlighted the possibility of developing different morphologies and surface properties from a limited number of chemicals by tuning some formulation parameters: a crucial point for economical and environmental reasons and toward nowadays strict regulatory requirements during new products development. Dialysis-desalting processes are reproducible. But a longer associated processing time scale does not necessarily meet industrial constraints (couple of hours for dialysis). Direct
mixing processes do not have such drawbacks, but they are very sensitive to experimental conditions such as the mixing pathway.1 Both approaches call then for more efficient and better controlled processes. Simple Micromixer. In the micromixer configuration, the mixing volume ratio X should equal the ratio between the flow rates. Compared with the previous “manual” formulation, this well-controlled process makes possible the building of a continuous phase diagram for a given system in a very short time. We made a test experiment with a 1 wt % CeO2-PAA2K solution to verify this hypothesis. The mixing process is here a simple dilution of the CeO2-PAA2K stock solution. We chose three different flow rates VCeO2-PAA2K/VH2O ) 120/480, 300/300, and 420/180 (with the overall flow rate kept constant at 600 mL/h). We then measured the Rayleigh ratio Rθ of the mixture prepared at each different rate by light scattering. Mixtures with the same X values were also prepared manually and the corresponding Rθ(c) were measured. The micromixer results matched perfectly the manual reference curve as seen in the
Assembly of Nanoparticles and Macromolecules
Figure 8. Schematic representation of the microfluidic chip coupled with online DLS.36 It consists of a 1 cm Y-shaped microchannel of height 40 µm, which drives two reactants into a 1 cm long channel of width w ) 200 µm. Chaotic mixing37 in such a laminar flow is performed with grooves of height 20 µm engineered on the main channel. DLS measurements are performed in the large channel (observation window 500 × 500 µm2) at an apparent angle θ ) 60°.
inset of Figure 7. X is hence equal to the flow rate ratio. Furthermore, we plotted the CeO2-PAA2k/PTEA11K-b-PAM30K phase diagrams at 0.1 wt % obtained both manually and with the micromixer. Clearly both approaches are in very good agreement (Figure 7). Generally the dilution curves can help us determine several important bulk parameters for a given system, such as the phase diagram, the coacervate critical aggregation concentration (CAC), the diffusion coefficient of the particles at c f 0, etc. With a micromixer, the “mixing order” parameter is not pertinent anymore. The two initial solutions are supposed to come into contact always at the same volume ratio X (small volume). This process should then minimize any gradient during the mixing and be similar to the “high speed injection” process.1 Experimentally, however, the flow rate and the stirring speed will influence the final result. For example, when the flow rate is too high or the stirring speed is too low, both feeding solutions will not mix inside the chamber postponing the complexation
J. Phys. Chem. C, Vol. 114, No. 39, 2010 16379 further down in the outlet tube generating large polydisperse aggregates. One needs definitely to pay specific attention to the hydrodynamic conditions during the complexation. Microfluidic Chip Coupled with DLS. Recently Destremaut et al.36 have developed a dynamic light scattering setup around a microfluidic chip enabling the measurement of the size of Brownian scatterers flowing in a PDMS-based microchannel. This particular chip can mix two reactants in 200 ms, and allows size measurements using DLS at about 300 ms after complete mixing. It is typically possible to measure sizes up to ∼500 nm at a flow rate of Q ≈ 2 mL/h in a microchannel of crosssection h × w ≈ 500 × 500 µm2 (Figure 8). Besides the control of the transport phenomena, the development of such laboratories on chip offers new interesting possibilities such as the high-throughput screening at the nL scale like the continuous monitoring of the viscosity of a twofluid mixture for example. This setup was used here to generate and characterize the complexation of the CeO2-PAA2k nanoparticles with both the block copolymers (PTEA11k-PAM30K) and the homopolyelectrolytes (PDADMAC100K) via the desalting-dilution approach. CeO2-PAA2K/PTEA11K-b-PAM30K. If the two nonsalted stock solutions (c ranging from 0.1 or 1 wt %) are injected into the chip, the interaction is so strong and rapid that the complexation happens before the two species are fully mixed leading to the formation of large frozen fractal structures. To avoid local inhomogeneities and to weaken the interaction, individual solutions of nanoparticles and polymers were mixed in the presence of 0.6 M NH4Cl, where the interaction is totally screened. A homogeneous solution was easily obtained as described previously. Both the dormant solution and DI water were then injected via the two entries into the chip. The interaction can in this manner be fine-tuned by adjusting the flow rate of both feeding solutions. Keeping the total flow rate constant at 300 µL/h, we tuned the ratio between the dormant solution and the DI water rate, and monitored the formed complexes with the online DLS. By plotting the scattering light intensity and the hydrodynamic radius RH vs the ionic strength I or the “mass fraction of colloids” (equals the ratio “rate of the premixed solution”/“rate of the premixed solution + DI water”), we obtained the phase diagram of Figure 9. It is a
Figure 9. Light scattering intensity (Int) and hydrodynamic radius (RH) as a function of the ionic strength (I) and the corresponding colloid mass fraction through online DLS measurements coupled with microfluidics chip for CeO2-PAA/PTEA11K-b-PAM30K system. The ionic strength at which an abrupt transition between an unassociated and a clustered state is observed at I ≈ 0.42 M. Inset: Example of a typical DLS correlation function taken in the microchip.
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Figure 10. Light scattering intensity (Int) and hydrodynamic radius (RH) as a function of the ionic strength (I) and the corresponding particle fraction through online DLS measurements coupled with microfluidics chip for the CeO2/PDADMAC system. The transition occurs at I ≈ 0.45 M.
typical phase diagram for electrostatic complexation with the presence of a peak around I ) 0.40 M. Unassociated nanoparticles and polymers are present in the solution at I > 0.42 M, and coacervates of ∼100 nm are formed at I < 0.40 M. In both cases, the intensity varies linearly with the colloid fraction (and ionic strength) following a simple dilution law. A transition zone was observed at I ) 0.40 to 0.42 M, where the interaction was switched back on but remained very weak. Some polydisperse structures were likely to be formed in such condition. A slow kinetics implies furthermore that not all the individual components can be integrated in the complexes within the observation time scale (∼500 ms) of the microfluidic chip, leading to an apparent lower scattered intensity. Interestingly, the critical ionic strength Ib (0.42 M) found in this phase diagram was very similar to one found “manually” by Fresnais et al.26 on the same system (Figure 2), which highlights the potential of such microfluidic tools for investigating interacting systems. CeO2-PAA2K/PDADMAC100K. The stock solutions were prepared and mixed at X ) 1 in the presence of 0.8 M NH4Cl, where the electrostatic interaction is totally screened for this system (see Figure 3). The phase diagram shown in Figure 10 is very similar to that of the CeO2-PAA2K/PTEA11K-b-PAM30K system. A peak in the intensity and a critical ionic strength Ib were found at 0.4 and 0.45 M, respectively, in agreement with manual off-line DLS measurements shown in Figure 3. Charged complexes were obtained with a size around 140 nm, a result different from the manual formulation in which much larger (∼>1000 nm) and polydisperse aggregates were generated due to a relatively slow dilution stage (drop-by-drop addition of DI water in the vial test). This drawback was avoided with the microfluidics formulation, where a minute amount of materials and a specific mixing geometry enabled a better and faster homogenization. These experiments showed that microfluidics setup coupled with DLS is a powerful tool for generating phase diagrams for electrostatic coassembly of nanoparticles and different charged macromolecules in the presence of a variable ionic strength. This microfluidics approach enables fast, reproducible, and accurate results to be obtained with a minute amount of materials: a key advantage over manual formulation in vial tests.
Conclusions and Outlook Different formulation pathways and mixing protocols were investigated in three coassembled systems composed of cerium oxide based nanoparticles and oppositely charged double hydrophilic diblock copolymers or homopolyelectrolytes. Under a strong driving force like the electrostatic interaction (along with other interactions like hydrogen bonding etc.), the key factor that controls the final morphology of the complexes (size, structure, and polydispersity) is the competition between the reaction and the mixing time needed to homogenize the formulation. Two approaches were put under scrutiny to shed some light on the crucial role of the formulation process: (i) Tuning the mixing stage (or the way individual components come into intimate contact) with specific protocols or tools was investigated in a former publication.1 (ii) Tuning the strength of the (electrostatic) interaction by varying the ionic strength in the initial stock solutions containing each individual building block was investigated in the current publication. The resulting structures from various formulation processes were characterized by cryo-TEM, AFM, light, and neutron scattering techniques. Some bulk and surface properties were also evaluated via rheology, wetting, and optical reflectometry measurements. Better controlled mixing processes were put forward at the end to improve the productivity and reproducibility of the complexes generation. In particular, a µ-fluidic chip coupled with dynamic light scattering was used to better control the hydrodynamics of the complexation process. The impact of the formulation pathway clearly evidenced the resulting complexes morphologies at different length scale leading to different bulk and surfaces properties. All these results suggest that a process-dependent formulation seen a priori as a drawback can be turned into an advantage: different properties can be developed from different morphologies while keeping the chemistry constant, certainly a key advantage for any product development in today’s strictly regulated world. References and Notes (1) Qi, L.; Fresnais, J.; Berret, J.-F.; Castaing, J.-C.; Grillo, I.; Chapel, J.-P. J. Phys. Chem. C 2010, 114 (30), 12870–12877.
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