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Double Emulsion Templated Microcapsules with Single Hollow Cavities and Thickness-Controllable Shells Fei Gao,†,‡ Zhi-Guo Su,† Ping Wang,*,§ and Guang-Hui Ma*,† National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China, Graduate UniVersity of Chinese Academy of Sciences, Beijing 100049, China, and Biotechnology Institute, UniVersity of Minnesota, St. Paul, Minnesota 55108 ReceiVed December 18, 2008. ReVised Manuscript ReceiVed January 14, 2009 A novel form of microcapsules, which possess single hollow cavities and thickness-controllable shells, were prepared by a two-step emulsification, emulsion ripening, and suspension polymerization. Parameters on morphology control of water-in-oil-in-water (W/O/W) emulsion globules were particularly investigated in this study, and a universal strategy to prepare single-core water-in-oil (W/O) globules from their multicore precursors was proposed. These single-core globules were further utilized as templates for solid microcapsules by the suspension polymerization, during which the phase-separation mechanism could be employed to form nanochannels across the shells. Such microcapsules could be further exploited as microreactors with functional cores to be loaded and would be especially suitable to encage bioactive materials.
Introduction During the past three decades, polymeric capsules in microscale or in even smaller size were proved to be of crucial importance in many practical performances and have been intensively studied for the existing and promising values in the fields of industry, agriculture, health care, daily consumption, and scientific research.1,2 This study aimed to develop a novel form of polymeric microcapsules which possess capacious compartments and designable shells. These microcapsules could be further exploited as microreactors with functional materials assembled inside. Quite a number of elegant methods have been reported to prepare polymeric microcapsules suitable for a variety of core materials. However, it is still hard to find a strategy to accommodate mechanical strength, capability, permeability, and biocompatibility by a simple procedure. Traditionally, polymeric microcapsules could be fabricated by phase separation between polymers and a hydrophobic solvent during either polymerization or solvent evaporation.2-7 These polymers formed the shell, and the solvent was engulfed as a liquid core, which could easily be removed by subsequent extraction. Some other researchers employed existing microspheres as raw materials, internal compartments of which were fabricated by further thermal or * To whom correspondence should be addressed. (G.-H.M.) E-mail:
[email protected]. Phone: 8610-8262-7072. Fax: 8610-8262-7072. (P.W.) E-mail:
[email protected]. Phone: (612) 624-4792or (612) 624-3264. Fax: (612) 625-6286. † Institute of Process Engineering, Chinese Academy of Sciences. ‡ Graduate University of Chinese Academy of Sciences. § University of Minnesota. (1) McDonald, J. C.; Devon, J. M. AdV. Colloid Interface Sci. 2002, 99, 181– 213. (2) Ma, G.-H.; Su, Z.-G.; Omi, S.; Sundberg, D.; Stubbs, J. J. Colloid Interface Sci. 2003, 266, 282–294. (3) Ma, G.-H.; Chen, A.-Y.; Su, Z.-G.; Omi, S. J. Appl. Polym. Sci. 2003, 87, 244–251. (4) Okubo, M.; Minami, H. Colloid Polym. Sci. 1996, 274, 433–438. (5) Ma, G.-H.; Omi, S.; Dimonie, V. L.; Sudol, E. D.; El-Aasser, M. S. J. Appl. Polym. Sci. 2001, 85, 1530–1543. (6) Okubo, M.; Konishi, Y.; Minami, H. Prog. Colloid Polym. Sci. 2004, 124, 54–59. (7) Okubo, M.; Konishi, Y.; Inohara, T.; Minami, H. J. Appl. Polym. Sci. 2002, 86, 1087–1091.
Figure 1. Flowchart for the preparation of double emulsion templated microcapsules.
chemical treatment.1,4,8 These procedures could be well explained by thermodynamic principles and would therefore lead to easy and accurate controls. However, applications, especially the ones with biomaterials incorporated, were often restricted by the hydrophobic compartments or by the harsh preparations. To encage delicate biomolecules during some particular encapsulations, aqueous environments were often desired during assembly, such as the procedure for calcium alginate capsules. Although these gel-based capsules could be prepared through a simple and mild procedure in water solutions, they usually suffered a lot from soft texture and poor stability in practice.9 Another widely applied approach to encapsulate water cores is the two-step emulsification, producing liquid microcapsules such as water-in-oil-in-water (W/O/W) emulsion globules, which have been studied since as early as 1925.10 With combination of this approach with subsequent solidification, such as suspension polymerization or solvent evaporation,11-13 liquid globules in W/O/W could be transformed into solid microcapsules. For the biocompatible compartments and controllable release behaviors, a variety of applications have shown great interest in such multiple (8) Im, S. H.; Jeong, U.; Xia, Y. Nat. Mater. 2005, 4, 671–675. (9) George, M.; Abraham, T. E. J. Controlled Release 2006, 114, 1–14. (10) Seifriz, W. J. Phys. Chem. 1925, 29, 738–749. (11) Zydowicz, N.; Nzimba-Ganyanad, E.; Zydowicz, N. Polym. Bull. 2002, 47, 457–463. (12) Liu, R.; Ma, G.; Meng, F.-T.; Su, Z.-G. J. Controlled Release 2005, 103, 31–43. (13) Meng, F. T.; Ma, G. H.; Qiu, W.; Su, Z. G. J. Controlled Release 2003, 91, 407–416.
10.1021/la804173b CCC: $40.75 2009 American Chemical Society Published on Web 02/19/2009
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Figure 2. Images of the W/O/W emulsion with different ripening times: a, 5 min; b, 30 min; c, 90 min; d, 180 min.
Figure 3. Schematic representation of ripening of W1/O globules. Table 1. Typical Recipe for Preparation of Double Emulsion Templated Microcapsules phase/ingredient
amount (g)
(W1) internal aqueous phase PVA Tween 20 NaCl (O) oil phase MMA EGDMA TMA Span 85 HA + chloroform (1.01 g/mL in density) ADVN (W2-1) initial part of the external aqueous phase SDS PVA NaCl (W2-2) additional part of the external aqueous phase PVA
0.5 0.005 0.005 0.001-0.005 3.5-5.5 1.0 1.5 0.5 0.5 0-2.0 0.03 30 0.03 0.75 0.088 120 3.0-4.8
systems.14 Although a rather large literature existed, there was still a lack of general formulas or quantitative criteria on the multiple emulsions,15 and the preparations were therefore often determined empirically and varied from case to case. Generally, in preparing a double emulsion, the primary emulsification should be more intensive than the secondary one, leading to each oil globule containing quite a number of water droplets. This was the reason why the freshly obtained globules were usually endowed with a multiple-core morphology, which was believed to be the most stable form and an ideal style for controlled release.16 However, multicell microcapsules templated by multicore globules would be short in volume of each compartment and would slow mass transportation across the shells. In this paper, we present a novel type of hollow microcapsule, taking advantage of both the biocompatibility of the W/O/W and the rigid polymeric texture obtained from suspension polymerization. Morphologies of the microcapsules were determined by their W/O/W globule templates. As a metastable system, a double emulsion tends to collapse into a more stable form, an (14) Engel, R. H.; Riggi, S. J.; Fahrenbach, M. J. Nature 1968, 219, 856–857. (15) Ficheux, M.-F.; Bonakdar, L.; Leal-Calderon, F.; Bibette, J. Langmuir 1998, 14, 2702–2706. (16) Florence, A. T.; Whitehill, D. J. Colloid Interface Sci. 1981, 79, 243–256.
Figure 4. Example ternary proportion diagram exhibiting evolution profiles of W1/O/W2 emulsions: (b, 9) morphology distribution of the three categories with a certain recipe and ripening time; (∆) evolution direction with time lapsing. The only difference among the dots in each curve is the ripening time. Example images attached aside exhibit the exclusive morphology of the three different forms.
O/W or a W/O simple emulsion.15 It was found that, under the given conditions, entrapped water (W1) droplets prefer to coalesce with each other than to escape to the external aqueous phase (W2). Such a preference would lead to a majority proportion of single-core globules in a unique process of evolution, which we named “emulsion ripening”. Parameters affecting such evolution were particularly investigated, and an effective strategy was first proposed to prepare single-core globules and single-cell microcapsules templated from such globules. To our best knowledge, it was the first attempt to manipulate the morphology of the W/O/W globules by guiding their inherent destruction and to produce single-core globules from their multicore precursors. Previously, such single-core globules could only be fabricated one by one in microfluidic devices.17 On the basis of single-core globules, the thickness of oil membranes could be designed by an osmotic gradient between W1 and W2, which would also determine the shell thickness of the final microcapsules. In addition, the phase-separation mechanism was employed to produce nanochannels across the shells, with midchain alcohols employed as the primary porogens.18-22 Besides acrylate monomers, cross-linkers which possess more than one vinyl group of each molecule were essentially employed to form porous and rigid shells. The eventual polymerization of the oil membranes was initiated by ultraviolet radiation with a wavelength of 380 nm, as had also been proved compatible for biomolecules.23
Experimental Section Materials. Methyl methacrylate (MMA) (Beijing Chemical Reagents Co.), ethylene dimethacrylate (EDMA) (Sigma-Aldrich), and trimethylolpropane trimethacrylate (TMA) (Sigma-Aldrich) were (17) Shum, H. C.; Lee, D.; Yoon, I.; Kodger, T.; Weitz, D. A. Langmuir 2008, 24, 7651–7653. (18) Omi, S.; Katami, K. i.; Taguchi, T.; Kaneko, K.; Iso, M. Macromol. Symp. 1995, 92, 309–320. (19) Omi, S.; Katami, K. i.; Taguchi, T.; Kazuyoshi, K.; iso, M. J. Appl. Polym. Sci. 1995, 57, 1013–1024. (20) Omi, S. Colloids Surf., A 1996, 109, 97–107. (21) Zhou, W.-Q.; Gu, T.-Y.; Su, Z.-G.; Ma, G.-H. Eur. Polym. J. 2007, 43, 4493–4502. (22) Wang, R.-W.; Zhang, Y.; Ma, G.-H.; Su, Z.-G. Colloids Surf., B 2006, 51, 93–99. (23) Wang, P.; Sergeeva, M. V.; Lim, L.; Dordick, J. S. Nat. Biotechnol. 1997, 15, 789–793.
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Figure 5. Evolution profiles of microcapsule morphologies with different PVA concentrations in W2: (1, 0) 2.5%, (b, O) 1.5%, ([, ]) 0.5%. The arrows indicate the evolution direction. The left-end dots of each curve represent the beginning morphologies with no ripening; each solid dot followed cost 60 min more in ripening, while each hollow dot cost only 30 min more. OM pictures attached aside record the typical morphology evolution with 2.5% PVA in W2.
Figure 6. Evolution profiles of microcapsule morphologies fabricated with different PVA concentrations in W1: (b, O) 0.0%, (9, 0) 1.0%, (2, 0) 2.0%. The arrows indicate the evolution direction. The left-end dots of each curve represent the beginning morphologies with no ripening; each solid dot followed cost 60 min more in ripening, while each hollow dot cost only 30 min more. OM pictures attached aside record the typical morphology evolution with 1.0% PVA in W1.
Figure 7. Evolution profiles of microcapsule morphologies fabricated with different NaCl concentrations in W2: (b, O) 0 mM, (9, 0) 10 mM, (2, 0) 13 mM. The arrows indicate the evolution direction. The left-end dots of each curve represent the beginning morphologies with no ripening; each solid dot followed cost 60 min more in ripening, while each hollow dot cost 30 min more. OM pictures attached aside show the typical morphology evolution with no NaCl in W2.
distilled under reduced pressure to remove the inhibitor. 2,2′Azobis(2,4-dimethylvaleronitrile) (ADVN; V65) was analysis grade and was used as an initiator. Hexanol (HA), octanol (OA), heptane (HP), and chloroform were all analysis grade and were purchased from Beijing Chemical Reagents Co. Sorbitan trioleate (Span 85)
(Shanghai Chemical Reagent Co.) was reagent grade. Poly(oxyethylene) sorbitan monooleate (Tween 20) was provided by Kishida Chemical Co. (Osaka, Japan). Poly(vinyl alcohol) (PVA-217; degree of polymerization 1700, degree of hydrolysis 88.5%, Kuraray) was used as a stabilizer. Sodium dodecyl sulfate (SDS) was of the grade
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Figure 8. Evolution profiles of microcapsule morphologies fabricated with different NaCl concentrations in W1: ([, ]) 45 mM, (9, 0) 60 mM, (1, 0) 120 mM. The arrows indicate the evolution direction. The left-end dots of each curve represent the beginning morphologies with no ripening; each solid dot followed cost 60 min more in ripening, while each hollow dot cost only 30 min more. OM pictures attached aside show the typical morphology evolution with 120 mM NaCl in W1.
Figure 9. Shell thicknesses of single-hollow microcapsules obtained with different NaCl concentrations in W1.
for biochemical use (Merck). NaCl was reagent grade (Beijing Chemical Reagents Co.) and was used to adjust the electrolyte concentration of the aqueous phase. Ethyl alcohol was commercial grade (Beijing Chemical Regents Co.) and was used to precipitate and wash the particles obtained. Flourescein isothiocyanate (FITC; MW 389.4) and Rhodamine B were from Beijing Xin Jing Ke Biotechnology Co. Ltd., China. Water was deionized using ionexchange resins. PMMA latex particles 390 nm in diameter, which were commonly applied as enzyme carriers, were provided by Biochemical Engineering Center (Beijing). Microcapsule Preparation. Novel microcapsules were fabricated by a two-step emulsification, emulsion ripening, and suspension polymerization (Figure 1). W1/O globules in W2 as templates of hollow microcapsules were first prepared. A typical recipe on the preparation is shown in Table 1. PVA was dissolved in both internal and external water phases as a hydrophilic stabilizer. Other low molecular weight surfactants were also employed, with Tween 20 in W1, SDS in W2, and Span 85 in the oil. NaCl was dissolved in both water phases (W1 and W2) to adjust the osmotic pressure. The density of the inner water was tested to be 1.01 g/mL at room temperature. To prevent the sinking or creaming of the entrapped droplets (W1 in O), the density of the oil phase was adjusted to the same value by a portion of chloroform. The procedure is detailed as follows. (1) Primary emulsification: A certain amount of internal aqueous phase (W1) was emulsified into an oil phase (O) by an ultrasonifier, which was conducted at 100 W for 20 s. During the primary emulsification, W1 was broken into fine droplets, less than 1 µm in diameter. (2) Secondary emulsification: The primary emulsion W1/O was further dispersed into an external water phase (W2-1, initial part of
W2) by mechanical stirring at 400-600 rpm for 120 min, and the W1/O globules in W2-1 were prepared at tens of micrometers in diameter and endowed with multicore morphologies. (3) Emulsion ripening: After the two-step emulsification, an additional part of W2 (W2-2) was added to W2-1 to adjust the concentrations of salt and stabilizer in the external phase. The emulsion was then loaded into several quartz vessels, sealed with a nitrogen atmosphere, and rotated vertically. It would cost tens of minutes to get the maximal proportion of the single-core W1/O globules. (4) Suspension polymerization: The polymerization of the oil phase was initiated by ultraviolet irradiation (380 nm in wavelength), with ADVN as the initiator. The oil globules would be solidified in a few minutes, and the monomer conversion would exceed 99% in 6 h. (5) Collection: The solidified microcapsules were collected by filtration and washed with 1% Tween 20 in a water-bath ultrasonifier to remove the stabilizer and most of the solvent. Further extraction by ethyl alcohol could remove the organic residues deeply entrapped. Characterization. The morphologies of W1/O globules were observed under an optical microscope (OM) and pictured by a digital camera at different ripening times, from a few minutes to several hours (Figure 2). After preparation, microcapsules between 30 and 50 µm in diameter, which occupied about 60% of the total products, were sieved out for morphology evaluation. According to the number of internal compartments, we sorted the microcapsules into three categories in general: multicell microcapsules (containing several to hundreds of internal compartments), single-cell microcapsules (containing only one visible compartment), and no-cell microcapsules (containing no visible compartment). The proportions of the three categories prepared with a certain recipe and after a certain evolution were summarized by observing and sorting more than 300 microcapsules. The three proportions obtained with a given recipe changed along with the evolution time, their tendency curves recording the whole process of evolution. Optical images were also employed to calculate the shell thickness of single-cell microcapsules, by deducting the internal mean diameter from the external one, with more than 300 single-cell microcapsules measured to get the mean value. Scanning electron microscopy (SEM) (JSM-6700F, JEOL, Japan) was used to observe the surface features of polymer microcapsules. An Autosorb-1 automatic surface area and pore size analyzer (Quantachrome Corp.) was used to determine the porosities of the microspheres. In the vacuum sorption system, ultrapure nitrogen gas and liquid nitrogen gas were used as the adsorbate and coolant. The specific surface area was determined with the intrusion and extrusion curves by the multipoint Brunauer-Emmett-Teller (BET) method. Average pore diameters were determined by the Barrett-Joyner-Halenda (BJH)
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Figure 10. SEM images of hollow microspheres in a dry condition: A, nonporous shell of the microcapsule; B, porous shell of the microcapsule; C, a single-cavity microcapsule; D, detailed image of the porous surface of a hollow capsule.
Figure 12. LCSM images of a capsule-encaged latex particle (390 nm) dyed with rhodamine B: A, fluorescent image; B, bright field image.
Results and Discussion Figure 11. LCSM images of a hollow capsule in an FITC solution captured at the same time, shell dyed with rhodamine B: A, fluorescent signal of FITC molecules in solution; B, fluorescent signal of rhodamine B attached on polymer shells; C, combined image. Table 2. Pore Characteristics with Different Oil Recipes specific amount pore amount amount amount surface of MMA of EDMA of TTMA of solvent size (nm) area (m2/g) (g) (g) (g) (g) no. 1 2 3 4 5
2 2 2 0 0
3 3 3 4 6
1 1 1 2 0
0 2 4 4 4
0 17.68 40.65 25.92 41.56
23.57 43.60 93.38 69.40
method. Laser confocal scanning microscopy (LCSM) (TCS SP5, Leica Ltd., Germany) was used to test the permeability of solid particles with FITC as a fluorescent tracer, and rhodamine B was employed to dye the polymeric texture. Particle size distributions were measured by laser diffractometry with a Mastesizer 2000A (Malvern Instruments Ltd., U.K.).
Emulsion Ripening. Stability is often the primary consideration associated with the application of a W/O/W emulsion. As metastable systems, multiple emulsions are destined to break down into simple ones. There are several possible destruction mechanisms for the original multicore emulsion: route i, internal coalescence (between W1 droplets); route ii, external coalescence (between O globules); route iii, burst escape (between W1 and W2); route iv, diffusion escape (from W1 to W2).16 It was found that under some particular conditions internal coalescence could dominate the emulsion evolution (Figure 2); entrapped W1 preferred to coalesce to a single water core before burst escape. Usually, it costs tens of minutes to obtain the maximal singlecore proportion and even more time to collapse entirely. For the similarity to the ripening process of fruit cells, with all vacuoles coalescing within each cell during ripening, we named the route i dominant process as emulsion ripening, as further illustrated in Figure 3. Such single-core W1/O globules could then serve as templates to produce single-cell microcapsules.
Double Emulsion Templated Microcapsules
Surfactants, stabilizer, and salt are essential components in both W1 and W2, the recipes of which are of guiding significance to the subsequent emulsion ripening. To monitor the ripening of a given recipe, a batch of W1/O/W2 emulsion was first separated into several vessels, and the evolution in each vessel was terminated after a unique ripening time by UV-initiated polymerization. Microcapsules fabricated by a given ripening time were then collected, and the proportions of the three typical morphologies (multicell, single-cell, and no-cell) were calculated. Ternary diagrams were employed to exhibit the profiles of emulsion evolutions, and an example diagram is illustrated in Figure 4. Every peak of the triangle diagram stands for an exclusive morphology distribution, as illustrated by example images attached aside. The morphology information with a certain ripening time could be represented by a single dot on the diagram (Figure 4), and curves connecting these dots record the evolution tendency of the W/O/W emulsion. The higher multicell proportion stands for the better stabilized system, while the higher no-cell proportion represents the less stabilized system. As the singlecell microcapsules were our target products, a sharper curve with a higher single-cell proportion was expected. Effect of Stabilizer on Morphology Control. In this study, polymeric stabilizer PVA was proved an essential additive in water phases, more effective in morphology control than low molecular surfactants. PVA, one of the most popular stabilizers applied in emulsion systems, had also been reported as a W1 additive to improve loading efficiency in W1/O/W2 emulsions.12 It is believed that PVA in both W1 and W2 played similar roles, adhering to the W/O interface and anchoring oil-soluble surfactants on the other side, and the lifetime of the oil membrane between W1 and W2 is severely affected by the surfactant/stabilizer complex. By employing different PVA concentrations in W1 and W2, internal (W1/O) and external (O/W2) interfaces could be designed with different stabilities, which was the way that we proposed to guide the preference of the evolution tendency. Figure 5 shows the evolution profiles of microcapsule morphologies fabricated with different PVA concentrations in W2. OM pictures attached aside show typical morphologies with 2.5% PVA in W2; each image was pictured with a unique ripening time. To emphasize the effects of PVA in W2, no stabilizer was involved in W1. NaCl concentrations were fixed at 60 mM in W1 and 10 mM in W2. With the increase of PVA in W2, the stability of the double emulsion was increased, which can be evidenced from the increased single-cell proportion and decreased no-cell proportion in Figure 5. From the differences of no-cell proportions during the latter phases of ripening, we believed that the main contribution of PVA in W2 was to strengthen the external interfaces and restrict burst escape (route iii) from the outside. The highest single-cell proportion could reach 78%, with 2.5% PVA in W2 and 60 min of ripening. Figure 6 shows the evolution profiles of microcapsule morphologies fabricated with different PVA concentrations in W1. OM pictures attached show the typical morphologies with 1.0% PVA in W1; each picture was obtained with a given ripening time. To emphasize the effects of PVA in W1, only 1.5% PVA was incorporated in W2, and the NaCl concentrations were fixed at 60 mM in W1 and 10 mM in W2. With the addition and increase of PVA in W1, the stability of the internal emulsions was enhanced, as can be evidenced from the slowed evolution, especially the slowed destruction speed, Figure 6. Compared to the effect of PVA in W2, the main contribution of PVA we believed in W1 was to stabilize the internal interfaces, with both the internal coalescence and the burst escape (routes i and iii) being obviously
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restricted. The highest proportion of single-cell morphology (73%) was obtained with 1.0% PVA in W1 and 4 h of ripening. The batch with no PVA involved in W1 led to the fastest collapse, while the one with the highest concentration of 2% led to an even slower ripening than collapse. In short, the highest single-cell proportion would be obtained with an optimal PVA concentration in W1, a mild value such as around 1%. Effect of the Osmotic Gradient on the Morphology Control. The osmotic gradient between W1 and W2 is another important factor to affect the emulsion evolution and the final morphology. Solute in the water phases of a multiple emulsion was usually employed to counteract Ostwald ripening and maintain osmotic equilibrium.24 Under a nonequilibrium condition, water molecules would migrate across the oil driven by an osmotic gradient or by the Laplace effect,25-28 and such migration is crucial to evolution behavior. The osmotic pressure generated by an electrolyte is a notable part of the composite force on water diffusion and could be an easy factor to manipulate. In this study, NaCl was applied to adjust the osmotic gradient between W1 and W2 and to affect the emulsion ripening eventually. Figure 7 shows the evolution profiles of microcapsule morphologies with three NaCl concentrations in W2, with a fixed NaCl concentration of 60 mM in W1. Figure 8 shows the evolutions with three NaCl concentrations in W1, with a fixed NaCl concentration of 10 mM in W2. OM pictures attached aside record the typical morphology evolution with given recipes. According to the optimization on PVA concentrations, 2.5% PVA and 1.0% PVA were incorporated in W2 and W1, respectively. From the results illustrated, the higher NaCl concentration in W1 than W2 led to faster evolution and to a higher single-cell proportion; however, it also led to faster collapse during the latter periods. During the tests on NaCl in W2, the highest single-cell proportion obtained was 89%, with no salt incorporated in W2, and during the tests on NaCl in W1 the single-hollow proportion could reach as high as 98%, with 120 mM NaCl in W1. According to the diffusion mechanism, the higher osmotic gradient leads to faster water migration; this is the reason why more salt in W1 than W2 will lead to faster swelling of entrapped W1 droplets and to a more vulnerable system. With the external interfaces better stabilized than the internal ones, ripening for the single-cell morphology would dominate the evolution during the early period and the higher single-cell proportion could be guaranteed. Although the initial loading capacity was limited by the water/ oil ratio of the primary emulsion, large cavities with thin shells could still be fabricated by the swelling mechanism. Figure 9 shows the relation between the initial NaCl concentration in W1 and the shell thickness of obtained microcapsules, 30-50 µm in diameter, with W2 NaCl fixed at 0.1 mM. PVA concentrations of 4.0% and 1.0% were employed in W2 and W1, respectively, to guarantee at least 90% single-cell proportion. Pictures attached above show the typical images obtained with different initial NaCl concentrations in W1. With an increase of the initial salt in W1, the volume of the hollow capacities became larger and the shell became thinner. Nanopores Fabricated in the Shell. The phase-separation mechanism was employed to produce permeable pores in the (24) Rosano, H. L.; Gandolfo, F. G.; Hidrot, J.-D. P. Colloids Surf., A 1998, 138, 109–121. (25) Matsumoto, S.; Inoue, T.; Kohda, M.; Ikura, K. J. Colloid Interface Sci. 1980, 77, 555–563. (26) Wen, L.; Papadopoulos, K. D. J. Colloid Interface Sci. 2000, 235, 398– 404. (27) Mezzenga, R. Food Hydrocolloids 2006, 21, 674–682. (28) Wen, L.; Papadopoulos, K. D. Colloids Surf., A 2000, 174, 159–167.
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shells, during the eventual suspension polymerization. According to the well-stabilized principles on phase separation, the pore size of polymer microspheres can be modulated by choosing suitable porogens and cross-linking degree of the polymeric matrix.5-7 Microstructures of the microcapsules are illustrated in Figure 10 with SEM images. The pore size could be adjusted from zero to tens of nanometers in diameter, analyzed by the BET nitrogen adsorption method. The general relations between the oil components and the porosity are illustrated in Table 2. When no solvent was added to the oil, nonporous shells were obtained. An image of the nonporous capsules is illustrated in Figure 10A. With an increase of the mixture solvent proportion in the oil phase, from 25% to 40%, the pore size increased to as high as 40.65 nm, with the specific surface area increasing from 23.57 to 43.60 m2/g. Typical images of porous microcapsules, which were fabricated according to recipe no. 3 listed in Table 2, are illustrated in Figure 10B-D, with different magnifications. Nanopores could be clearly distinguished on both sides of the shells and in the cross section as well, parts B and C. Part D shows the coarse surface with an irregular pore shape of a microcapsule with a higher magnification. The greater the amount of cross-linker incorporated, the smaller pore size with a larger specific area obtained. When EDMA, which possesses two vinyl groups on each molecule, was employed as the only monomer, 41.56 nm pores with 69.40 m2/g specific surface area were obtained with 40% mixture solvent in the oil, and when 1/3 (w/w) EDMA was replaced by TTMA, which possesses three vinyl groups on each molecule, 25.92 nm pores with 93.38 m2/g was the result. The porosity of the cross-linked texture could be manipulated effectively by oil ingredients according to the phaseseparation mechanism, and diverse shell structures could be expected in future development. LCSM was employed to measure the permeability of the porous shells for small molecules. When a fluorescent dye, FITC (MW 389.4), was added to the external aqueous solution of microcapsules, which were fabricated according to recipe no. 3 in Table 2 and could possess the same morphology as illustrated in Figure 10B-D, an FITC signal with the same fluorescent intensity was soon detected inside the microcapsules, as soon as the microcapsules were focused by the LCSM instrument. Figure 11 shows porous microcapsules merged in an FITC solution, with three images featured with different fluorescent signals captured by LCSM at the same time. The porous shells were dyed with rhodamine B in advance to show the shell location. The rapid diffusion of FITC suggested that such porous shells possess excellent permeability and offer convenient channels for molecule exchange across the polymeric boundary. Moreover, PMMA latex particles 390 nm in diameter were employed as core material to test the holding capability. Latex particles were predispersed into W1 with 3.0% solid content and then followed the encapsulation procedure herein presented,
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according to recipe no. 3 listed in Table 2. To facilitate the observation, latex particles and microcapsules were dyed with rhodamine B after encapsulation. Figure 12 shows the image of capsule-encaged nanoparticles. After the encapsulation, latex particles were well restricted by the porous shells and enjoyed a confined freedom with the original mobility well preserved in such hollow cavities. In the successive development, nanoparticlebased catalysts were successfully assembled within such microcapsules, leading to nanoparticle-assisted reactors in microscale.
Conclusions A novel form of single-cavity microcapsules, which were prepared by a two-step emulsification, emulsion ripening, and subsequent suspension polymerization, was herein presented, and so was a universal strategy to produce single-core W1/O globules from their original multicore precursors in W1/O/W2 emulsions. To obtain as many single-core W1/O globules as possible, the coalescence between entrapped W1 droplets should be adjusted much faster than coalescence between W1 and W2 through a typical evolution we named “emulsion ripening”. To promote the emulsion ripening, the external interface (between O and W2) should be better stabilized than the internal one (between W1 and O). PVA was employed in both W1 and W2 to enhance the stability of the interfaces, and the difference in PVA concentrations was proved to be of guiding significance to the evolution. The osmotic gradient between W1 and W2, as could be easily adjusted by salt concentrations, was proved to be another crucial parameter in evolution control. More salt dissolved in W1 than W2 would lead to faster swelling of the entrapped volume and sequentially to faster ripening and a higher single-core proportion. It was also found that the final volume of entrapped water cores and the shell thickness of the oil membranes could be effectively adjusted by such nonequilibrium osmotic conditions. Suspension polymerization initiated by UV irradiation was eventually employed to transform the W1/O globules into solid microcapsules, and nanochannels across the shells could be formed with proper porogen(s) and cross-linker(s) incorporated. During the encapsulation process, the entrapped water phase was guaranteed biocompatible, leading to a promising system for encapsulation of bioactive materials in future development. Acknowledgment. We thank the National Basic Research Program of China (973 Program, Contract No. 2009CB724705) and Chinese National Science Foundation of China (Grant Nos. 20576135, 20536050, and 20728607) for support. The support from the Chinese Academy of Sciences for international collaboration is also greatly appreciated. LA804173B
