Article pubs.acs.org/Langmuir
Monodisperse w/w/w Double Emulsion Induced by Phase Separation Yang Song and Ho Cheung Shum* Department of Mechanical Engineering, The University of Hong Kong, Pokfulam Road, Hong Kong S Supporting Information *
ABSTRACT: We develop an approach to fabricate monodisperse water-in-water-in-water (w/w/w) double emulsion in microfluidic devices. A jet of aqueous solution containing two incompatible solutes, dextran and polyethylene glycol (PEG), is periodically perturbed into water-in-water (w/w) droplets. By extracting water out of the w/w droplet, the solute concentrations in the droplet phase increase; when the concentrations exceed the miscibility limit, the droplet phase separates into two immiscible phases. Consequently, PEG-rich droplets are formed within the single emulsion templates. These PEG-rich droplets subsequently coalesce with each other, resulting in transiently stable w/w/w double emulsions with a high degree of size uniformity. These double emulsions are free of organic solvents and thus are ideal for use as dropletvessels in protein purification, as microreactors for biochemical reactions, and as templates for fabrication of biomaterials. diagrams.7 This particular ATPS has been widely used for preparing microspheres8 as well as for separating proteins, cells, and particles.9−11 Recently, a microfluidic approach has been introduced to produce all-aqueous emulsions using ATPS.12,13 At typical flow rates of 10−1−103 μL/h, a dispersed phase forms a jet surrounded in an immiscible continuous phase in microfluidic devices. By applying oscillating pressure, the jet can be perturbed to break up into droplets. Using similar approaches, double emulsions have been successfully fabricated by sheathing the inner jet inside a middle jet and subsequently breaking up the compound jet.12 These double emulsion droplets typically have a polydispersity of around 10%. Further reduction in the polydispersity of the all-aqueous emulsion droplets is desired, especially for biomedical applications that are sensitive to the droplet size; these include quantitative analysis of biomolecules and screening of enzyme bioactivity in biochemical reactions. In this paper, we demonstrate the generation of w/w/w double emulsions with an excellent polydispersity of less than 4%. A single-phase solution of 5% dextran/1% PEG/94% water is injected as the dispersed phase, which is surrounded by a concentrated PEG solution in a microchannel. When the driving pressure of the dispersed phase varies periodically, the jet breaks up into droplets. We investigate the polydispersity of the resultant w/w droplets as a function of the perturbation frequency and flow rates. Droplets that are formed directly at the nozzle show superior monodispersity when compared with those formed due to the breakup of jets at some distance
1. INTRODUCTION Double emulsions, which are suspensions of droplets containing smaller droplets of an immiscible phase within them, are important templates for synthesizing core−shell structured materials,1 such as vesicles, capsules, and particles.2−4 In conventional double emulsions, organic solvents are frequently used as at least one of the emulsion phases.5,6 As concerns over the safety and biocompatibility of these emulsions are increasing, a route of producing all-aqueous double emulsion without any organic solvent phase is desired. Moreover, when these emulsions are used as liquid vessels for protein delivery, or microreactors for enzyme reactions, the bioactivity of these biomolecules may be impaired due to denaturation by harmful organic solvents. In addition, hydrogel particles that are templated by water-in-oil (w/o) emulsion droplets often need to be washed repeatedly to remove any remaining oil phase before use with biological tissues and cells. This tedious step severely limits the attractiveness of microfluidic approaches for fabricating hydrogel particles for biomedical applications. An organic-solvent-free approach to fabricate droplets is needed, not only for biomedical applications, but also for food industries, cosmetics formulation, and bioinspired studies. Formation of all-aqueous emulsions requires two immiscible aqueous phases, which are commonly known as aqueous twophase systems (ATPSs). The two immiscible aqueous phases are formed by dissolving two incompatible solutes in water above critical concentrations of phase separation. Dextran and polyethylene glycol (PEG) are two such examples of incompatible solutes; a solution with high concentrations of dextran and PEG spontaneously separates into a dextran-rich phase and a PEG-rich phase. The equilibrium compositions of these two phases have been measured and presented in phase © 2012 American Chemical Society
Received: July 3, 2012 Revised: July 30, 2012 Published: July 31, 2012 12054
dx.doi.org/10.1021/la3026599 | Langmuir 2012, 28, 12054−12059
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Article
Figure 1. (a) Phase diagram of dextran (Mw = 500 000 Da)/PEG (Mw = 8000 Da)/H2O at room temperature (22 °C), redrawn from ref 7, and the blue dashed lines are tie lines. Compositions of the outer and inner fluids used to form w/w/w double emulsion are denoted by points A and B, respectively. The line BD represents the change in emulsion compositions upon mixing of the two fluids. The line AE is a tie line estimated by drawing a line parallel to the measured tie lines. (b) Schematic of a capillary device used for preparing w/w/w double emulsions. The configuration of the capillaries inside the device is magnified in the schematic at the bottom half of panel (b).
2. EXPERIMENTAL SECTION
produces periodical pressure fluctuation in the inner tubing, leading to an oscillatory flow of the inner fluid. Consequently, the jet was forced to break up at certain frequencies. After the jet was broken up into droplets, the diameters of the fabricated droplets were measured using an open-source image-processing software, ImageJ. The uncertainty of the measured diameter was about ±1 μm. The polydispersity was obtained by measuring the diameters of at least 100 droplets. To observe the dynamics of phase separation, we monitored the shapes of droplets inside the microchannel using a camera connected to the optical microscope. The droplets were dyed with 0.625 wt % methylene blue to improve image contrast.
2.1. Emulsion Preparation. An aqueous solution with 5 wt % dextran (Mw = 500 000 Da, Shanghai Kayon Biological Technology Co.) and 1 wt % PEG (Mw = 8000 Da, Aldrich) was chosen as the inner fluid (disperse phase). This fluid remains a single phase at room temperature and atmospheric pressure, according to the phase diagram in Figure 1a. The outer fluid, which extracts water from the inner fluid upon mixing, was an aqueous phase with 8 wt % PEG and 20 wt % glycerol. Glycerol increases the viscosity and density of the outer fluid. 2.2. Fabrication of the Glass Capillary Microfluidic Device. We used a capillary microfluidic device to generate all-aqueous emulsions.14 Briefly, a cylindrical glass capillary with inner and outer diameters of 0.86 mm and 1.0 mm, respectively, was pulled to achieve a tapered tip geometry using a pipet puller (Sutter Instrument Co., USA). Then we polished the tips of two capillaries on an abrasive sand paper until their inner diameters reach about 30 and 125 μm. Subsequently, the two capillaries were coaxially positioned inside another square capillary with an inner diameter of 1.0 mm, as shown by the schematic in Figure 1b. The inner and outer fluids were separately injected into the device through two inlets, each of which was connected through plastic tubing to a syringe driven by a syringe pump (Longer pump, model LSP01-2A). 2.3. Perturbation-Induced Droplet Formation and Phase Separation within the Droplets. We tuned the velocity of the inner fluid Qin from 10 to 200 μL/h, and fixed the velocity of the outer fluid Qout at 1000 μL/h. The resultant jets were observed under an optical microscope (DM 180, Motic Inc.). Then we perturbed the jet using a mechanical vibrator (PASCO scientific, SF-9324), which was controlled by a sinusoidal-wave generator (RIGOL, DG1012). The inner tubing is firmly inserted between a tubing holder and the vibrator. This vibrator oscillates under the control of the generator, and thus, the inner plastic tubing is squeezed and relaxed accordingly. As a result, the instantaneous flow rate of the inner fluid also changes periodically. The input voltage of vibrator was fixed at 10 V, and the perturbation frequency f p was varied from 2 to 30 Hz. The vibrator
3. RESULTS AND DISCUSSION 3.1. Perturbation-Induced Formation of Monodisperse Droplets. The inner fluid forms a jet surrounded by the outer fluid upon injection inside the microfluidic device. In the absence of perturbation, the jet spontaneously breaks up into polydisperse w/w droplets with a broad size distribution. To control the droplet sizes and improve the droplet size distribution, the jet is periodically perturbed using a vibrator and forced to break up into droplets.13,15,16 As the instantaneous flow rate of the inner fluid changes sinusoidally, the resultant jet diameter varies and forms a corrugated interface. This instantaneous deviation of the jet diameter from its average diameter is defined as the perturbation amplitude of the system. With an increase in the input voltage of perturbation Up, the perturbation amplitude of the jet also rises.17 When this perturbation amplitude reaches a length scale comparable with the jet radius, the jet breaks up into droplets immediately at the nozzle17 and no satellite drops are observed. The resultant droplets are highly uniform in size. By contrast, with a smaller perturbation amplitude obtained at lower Up, the jet breaks up into droplets at some distance from the nozzle of the receiving tube. Satellites drops are occasionally observed along with the perturbed jet; this raises the polydispersity of the droplets downstream. The resultant droplets have a larger polydispersity (around 5%) than those formed directly at the nozzle (
