Generation Of Double Emulsions Research Articles

Double emulsion generation in shear-thinning fluids under electric field effects

Double emulsions are encapsulated microstructures with medical, food, and cosmetics applications. Precise control over their parameters, such as volume, thickness, and size distribution, is crucial. This study numerically investigates, for the first time, the generation of double emulsions in a shear-thinning outer fluid under the influence of an external DC electric field. A dual co-flowing microfluidic device is chosen as the flow domain, and the shear-thinning behavior of the outer fluid is approximated by the Carreau–Yasuda rheological model. The governing equations are discretized on structured numerical grids using the VOF-CSF model and the finite volume method. The combined effects of the shear-thinning behavior and electric forces on compound droplet formation parameters, including volume, eccentricity, maximum thickness, and jet length, are studied carefully. Results show that for highly shear-thinning fluids, the application of electric forces can ensure the generation of desired compound droplets, which is not feasible without electric field effects. However, excessive electric forces can disrupt the droplet generation process, especially for moderate to low shear-thinning fluids. These findings provide new insights into the role of external electric fields in double emulsion generation and highlight their potential to optimize the production process.

Micro‐Patterning Wettability in Very Large Scale Microfluidic Integrated Chips for Double Emulsion Generation

AbstractThe fabrication of microfluidic chips is becoming a mature field and channels can be reliably defined with micrometer‐scale precision in a variety of substrates. In addition to channel geometry, micrometer‐scale patterning of wettability in microfluidic channels is essential for many applications including multi‐phase flow stabilization and multiple emulsion generation. Unfortunately, current methods to pattern wettability in microfluidics suffer from low spatial resolution, inability for patterns to be arbitrarily defined, cumbersome procedures, and incompatibility with parallelized architectures for scaled‐up production of microfluidic generated materials. To address these issues, a method is developed to lithographically define micrometer‐scale resolution patterns of wettability on all channel surfaces (ceiling, floor, and walls) in silicon and glass microfluidic devices with complex 3D geometry. A process is reported to pattern silanes on microfluidic chips that uses photolithography and an optimized process that keeps silanized surfaces stable through the microfabrication process, including anodic bonding. The versatility of this approach is highlighted by patterning wettability of a silicon/glass device to generate both highly uniform water–in‐oil–in‐water and oil–in‐water–in‐oil double emulsions. The applicability of this process is demonstrated to the parallel generation of materials in a microfluidic chip with complex geometry, by fabricating and successfully validating parallelized double emulsion generators.

Single-step generation of double emulsions in aqueous two-phase systems

This communication presents a simple yet straightforward method for preparing water-in-water-in-water particle-stabilized double emulsions, also known as Pickering double emulsions. The approach involves using oppositely charged nanoparticles (OCNPs) in two distinct fluid phases, promoting self-assembly and the formation of aggregates with varying sizes and compositions. By enhancing the interfacial area through the adsorption of aggregates at the interface, this method increases the Gibbs detachment energy of particles between the two aqueous phases, forming stable double emulsions. Furthermore, we investigated the impact of the molecular weight of polyethylene oxide and dextran in the respective fluid phases and the mass ratio (M) of the OCNPs on double emulsion formation. The results demonstrate that the molecular weight of the polymers used in the aqueous phase is a critical parameter influencing the structural formation of the emulsion and the generation of double emulsions. Consequently, double emulsions are formed when equal molecular weight polymer mixtures are employed at an appropriate M, with the dispersed phase placed in the highly viscous continuous phase. The proposed method offers a one-step synthesis process, enabling easy preparation, and exhibits excellent stability for at least 30 days. This study represents the first reported approach for the one-step synthesis of multiple emulsions in an aqueous two-phase system utilizing a Pickering emulsion template.

Ferrofluid double emulsion generation and manipulation under magnetic fields

Magnetically controlled double emulsions offer a promising tool for targeted drug delivery. Here, the effect of a non-uniform magnetic field induced by a permanent magnet on ferrofluid double emulsion generation and morphology is studied. In the numerical part, the double emulsion (and its ferrofluid core) migration was analyzed for different transverse distances of the magnet from the channel centerline (Ly) and different core and shell diameters. In the experimental part, water-based ferrofluid is encapsulated in a UV-curable polymer phase in water, forming a W/O/W double emulsion. As the magnet’s downstream distance from the ferrofluid junction reduces, the core aspect ratio increases while its diameter decreases. Finally, the core eccentricity was calibrated with Ly, and by shining a UV light, this core eccentricity was affixed inside the UV-curable medium. These double emulsions can then rearrange and orient under a uniform magnetic field for more efficient targeted drug delivery.

Microfluidic formation and manipulation of OLA-based liposomes for synthetic biology research.

Bottom-up synthetic biology aims to create biomimetic systems from the basic elements of cellular life. Such synthetic cell analogs may support scientists in understanding the fundamental biological behaviors at the cellular level. As microscopic aqueous compartments confined within a lipid bilayer, liposomes have been widely used as simplistic cellular mimics for synthetic cell studies. Here, we describe the manufacturing protocol for octanol-assisted liposome assembly (OLA) to produce unilamellar, monodispersed, cell-sized, and biocompatible liposomes as well as in situ liposome experiments. We use maskless, direct-write lithography, which allows for rapid prototyping, to fabricate PDMS-based lab-on-a-chip devices. OLA makes use of the double emulsion generation and dewetting purification approach, with the membrane composition of the formed liposomes and the nature of the encapsulated solution widely adjustable. Using highly responsive pressure-driven flow, the liposome size is tuneable with a steady liposome generation, and the experiment requires very low sample volumes (∼50 µl). We enrich the liposomes from the waste products using density-based on-chip separation techniques, which allows us to conduct experiments on the same device with real-time observation, right after production. As a proof-of-principle synthetic biology application, we demonstrate the capability of adjusting the microenvironment by inducing pH-controlled coacervation between ATP and poly-Llysine molecules inside the liposomes via transmembrane proton flux. Such liquid-liquid phase separation, creating multiple distinct phases from a single homogeneous mixture, is known to play a key role in a variety of cellular processes, including the formation of membraneless organelles as well as numerous other supramolecular assemblies. Our technique shows promising potential for synthetic cell research, particularly in terms of step-by-step construction and experimental control.

Generation of double emulsions from commercial single-emulsion microfluidic chips: a quality-control study

Generation of double emulsions from commercial single-emulsion microfluidic chips: a quality-control study

Generation and evolution of double emulsions in a circular microchannel

Double emulsions are attracting much attention from academic community and industry fields. Emerging microfluidics technology offered new ways to prepare unprecedentedly controllable double emulsions. Here a numerical simulation method, volume of fluid (VOF) technique, was adopted to study the generation and evolution of double emulsions in a circular microchannel, which is helpful for understanding the underlying physics of double emulsion preparation and optimizing the operation conditions. The flow pattern regions where double emulsions existed were found and classified according to dripping and jetting models of double emulsion generation. Velocity fields inside and outside double emulsions were investigated, and several distinct velocity vortexes were observed. Several characteristic parameters involving pressure distribution of fluids and deformation index of double emulsions were studied. At last, preparation of double emulsions with desired configurations, including same inner droplets engulfed by different outer droplets and same outer droplets encapsulating different inner droplets, were explored in depth.

Simplified, Shear Induced Generation of Double Emulsions for Robust Compartmentalization during Single Genome Analysis.

Drop microfluidics has driven innovations for high throughput, low input analysis techniques such as single-cell RNA-seq. However, the instability of single emulsion (SE) drops occasionally causes significant merging during drop processing, limiting most applications to single-step reactions in drops. Here, we show that double emulsion (DE) drops address this critical limitation and completely prevent drop contents from mixing. DEs show excellent stability during thermal cycling. More importantly, DEs undergo rupture into the continuous phase instead of merging, preventing content mixing and eliminating unstable drops from the downstream analysis. Due to the lack of drop merging, the monodispersity of drops is maintained throughout a workflow, enabling the deterministic manipulation of drops downstream. We also developed a simple, one-layer DE drop maker compatible with simple surface treatment using a plasma cleaner. The device allows for the robust production of single-core DEs at a wide range of flow rates and better control over the shell thickness, both of which have been significant limitations of conventional two-layer devices. This approach makes the fabrication of DE devices much more accessible, facilitating its broader adoption. Finally, we show that DE droplets eliminate content mixing and maintain compartmentalization of single virus genomes during PCR-based amplification and barcoding, while SEs mixed contents due to merging. With their resistance to content mixing, DE drops have key advantages for multistep reactions in drops, which is limited in SEs due to merging and content mixing.

A double-step emulsification device for direct generation of double emulsions.

In microfluidic step emulsification, the size of droplets generated in the dripping regime is predominantly determined by the nozzle's height and only weakly depends on the applied flow rates or liquid properties. While the generation of monodisperse emulsions at high throughput using step emulsifiers has been well established, the generation of double emulsions, i.e., liquid core-shell structures, is still challenging. Here, we demonstrate a novel double-step emulsification method for the direct generation of multi-core double-emulsions and provide a predictive model for the number of cores. While the mechanism of the formation of the core droplets or empty shell droplets follows the well-established scenario of simple step emulsification, the formation of double-emulsion droplets is strongly affected by the presence of the cores. Passing of the cores through the narrowing neck of the shell postpones shell pinch-off. In particular, we demonstrate that our system can be used for the generation of arbitrary large, tightly packed droplet clusters consisting of a controllable number of droplets. Finally, we discuss the options of upscaling the system for high-throughput generation of tailored double emulsions.

Rapid, Simple, and Inexpensive Spatial Patterning of Wettability in Microfluidic Devices for Double Emulsion Generation.

Water-in-oil-in-water (w/o/w) double emulsion (DE) encapsulation has been widely used as a promising platform technology for various applications in the fields of food, cosmetics, pharmacy, chemical engineering, materials science, and synthetic biology. Unfortunately, DEs formed by conventional emulsion generation approaches in most cases are highly polydisperse, making them less desirable for quantitative assays, controlled biomaterial synthesis, and entrapped ingredient release. Microfluidic devices can generate monodisperse DEs with controllable size, morphology, and production rate, but these generally require multistep fabrication processes and use of different solvents or bulky external instrumentation to pattern channel wettability. To overcome these limitations, we propose a rapid, simple, and inexpensive method to spatially pattern wettability in microfluidic devices for the continuous generation of monodisperse DEs. This is achieved by applying corona-plasma treatment to a select zone of the microchannel surface aided by a custom-designed corona resistance microchannel to strictly confine the plasma-treatment zone in a single polydimethylsiloxane (PDMS) microfluidic device. The properties of PDMS channel surfaces and key microchannel regions for DE generation are characterized under different levels of treatment. The size, shell thickness, and number of inner cores of generated DEs are shown to be highly controllable by tuning the phase flow rate ratios. Using DEs as templates, we successfully achieve a one-step generation and collection of gelatin microgels. Additionally, we demonstrate the biological capability of generated DEs by flow cytometric screening of the encapsulation and growth of yeast cells within DEs. We expect that the proposed approach will be widely used to create microfluidic devices with more complex wettability patterns.

Controllable preparation of double emulsion droplets in a dual-coaxial microfluidic device

This article investigates the formation process of double emulsions, controlling their sizes, and manipulating the encapsulation process in a three nested microcapillary tube configuration. In the proposed geometry, as a result of the interaction of liquids using dripping instability, double emulsions are generated. Governing equations are discretized using the finite-element method utilizing COMSOL Multiphysics 5.4 software, employing axisymmetric assumption, and the interface position is tracked by solving additional transport equations of the phase-field method. Influence of phase flow rates in the form of inner phase Weber number, Wein, intermediate phase capillary number, Caint, and outer phase capillary number, Caout, are studied on the flow regimes and characteristics of double emulsions. Results exhibit that the outer phase flow rate has the most influence among other phase flow rates. Additionally, three distinct regimes, named single emulsion, decussate, and double emulsion, are observed by changing intermediate phase flow rare. Furthermore, satellite droplets are detected at relatively high flow rates of the outer phase due to the second instability induced by the continuous phase to the intermediate phase jet. A comparison between the formation frequency of the present study with some similar works, including both capillary-based and channel-based devices, unveils that the existing configuration has a higher throughput of double emulsion generation in a broad range of flow rates.

Rapid generation of double-layer emulsion droplets based on microfluidic chip

In vitro compartmentalization (IVC) links genotype and phenotype by compartmentalizing individual genes (including expression system) or cells into a micro-droplet reaction region. Combined with fluorescence-activated cell sorting (FACS), it can detect and separate single droplets in ultra-high throughput. IVC-FACS screening method has been widely used in protein engineering, enzyme directed evolution, etc. However, it is difficult to control the homogeneity of droplet size by mechanical dispersion method in previous studies, which seriously affects the quantitative detection of droplets and reduces the efficiency and accuracy of this screening method. With the rapid development of microfluidic chip manufacturing technology, the microfluidic chip-based methods for droplet generation are becoming more efficient and controllable. In this study, firstly, the water-in-oil (W/O) single-layer droplet generation chip was used to prepare single-layer monodisperse W1/O droplets at a high generation frequency, and then the W1/O droplets were reinjected into water-in-oil-in-water (W/O/W) double-layer droplet generation chip to prepare uniform W1/O/W2 double-layer emulsion droplets. By optimizing the flow rate and ratio of the oil and water phases, a single-layer micro-droplet can be generated with a diameter range from 15.4 to 23.2 μm and remain stable for several days under normal incubation. Then the single-layer droplets were reinjected into the double emulsion generation chip. By adjusting the flow rate of drop phase, oil phase and water phase, the double-layer emulsion droplets with a diameter range from 30 to 100 μm at a rate of 1 000 droplets/s could be obtained. Escherichia coli embedded in the double-layer emulsion droplets could be cultured and induced for protein expression. This study lays a foundation for the establishment of a high-throughput screening method based on the droplet and flow cytometry.

Lipid-Stabilized Double Emulsions Generated in Planar Microfluidic Devices

Microemulsions have found a wide range of applications exploiting their chemical and physical properties. Development of microfluidic-based approaches has allowed for the controlled production of highly monodispersed emulsions, including the formation of multiple and hierarchical emulsions. Conventional poly(dimethylsiloxane)-based microfluidic systems require tight spatial control over the surface chemistry when used for double emulsion generation, which can be challenging to achieve on the micrometer scale. Here, we present a two-dimensional device design, which can selectively be surface-treated in a straightforward manner and allows for the formation of uniform water/oil/water double emulsions by combining two distinct hydrophilic and hydrophobic surface properties. These surfaces are sufficiently separated in space to allow for imparting their functionalization without the requirement for lithographic approaches or complex flow control. We demonstrate that a mismatch between the wettability requirements of the continuous phase and the channel wall inherent in this approach can be tolerated over several hundreds of micrometers, opening up the possibility to use simple pressure-driven flows to achieve surface functionalization. The design architecture exhibits robust efficiency in emulsion generation while retaining simple device fabrication. We finally demonstrate the potential of this approach by generating water in oil in water emulsions with lipid molecules acting as surfactants.

Coaxial oblique interface shearing: tunable generation and sorting of double emulsions for spatial gradient drug release.

We propose a coaxial oblique interface shearing (COIS) process for one-step generation of double emulsions which are synchronously sorted with spatial gradient distributions. As a coaxial needle supplying the inner and outer liquids obliquely vibrates across an air-liquid interface, the pinch-off of the compound liquid neck arises and the resultant double emulsions moves with tunable lateral displacements in the receiving phase. In the COIS process, the morphology and size of the double emulsions are heavily dependent on the vibration frequency and the inner and outer liquid flow rates. The lateral droplet displacements changing with process parameters can be precisely controlled in experiments and predicted theoretically by the Stokes drift model. Furthermore, the feasibility of the COIS process in spatial gradient drug release is verified. The double emulsions sorted along a specific direction are available for spatial gradient release under thermal and chemical environments, respectively. The COIS technique has great potential in fields of sensors, spatial gradient materials, advanced drug delivery and biomedical applications.

Parallelizable microfluidic dropmakers with multilayer geometry for the generation of double emulsions.

Microfluidic devices enable the production of uniform double emulsions with control over droplet size and shell thickness. However, the limited production rate of microfluidic devices precludes the use of monodisperse double emulsions for industrial-scale applications, which require large quantities of droplets. To increase throughput, devices can be parallelized to contain many dropmakers operating simultaneously in one chip, but this is challenging to do for double emulsion dropmakers. Production of double emulsions requires dropmakers to have both hydrophobic and hydrophilic channels, requiring spatially precise patterning of channel surface wettability. Precise wettability patterning is difficult for devices containing multiple dropmakers, posing a significant challenge for parallelization. In this paper, we present a multilayer dropmaker geometry that greatly simplifies the process of producing microfluidic devices with excellent spatial control over channel wettability. Wettability patterning is achieved through the independent functionalization of channels in each layer prior to device assembly, rendering the dropmaker with a precise step between hydrophobic and hydrophilic channels. This device geometry enables uniform wettability patterning of parallelized dropmakers, providing a scalable approach for the production of double emulsions.

On-Demand Generation of Double Emulsions Based on Interface Shearing for Controlled Ultrasound Activation.

Stimuli-responsive microcarriers (SRMs) based on multiple emulsions can be widely used in advanced drug delivery, tissue engineering, biosensing, and cell biology. Here a simple and effective compound interface shearing (CIS) method is proposed to one-step produce SRMs for controlled ultrasound (US) activation. In the CIS process, a coaxial needle supplying the core and shell liquids vibrates periodically across a free gas-liquid surface, resulting in the pinch-off of a compound liquid neck for on-demand generation of multiple emulsions. The CIS process is free of confined walls with a pure interface shearing mechanism. Perfectly uniform SRMs with tunable core-shell volume ratios can be produced, following a scaling law of their size as a function of the liquid flow rates and the vibration frequency. US- and magnetic-responsive microcapsules are prepared for magnetic-guided site-targeting delivery, and acid-aided sequential US activation realizes the synergistic delivery of hydrophilic and hydrophobic payloads. It can be concluded that the CIS technique is able to generate multifunctional SRMs at low cost, high uniformity, high flexibility, and effective process control for various fields of potential applications.

One-step wettability patterning of PDMS microchannels for generation of monodisperse alginate microbeads by in Situ external gelation in double emulsion microdroplets

Droplet-based microfluidic systems are promising tools for many biological applications from digitalized analysis to drug delivery and biomolecules synthesis. Among these, fabrication of microgels, particularly alginate microbeads, has substantial importance due to their capability for three-dimensional (3D) encapsulation of cells, drugs and other bioactive materials. However, biocompatible generation of monodisperse alginate microbeads is still challenging. In this study, a new droplet-based microfluidic strategy is developed for cell and chip friendly fabrication of monodispersed alginate microbeads. The polydimethylsiloxane (PDMS) microfluidic device is first treated using on-chip plasma-assisted deposition of polyvinyl alcohol, to selectively modify the hydrophobicity of microchannels. The proposed wettability patterning approach is permanent, simple, reliable, and time-effective, which makes the resulting microfluidic device highly stable and controllable for generation of double emulsions (DEs). The device is then used for fabrication of monodisperse alginate microbeads using external gelation in DE microdroplets. The reported microbead fabrication method offers both biocompatibility as a result of external gelation and rapid recovery of beads from processing solutions, as well as chip-compatibility due to clog-free gelation. This simple and robust DE strategy provides new opportunities not only for 3D micro-encapsulation but also for other droplet-based microfluidic applications in diagnostic testing and drug delivery.

A Modular Microfluidic Device via Multimaterial 3D Printing for Emulsion Generation

3D-printing (3DP) technology has been developing rapidly. However, limited studies on the contribution of 3DP technology, especially multimaterial 3DP technology, to droplet-microfluidics have been reported. In this paper, multimaterial 3D-printed devices for the pneumatic control of emulsion generation have been reported. A 3D coaxial flexible channel with other rigid structures has been designed and printed monolithically. Numerical and experimental studies have demonstrated that this flexible channel can be excited by the air pressure and then deform in a controllable way, which can provide the active control of droplet generation. Furthermore, a novel modular microfluidic device for double emulsion generation has been designed and fabricated, which consists of three modules: function module, T-junction module, and co-flow module. The function module can be replaced by (1) Single-inlet module, (2) Pneumatic Control Unit (PCU) module and (3) Dual-inlet module. Different modules can be easily assembled for different double emulsion production. By using the PCU module, double emulsions with different number of inner droplets have been successfully produced without complicated operation of flow rates of different phases. By using single and dual inlet module, various double emulsions with different number of encapsulated droplets or encapsulated droplets with different compositions have been successfully produced, respectively.

Controlled Generation of Double Emulsions for Laser Fusion Target Fabrication Using a Glass Capillary Microfluidic Device

We report the controllable generation of double emulsions for target fabrication using glass capillary microfluidic devices. Instead of a conventional triple-orifice droplet generator, user-friendly glass capillary devices are used to produce micrometer to millimeter-sized water-in-oil-in-water emulsions. The double emulsions have a relatively uniform size distribution with an average outer diameter of 1420 μm. The sizes of the emulsions can also be varied by changing the ratio of the inner, middle, and outer fluids. Increasing the flow rate ratio of the outer fluid to the other fluids [Qo/(Qm+Qi)] from 3 to 11, the outer radii of the emulsions decrease from 1120 to 950 μm. On the other hand, increasing the flow rate ratio of the middle fluid to the inner fluid (Qm/Qi) from 0.7 to 1.6, the aspect ratio of the emulsions increases from 4 to 8. Our experimental values are in good agreement with a simple theoretical model. These results suggest that our present method to control the generation of double emulsions can be used as an alternative approach to fabricate polystyrene targets for future laser fusion experiments.

Molecular and functional assessment of multicellular cancer spheroids produced in double emulsions enabled by efficient airway resistance based selective surface treatment

Multicellular spheroids have garnered significant attention as an in vitro three-dimensional cancer model which can mimick the in vivo microenvironmental features. While microfluidics generated double emulsions have become a potential method to generate spheroids, challenges remain on the tedious procedures. Enabled by a novel ‘airway resistance’ based selective surface treatment, this study presents an easy and facile generation of double emulsions for the initiation and cultivation of multicellular spheroids in a scaffold-free format. Combining with our previously developed DNA nanosensors, intestinal spheroids produced in the double emulsions have shown an elevated activities of an essential DNA modifying enzyme, the topoisomerase I. The observed molecular and functional characteristics of spheroids produced in double emulsions are similar to the counterparts produced by the commercially available ultra-low attachment plates. However, the double emulsions excel for their improved uniformity, and the consistency of the results obtained by subsequent analysis of the spheroids. The presented technique is expected to ease the burden of producing spheroids and to promote the spheroids model for cancer or stem cell study.