Letter pubs.acs.org/macroletters
Stabilization of Water-in-Water Emulsions by Nanorods Karthik R. Peddireddy,† Taco Nicolai,*,† Lazhar Benyahia,† and Isabelle Capron‡ †
LUNAM Université du Maine, IMMM UMR-CNRS 6283, 72085 Cedex 9 Le Mans, France UR1268 Biopolymères, Interactions et Assemblages, INRA, F-44316 Nantes, France
‡
ABSTRACT: Water-in-water (W/W) emulsions formed by mixing incompatible water-soluble polymers cannot be stabilized with molecular surfactants. However, they can be stabilized by particles through the so-called Pickering effect. Recently, it was shown that its stabilization can be achieved also with nanoplates. Here, we show for the first time that even nanorods in the form of cellulose nanocrystals (CNCs) can efficiently stabilize W/W emulsions. Static light scattering and confocal microscopy techniques were used to determine the surface coverage by CNCs. In the presence of 50 mM NaCl very weak gels were formed by excess CNCs in the continuous phase. In this way creaming of the dispersed phase could be arrested. The nontoxicity, sustainability, and low cost of CNCs and the abundant availability of cellulose render these nanorods potentially highly suited for preparing W/W emulsions.
A
ΔG(disc) = −πR2γ(1 − |cos θ|)
water-in-water (W/W) emulsion is commonly formed when two incompatible hydrophilic polymers are mixed in water above certain threshold concentrations with each phase enriched with one of the polymers.1 W/W emulsions play an important role in different areas such as green chemistry,2 cell biology,1b and food.1c,3 Compared to oil-in-water (O/W) emulsions, W/W emulsions have very low interfacial tensions (1 μN/m to 1000 μN/m) and a large interfacial thickness (at least several nanometers). Therefore, molecular surfactants cannot stabilize these emulsions. Until recently, the only way to avoid macroscopic phase separation was by gelling one or both phases. However, it has recently been shown that colloidal particles can stabilize W/W emulsions by forming a layer at the interface, which reduces the free energy.4 The stabilization of interfaces with particles is known as the Pickering effect and has been intensively investigated for O/W emulsions.3,5 The change of the free energy (ΔG) by particle adsorption depends strongly on the radius of the particles (R), the contact angle (θ), and the interfacial tension (γ) of the system ΔG(sphere) = −πR2γ(1 − |cos θ|)2
Here we address the question whether even nanorods can stabilize W/W emulsions. We investigated this issue using cellulose nanocrystals (CNCs) that have a highly anisotropic rectangular parallelepiped structure with average dimensions 160 nm × 6 nm × 6 nm (see Figure 1a). CNC is a very
Figure 1. (a) Transmission electron microscopy (TEM) image of CNCs. (b) Phase diagram of the polymers used for the experiments adapted from ref 4a. The solid line indicates the binodal, and the dashed lines indicate tie lines. The colored symbols indicate the compositions of the emulsions studied here.
(1)
For example if R = 100 nm, θ = 90°, and γ = 10 μN/m, the reduction in the free energy due to the adsorption of a particle at the interface is 75 kT. This shows that adsorption of the particles can be practically irreversible even for W/W interfaces. Recently, it has been shown that also gibbsite discs with a radius of 170 nm and a thickness of 7 nm adsorb at the interface and can stabilize W/W emulsions.4d As the thickness of these discs is comparable with the interfacial thickness, the interfacial area covered by the disks is independent of the contact angle. It was shown by Vis et al.4d that the gain in free energy for the adsorption of a disc with radius R is given by © XXXX American Chemical Society
(2)
promising material for applications, especially in the fields of packaging and healthcare due to its biodegradability and nontoxicity.2 The model W/W emulsion that was used consisted of mixtures of dextran and poly(ethylene oxide) (PEO). Received: December 30, 2015 Accepted: February 5, 2016
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DOI: 10.1021/acsmacrolett.5b00953 ACS Macro Lett. 2016, 5, 283−286
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ACS Macro Letters The phase diagram of the system used for this investigation was reported in ref 4a and is reproduced in Figure 1b. The critical point is situated at 1 wt % PEO and 1.7 wt % dextran, and at the polymer compositions used for the present study phase separation is practically complete. Depending on the composition, emulsions are formed in which droplets of the dextran phase are dispersed in a continuous PEO phase (D/P emulsions) or vice versa (P/D emulsions). The interfacial tension was shown to increase with the tie line length following a power law.4a We have studied D/P and P/D emulsions on the same tie-line for which γ = 75 μN/m at compositions indicated in Figure 1b. Figure 2 shows confocal laser scanning microscopy (CLSM) images of dextran droplets in PEO (D/P) and PEO droplets in
Figure 3. (a) P/D emulsions at steady state with the same polymer composition indicated by the violet square in Figure 1b and φ = 0.42 and different CNC concentrations between 0 and 4 g/L as indicated in the figure. (b) CLSM image (160 μm × 160 μm) of the interface between the creamed emulsion containing 0.5 g/L of CNC and the destabilized PEO top phase. The red color indicates the location of Nile blue A labeled CNC particles.
emulsion within a few minutes. Subsequently the stabilized PEO droplets creamed slowly until they formed a dense layer. The volume of the pure PEO phase layer decreased with increasing CNC concentration up until C = 1.0 g/L but remained constant at higher concentrations. For C > 1.0 g/L, an excess of CNC remained in the continuous dextran phase, causing an increasing turbidity of this phase with increasing CNC concentration. These observations suggest that CNC takes a few minutes to form an effective barrier at the interface and that the fraction of PEO droplets that coalesces into a pure PEO layer within this time no longer depends on the CNC concentration when CNC is present in excess. The fraction of the PEO phase that rapidly formed a homogeneous layer decreased if φ was reduced and disappeared completely for φ < 0.35 as is shown in Figure 2c. This is due to the decrease of the rate of phase separation with decreasing φ, allowing more time for the CNC to form stable barriers at the P/D interfaces. Figure 3b shows an image of the creamed emulsion at the interface with the pure PEO layer. The low fluorescence intensity from the pure PEO layer and the PEO phase within the droplets demonstrates that the fraction of CNC in the PEO phase is negligible. The amount of excess CNC in the dextran phase was determined as described in the methods section. By comparing the amount of excess CNC with the total amount in the system, we can deduce the fraction of CNC that was accumulated at the interface. In Figure 4a the fraction of accumulated CNC at the interface is plotted as a function of the total CNC concentration. It is very high at low CNC concentrations but decreases rapidly for C > 1 g/L. The average droplet size was determined from CLSM images, which allowed us to calculate the coverage of the droplet surface by CNC assuming that they lie flat on the
Figure 2. (a) Completely destabilized D/P emulsion with composition indicated by the red circle in Figure 1b and φ = 0.25. (b) CLSM image (160 μm × 160 μm) of the unstable D/P emulsion taken immediately after preparation. The red color indicates the location of Nile blue A labeled CNC particles. (c) A stable P/D emulsion with composition indicated by the green triangle in Figure 1b and φ = 0.33, at steady state with a fully creamed layer of stable PEO droplets. (d) CLSM (160 μm × 160 μm) image of the creamed dense P/D emulsion layer. The concentration of CNC was in all cases 0.5 g/L.
dextran (P/D). The volume fraction of the dispersed phase was φ = 0.25 for the D/P emulsion and φ = 0.33 for the P/D emulsion. CNC was visualized by labeling with the fluorophore Nile blue A. For both types of emulsion, a layer of CNC can be clearly observed at the interface between the two phases. The P/D emulsion was stable for months, which demonstrates that W/W emulsions can be stabilized effectively by one-dimensional nanorods. Notice that we use the expression stable here to denote the absence of coalescence. Creaming of PEO droplets occurred in all cases more or less quickly. In the case of the emulsion shown in Figure 2c it took 1 month to reach steady state after which the PEO droplets had formed a layer of densely packed droplets at the top (see Figure 2d). However, the formation of the interfacial CNC layer was not sufficient to stabilize dextran droplets in the continuous PEO phase. The D/ P emulsion phase separated into a homogeneous dextran bottom phase and a homogeneous PEO top phase, with the CNC situated almost exclusively in the dextran phase (see Figure 2a). We note that sedimentation of CNC does not play a role because the gravitational length of the CNC is about 15 cm, which is much larger than the sample height. CNC prefers the dextran phase, perhaps because both contain sugar groups. It is known that dextran does not adsorb to CNC,6 and the strong preference of CNC for the dextran phase renders it unlikely that PEO adsorbs to CNC. Figure 3a shows P/D emulsions at steady state with φ = 0.42 containing different CNC concentrations up to 4 g/L. The systems at this composition indicated by the violet square in Figure 1b formed a layer of the pure PEO phase on top of the
Figure 4. (a) Fraction of CNC at the droplet interface as a function of the overall CNC concentration. (b) Effect of overall CNC concentration on the PEO droplet size (circles) and their coverage at the P/D interfaces (squares). 284
DOI: 10.1021/acsmacrolett.5b00953 ACS Macro Lett. 2016, 5, 283−286
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ACS Macro Letters
of the semidilute PEO and dextran solutions (∼2.5 nm). Nevertheless, the interfacial thickness is still comparable to the width of the CNC. Considering also that CNC strongly prefers the dextran phase it is likely that the arrangement of the CNC at the P/D interface is significantly different from that at the D/ P interface, which may explain why CNC does not stabilize P/ D emulsions. A similar asymmetric capacity to stabilize P/D or D/P interfaces was noted earlier also for spherical colloids, the origin of which still needs to be elucidated.4c In conclusion, it is shown that even nanorods can stabilize water-in-water emulsions. Due to their high anisotropy, less material is needed to stabilize W/W emulsions than when homogeneous spherical colloids are used. Moreover, even a small amount of salt is enough to initiate aggregation and gelation of excess CNC and thereby inhibits creaming. The material used for this study is nontoxic, biodegradable, and available in abundance. Therefore, it may be viable to use CNCs for large-scale applications especially for biologically sensitive emulsions.
droplet surface as was shown recently for CNC at the O/W interface. 7 Figure 4b shows the effect of total CNC concentration in the mixture on the PEO droplet size as well as on the coverage at the interface. As expected, the PEO droplet size decreased with increasing CNC concentration, but the coverage of CNCs remained constant at approximately 50%. The density at the W/W interface is less than that reported for the O/W interface (85%) stabilized by the same particles.10 One possible reason for the lower density at the W/ W interface is the effect of electrostatic repulsion between the charged CNC which will make them align side by side instead of forming in a mesh-type structure as was found at the O/W interface.8 As might be expected, the droplet size decreased also when the volume fraction of the dispersed phase was decreased at a given CNC concentration (results not shown). A consequence of the reduction of the droplet size is that creaming slows down. However, the only way to avoid creaming altogether is to gel the continuous phase. For the system studied here this can be easily done by exploiting the excess CNC in the continuous phase. CNC does not aggregate in aqueous solution due to electrostatic repulsion, but addition of a small amount of salt (50 mM NaCl) induces random aggregation of the CNC particles. 9 Gels are formed even at very low CNC concentrations (C > 3 g/L) because the CNC is highly anisotropic. Elsewhere we will discuss the salt-induced aggregation of CNC in detail and show that the kinetics and the stiffness of CNC gels can be tuned by varying the CNC concentration and the salt concentration. Figure 5 shows a
METHODS
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AUTHOR INFORMATION
The dextran (5 × 105 g/mol), PEO (2 × 105 g/mol), sodium chloride (NaCl), and Nile blue A were purchased from Sigma-Aldrich. Dextran was used as it was received, but PEO contained a small amount of silica particles, which were removed through centrifugation for 4 h at 5 × 105g. The stock aqueous solutions of dextran (30 wt %) and PEO (10 wt %) were prepared with 200 ppm of azide added in order to suppress bacterial growth. Whatman filters (grade 20 Chr) were used as the cellulose source, and CNCs were obtained by the sulfuric acid hydrolysis method as described elsewhere.8 Conductometric titration and transmission electron microscopy (TEM) methods were used to determine the surface charge density (0.16 e/nm2) and the dimensions (∼160 nm × 6 nm × 6 nm) of the CNC, respectively. The density of the CNC is 1.6 g/cm3. Further details of the preparation and characterization of CNC can be found in ref 8. The emulsions were prepared by adding all ingredients together and then mixing using a sonicator (Bioblock Scientific). The order of mixing had no effect on the emulsion behavior. The emulsions were imaged by confocal microscopy (Leica TCS-SP2). CNC was labeled with Nile blue A dye by adding 10 ppm of the dye to the stock solution of 20 g/L of CNC. The excess dye partitioned preferentially to the PEO phase. The excess CNC concentrations in the continuous dextran phases were obtained by measuring the light-scattering intensity using an ALV/CGS-3 Compact Goniometer in combination with a helium neon laser operating at 633 nm. The CNC concentration was determined by comparing the scattering intensity with a calibration curve that was obtained by measuring the scattering intensity of pure dextran solutions with known amounts of CNC.
Figure 5. CLSM image (160 μm × 160 μm) of a P/D emulsion at steady state with polymer composition indicated by the green triangle mark in Figure 1b. The CNC concentration and the NaCl concentration are 1 g/L and 50 mM, respectively. The fluorescence from the PEO phase, which does not contain CNC, is caused by excess dye.
CLSM image of a P/D emulsion with φ = 0.33 containing 1 g/ L of CNC and 50 mM NaCl. The PEO droplets are connected by clusters of CNC forming a very weak gel. At this CNC concentration, the gel is strong enough to inhibit creaming of the droplets but flows easily when tilted. In order to calculate the reduction of the free energy after adsorption on the interface as a function of the dimensions of the nanorods, eq 2 can be modified ΔG(rod) = −lbγ(1 − |cos θ|)
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Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS The research was financially supported by the MATIERES project and the “Région Pays de la Loire”. The BIBS platform of INRA Angers-Nantes and Emilie Perrin are thanked for technical support with TEM microscopy.
(3)
where l is the length and b is the width of the rods. CNC will no longer adsorb to the interfaces if the reduction in the free energy is less than kT, i.e., for γ < 5 μN/m if θ = 90° using eq 3. A detailed investigation of this prediction is currently being conducted and will be reported elsewhere. The interfacial thickness is known to diverge close to the critical point, but the emulsions studied here were far from the critical point where the correlation length is comparable to that
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REFERENCES
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