A Microfluidic Strategy for Controllable Generation of Water‐in‐Water Droplets as Biocompatible Microcarriers

Droplet microfluidics has been widely applied in functional microparticles fabricating, tissue engineering, and drug screening due to its high throughput and great controllability. However, most of the current droplet microfluidics are dependent on water‐in‐oil (W/O) systems, which involve organic reagents, thus limiting their broader biological applications. In this work, a new microfluidic strategy is described for controllable and high‐throughput generation of monodispersed water‐in‐water (W/W) droplets. Solutions of polyethylene glycol and dextran are used as continuous and dispersed phases, respectively, without any organic reagents or surfactants. The size of W/W droplets can be precisely adjusted by changing the flow rate of dispersed and continuous phases and the valve switch cycle. In addition, uniform cell‐laden microgels are fabricated by introducing the alginate component and rat pancreatic islet (β‐TC6) cell suspension to the dispersed phase. The encapsulated islet cells retain high viability and the function of insulin secretion after cultivation for 7 days. The high‐throughput droplet microfluidic system with high biocompatibility is stable, controllable, and flexible, which can boost various chemical and biological applications, such as bio‐oriented microparticles synthesizing, microcarriers fabricating, tissue engineering, etc.     

Responsive Porous Microcarriers With Controllable Oxygen Delivery for Wound Healing

Microcarriers with oxygen‐delivering capacity have attracted increasing interest in the field of tissue regeneration. Here, a kind of molybdenum disulfide quantum dots (MoS₂ QDs) integrated responsive porous microcarriers with controllable oxygen‐delivering ability for wound healing is presented. The specific gelatin methacryloyl (GelMa) porous microcarriers are derived from inverse opal microparticles which can be decorated with the oxygen‐carrying protein hemoglobin. Because of their characteristic porous structure, interconnected nanochannels, and excellent biocompatibility, the resultant microcarriers could carry oxygen extensively and provide support for tissue repair physically and biologically. Besides, since the typical photothermal effect of 2D materials and their derived 2D QDs, the inverse opal particles integrated with MoS₂ QDs are imparted with photo‐responsive capacity, which makes them able to release oxygen photo‐controllably. It is demonstrated that the designed microcarriers can promote the repair of abdominal wall defects effectively with their multifunctional features. These remarkable properties point to the potential value of the microcarriers in wound healing and tissue engineering.     

Preparation and In Vitro Properties of N-Succinylchitosan- or Carboxymethylchitin-Mitomycin C Conjugate Microparticles with Specified Size

The preparation of cross-linked conjugate microparticles of N succinyl-chitosan (Suc) or 6-O-carboxymethylchitin (CM) with mitomycin C (MMC), which showed an adequate size for liver targeting (0.2-3 μm), was attempted by a combination of water-soluble carbodiimide (EDC) coupling and emulsification technique. As for Suc, microparticles with a diameter less than a few micrometers could be obtained easily, while the preparation of CM microparticles (CM-MPs) of the same diameter was not necessarily easy. First, preparation conditions were compared for CM-MPs, and some conditions gave CM-MPs with a diameter less than a few micrometers. As to CM-MMC conjugate microparticles, the method by addition of EDC after emulsification using CM with low molecular weight (CM_L) gave more appropriate microparticles with a mean diameter of 0.97 μm (CM_L-MP-MMC). Suc-MMC conjugate microparticles adequate for liver targeting could be produced by the addition of EDC both before and after emulsification; especially, the conjugate microparticles with a mean diameter of 0.45 μm (Suc-MP-MMC) were derived by the addition of EDC before emulsification. Suc-MP-MMC exhibited a higher drug content than CM_L-MP-MMC. CM_L-MP-MMC and Suc-MP-MMC exhibited 50% drug release times of 2.87 h and 42.1 h, respectively.     

Preparation and In Vitro Properties of N-Succinylchitosan– or Carboxymethylchitin–Mitomycin C Conjugate Microparticles with Specified Size

The preparation of cross-linked conjugate microparticles of N succinyl-chitosan (Suc) or 6-O-carboxymethylchitin (CM) with mitomycin C (MMC), which showed an adequate size for liver targeting (0.2–3 μm), was attempted by a combination of water-soluble carbodiimide (EDC) coupling and emulsification technique. As for Suc, microparticles with a diameter less than a few micrometers could be obtained easily, while the preparation of CM microparticles (CM-MPs) of the same diameter was not necessarily easy. First, preparation conditions were compared for CM-MPs, and some conditions gave CM-MPs with a diameter less than a few micrometers. As to CM-MMC conjugate microparticles, the method by addition of EDC after emulsification using CM with low molecular weight (CM_L) gave more appropriate microparticles with a mean diameter of 0.97 μm (CM_L-MP-MMC). Suc-MMC conjugate microparticles adequate for liver targeting could be produced by the addition of EDC both before and after emulsification; especially, the conjugate microparticles with a mean diameter of 0.45 μm (Suc-MP-MMC) were derived by the addition of EDC before emulsification. Suc-MP-MMC exhibited a higher drug content than CM_L-MP-MMC. CM_L-MP-MMC and Suc-MP-MMC exhibited 50% drug release times of 2.87 h and 42.1 h, respectively.     

Compound‐Droplet‐Pairs‐Filled Hydrogel Microfiber for Electric‐Field‐Induced Selective Release

The separate co‐encapsulation and selective controlled release of multiple encapsulants in a predetermined sequence has potentially important applications for drug delivery and tissue engineering. However, the selective controlled release of distinct contents upon one triggering event for most existing microcarriers still remains challenging. Here, novel microfluidic fabrication of compound‐droplet‐pairs‐filled hydrogel microfibers (C‐Fibers) is presented for two‐step selective controlled release under AC electric field. The parallel arranged compound droplets enable the separate co‐encapsulation of distinct contents in a single microfiber, and the release sequence is guaranteed by the discrepancy of the shell thickness or core conductivity of the encapsulated droplets. This is demonstrated by using a high‐frequency electric field to trigger the first burst release of droplets with higher conductivity or thinner shell, followed by the second release of the other droplets under low‐frequency electric field. The reported C‐Fibers provide novel multidelivery system for a wide range of applications that require controlled release of multiple ingredients in a prescribed sequence.     

A New Approach to In‐Situ “Micromanufacturing”: Microfluidic Fabrication of Magnetic and Fluorescent Chains Using Chitosan Microparticles as Building Blocks

An in situ microfluidic assembly approach is described that can both produce microsized building blocks and assemble them into complex multiparticle configurations in the same microfluidic device. The building blocks are microparticles of the biopolymer chitosan, which is intentionally selected because its chemistry allows for simultaneous intraparticle and interparticle linking. Monodisperse chitosan‐bearing droplets are created by shearing off a chitosan solution at a microfluidic T‐junction with a stream of hexadecane containing a nonionic detergent. These droplets are then interfacially crosslinked into stable microparticles by a downstream flow of glutaraldehyde (GA). The functional properties of these robust microparticles can be easily varied by introducing various payloads, such as magnetic nanoparticles and/or fluorescent dyes, into the chitosan solution. The on‐chip connection of such individual particles into well‐defined microchains is demonstrated using GA again as the chemical “glue” and microchannel confinement as the spatial template. Chain flexibility can be tuned by adjusting the crosslinking conditions: both rigid chains and semiflexible chains are created. Additionally, the arrangement of particles within a chain can also be controlled, for example, to generate chains with alternating fluorescent and nonfluorescent microparticles. Such microassembled chains could find applications as microfluidic mixers, delivery vehicles, microscale sensors, or miniature biomimetic robots.     

Microfluidics for Production of Particles: Mechanism,Methodology, and Applications

In the past two decades, microfluidics‐based particle production is widely applied for multiple biological usages. Compared to conventional bulk methods, microfluidic‐assisted particle production shows significant advantages, such as narrower particle size distribution, higher reproducibility, improved encapsulation efficiency, and enhanced scaling‐up potency. Herein, an overview of the recent progress of the microfluidics technology for nano‐, microparticles or droplet fabrication, and their biological applications is provided. For both nano‐, microparticles/droplets, the previously established mechanisms behind particle production via microfluidics and some typical examples during the past five years are discussed. The emerging interdisciplinary technologies based on microfluidics that have produced microparticles or droplets for cellular analysis and artificial cells fabrication are summarized. The potential drawbacks and future perspectives are also briefly discussed.     

Precision emulsification for droplet and capsule production

Emulsions and microcapsules are typical structures in various dispersion formulations for pharmaceutical, food, personal and house care applications. Precise control over size and size distribution of emulsion droplets and microcapsules are important for effective use and delivery of active components and better product quality. Many emulsification technologies have been developed to meet different formulation and processing requirements. Among them, membrane and microfluidic emulsification as emerging technologies have the feature of being able to precisely manufacture droplets in a drop-by-drop manner to give subscribed sizes and size distributions with lower energy consumption. This paper reviews fundamental sciences and engineering aspects of emulsification, membrane and microfluidic emulsification technologies and their use for precision manufacture of emulsions for intensified processing. Generic application examples are given for single and double emulsions and microcapsules with different structure features.     

Sequential Coalescence Enabled Two‐Step Microreactions in Triple‐Core Double‐Emulsion Droplets Triggered by an Electric Field

Advances in microfluidic emulsification have enabled the generation of exquisite multiple‐core droplets, which are promising structures to accommodate microreactions. An essential requirement for conducting reactions is the sequential coalescence of the multiple cores encapsulated within these droplets, therefore, mixing the reagents together in a controlled sequence. Here, a microfluidic approach is reported for the conduction of two‐step microreactions by electrically fusing three cores inside double‐emulsion droplets. Using a microcapillary glass device, monodisperse water‐in‐oil‐in‐water droplets are fabricated with three compartmented reagents encapsulated inside. An AC electric field is then applied through a polydimethylsiloxane chip to trigger the sequential mixing of the reagents, where the precise sequence is guaranteed by the discrepancy of the volume or conductivity of the inner cores. A two‐step reaction in each droplet is ensured by two times of core coalescence, which totally takes 20–40 s depending on varying conditions. The optimal parameters of the AC signal for the sequential fusion of the inner droplets are identified. Moreover, the capability of this technique is demonstrated by conducting an enzyme‐catalyzed reaction used for glucose detection with the double‐emulsion droplets. This technique should benefit a wide range of applications that require multistep reactions in micrometer scale.     

A Novel Acoustomicrofluidic Nebulization Technique Yielding New Crystallization Morphologies

A novel acoustic microfluidic nebulization platform is demonstrated, which, due to its unique ability to access intermediate evaporation rate regimes—significantly faster than that in slow solvent evaporation but considerably below that achieved in spray drying, is capable of producing novel crystal morphologies that have yet to be reported in both model inorganic and organic systems. In addition, the potential for simultaneously encapsulating single crystals within a biodegradable polymeric coating in a single simultaneous step together with the crystallization process as the solvent evaporates during nebulization is briefly shown. The platform not only has the potential to be highly scalable by employing a large number of these low‐cost miniature devices in parallel to achieve industrially relevant particle production rates, but could also be advantageous over conventional spray drying in terms of energy utilization, given the tremendous efficiency associated with the high‐frequency ultrasonic microdevice as well as its ambient temperature operation.     

Capillary‐Assisted Fabrication of Biconcave Polymeric Microlenses from Microfluidic Ternary Emulsion Droplets

In this study, a simple capillary‐based approach for producing biconcave polymeric microlenses with uniform size and shape from ternary emulsion droplets is presented. Monodisperse ternary emulsion droplets (0.6–4.0 nL) are produced which contain a photocurable segment of an acrylate monomer and two non‐curable segments of silicone oil (SO) by using a microfluidic sheath‐flowing droplet generator on a glass chip. The curvature radius of the interfaces separating the droplet segments, as well as the droplet size, and production rate can be flexibly varied by changing the flow conditions of the organic and aqueous phases. Subsequently, off‐chip suspension photopolymerization yields non‐spherical polymeric microparticles with two spherical concave surfaces templated by two SO segments at random positions. By ultraviolet light irradiation of ternary droplets with two SO segments trapped by the interior wall of a cylindrical microcapillary (internal diameter: 130 μm), biconcave microlenses can be produced with two spherical concave surfaces with a common lens axis. The produced lenses are suitable for use as optical diverging lenses.     

Cellulose‐Based Microparticles for Magnetically Controlled Optical Modulation and Sensing

Responsive materials with birefringent optical properties have been exploited for the manipulation of light in several modern electronic devices. While electrical fields are often utilized to achieve optical modulation, magnetic stimuli may offer an enticing complementary approach for controlling and manipulating light remotely. Here, the synthesis and characterization of magnetically responsive birefringent microparticles with unusual magneto‐optical properties are reported. These functional microparticles are prepared via a microfluidic emulsification process, in which water‐based droplets are generated in a flow‐focusing device and stretched into anisotropic shapes before conversion into particles via photopolymerization. Birefringence properties are achieved by aligning cellulose nanocrystals within the microparticles during droplet stretching, whereas magnetic responsiveness results from the addition of superparamagnetic nanoparticles to the initial droplet template. When suspended in a fluid, the microparticles can be controllably manipulated via an external magnetic field to result in unique magneto‐optical coupling effects. Using a remotely actuated magnetic field coupled to a polarized optical microscope, these microparticles can be employed to convert magnetic into optical signals or to estimate the viscosity of the suspending fluid through magnetically driven microrheology.     

Fast Dynamic Visualizations in Microfluidics Enabled by Fluorescent Carbon Nanodots

Microfluidic systems have become a superior platform for explorations of fascinating fluidic physics at microscale as well as applications in biomedical devices, chemical reactions, drug delivery, etc. Exploitations of this platform are built upon the fundamental techniques of flow visualizations. However, the currently employed fluorescent materials for microfluidic visualization are far from satisfaction, which severely hinders their widespread applications. Here fluorescent carbon nanodots are documented as a game‐changer, applicable in versatile fluidic environment for the visualization in microfluidics with unprecedented advantages. One of the fastest fluorescent imaging speeds up to 2500 frames per second under a normal contionous wave (CW) laser line is achieved by adopting carbon nanodots in microfluidics. Besides better visualizations of the fluid or interface, fluorescent carbon nanodots‐based microparticles enable quantitative studies of high speed dynamics in fluids at microscale with a more than 90% lower cost, which is inaccessible by traditionally adopted fluorescent dye based seeding particles. The findings hold profound influences to microfluidic investigations and may even lead to revolutionary changes to the relevant industries.     

Toward Efficient Wound Management: Bioinspired Microfluidic and Microneedle Patch (Small 3/2023)

Microneedle (MN) patches hold demonstrated prospects in intelligent wound management. Herein, inspired by the highly folded structure of insect wings, a three-dimensional (3D) origami MN patch with superfine miniature needle structures, microfluidic channels, and multiple functions was reported to detect biomarkers, release drugs controllably and monitor motions to facilitate wound healing. By simply replicating the pre-stretched silicone rubber (Ecoflex) molds patterned by a laser engraving machine, the superfine structure MN patch with microfluidic channels was obtained from the restored molds. The bioinspired origami structure endows the MN patch with a high degree of functional integration, including microfluidic channels and electrocircuits. The microfluidic channels combined with the pH value and glucose concentration indicators enable the patch with the capability of biomarker sensing detection. Porous structures, a temperature-responsive hydrogel, and a photothermal-sensitive agent are utilized to form a controllable drug release system on the MN patch. Meanwhile, MXene electrocircuits were printed on the MN patch for motion sensing. In addition, the ability of the MN patch to accelerate wound healing was demonstrated by a mouse model experiment with full-thickness skin wounds. These results indicate that the multifunctional 3D origami MN patch is a valuable intelligent strategy for wound management.     

Design and Development of Biomimetic Nanovesicles Using a Microfluidic Approach

The advancement of nanotechnology toward more sophisticated bioinspired approaches has highlighted the gap between the advantages of biomimetic and biohybrid platforms and the availability of manufacturing processes to scale up their production. Though the advantages of transferring biological features from cells to synthetic nanoparticles for drug delivery purposes have recently been reported, a standardizable, batch‐to‐batch consistent, scalable, and high‐throughput assembly method is required to further develop these platforms. Microfluidics has offered a robust tool for the controlled synthesis of nanoparticles in a versatile and reproducible approach. In this study, the incorporation of membrane proteins within the bilayer of biomimetic nanovesicles (leukosomes) using a microfluidic‐based platform is demonstrated. The physical, pharmaceutical, and biological properties of microfluidic‐formulated leukosomes (called NA‐Leuko) are characterized. NA‐Leuko show extended shelf life and retention of the biological functions of donor cells (i.e., macrophage avoidance and targeting of inflamed vasculature). The NA approach represents a universal, versatile, robust, and scalable tool, which is extensively used for the assembly of lipid nanoparticles and adapted here for the manufacturing of biomimetic nanovesicles.     

Processing of nanolitre liquid plugs for microfluidic cell-based assays

Plugs, i.e. droplets formed in a microchannel, may revolutionize microfluidic cell-based assays. This study describes a microdevice that handles nanolitre-scale liquid plugs for the preparation of various culture setups and subsequent cellular assays. An important feature of this mode of liquid operation is that the recirculation flow generated inside the plug promotes the rapid mixing of different solutions after plugs are merged, and it keeps cell suspensions homogeneous. Thus, serial dilutions of reagents and cell suspensions with different cell densities and cell types were rapidly performed using nanolitres of solution. Cells seeded through the plug processing grew well in the microdevice, and subsequent plug processing was used to detect the glucose consumption of cells and cellular responses to anticancer agents. The plug-based microdevice may provide a useful platform for cell-based assay systems in various fields, including fundamental cell biology and drug screening applications.     

Microalgae-Based Biohybrid Microrobot for Accelerated Diabetic Wound Healing

A variety of wound healing platforms have been proposed to alleviate the hypoxic condition and/or to modulate the immune responses for the treatment of chronic wounds in diabetes. However, these platforms with the passive diffusion of therapeutic agents through the blood clot result in the relatively low delivery efficiency into the deep wound site. Here, a microalgae-based biohybrid microrobot for accelerated diabetic wound healing is developed. The biohybrid microrobot autonomously moves at velocity of 33.3 µm s⁻¹ and generates oxygen for the alleviation of hypoxic condition. In addition, the microrobot efficiently bound with inflammatory chemokines of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) for modulating the immune responses. The enhanced penetration of microrobot is corroborated by measuring fibrin clots in biomimetic wound using microfluidic devices and the enhanced retention of microrobot is confirmed in the real wounded mouse skin tissue. After deposition on the chronic wound in diabetic mice without wound dressing, the wounds treated with microrobots are completely healed after 9 days with the significant decrease of inflammatory cytokines below 31% of the control level and the upregulated angiogenesis above 20 times of CD31⁺ cells. These results confirm the feasibility of microrobots as a next-generation platform for diabetic wound healing.     

Preparation of Monodisperse Biodegradable Polymer Microparticles Using a Microfluidic Flow‐Focusing Device for Controlled Drug Delivery

Degradable microparticles have broad utility as vehicles for drug delivery and form the basis of several therapies approved by the US Food and Drug Administration. Conventional emulsion‐based methods of manufacturing produce particles with a wide range of diameters (and thus kinetics of release) in each batch. This paper describes the fabrication of monodisperse, drug‐loaded microparticles from biodegradable polymers using the microfluidic flow‐focusing (FF) devices and the drug‐delivery properties of those particles. Particles are engineered with defined sizes, ranging from 10 µm to 50 µm. These particles are nearly monodisperse (polydispersity index = 3.9%). A model amphiphilic drug (bupivacaine) is incorporated within the biodegradable matrix of the particles. Kinetic analysis shows that the release of the drug from these monodisperse particles is slower than that from conventional methods of the same average size but a broader distribution of sizes and, most importantly, exhibit a significantly lower initial burst than that observed with conventional particles. The difference in the initial kinetics of drug release is attributed to the uniform distribution of the drug inside the particles generated using the microfluidic methods. These results demonstrate the utility of microfluidic FF for the generation of homogenous systems of particles for the delivery of drugs.     

Processing of nanolitre liquid plugs for microfluidic cell-based assays

Abstract

Plugs, i.e. droplets formed in a microchannel, may revolutionize microfluidic cell-based assays. This study describes a microdevice that handles nanolitre-scale liquid plugs for the preparation of various culture setups and subsequent cellular assays. An important feature of this mode of liquid operation is that the recirculation flow generated inside the plug promotes the rapid mixing of different solutions after plugs are merged, and it keeps cell suspensions homogeneous. Thus, serial dilutions of reagents and cell suspensions with different cell densities and cell types were rapidly performed using nanolitres of solution. Cells seeded through the plug processing grew well in the microdevice, and subsequent plug processing was used to detect the glucose consumption of cells and cellular responses to anticancer agents. The plug-based microdevice may provide a useful platform for cell-based assay systems in various fields, including fundamental cell biology and drug screening applications.     

Formulation and characterization of ethylcellulose microparticles containing .l-alanyl- l-glutamine peptide

Context: The l-alanyl-l-glutamine peptide (AGP) has been effective to promote acute glycemia recovery during long-term insulin-induced hypoglycemia (IIH), and the oral administration of AGP is suggested to prevent prolonged hypoglycemia, such as nocturnal hypoglycemia.

Objective: Considering the ability of AGP on glycemia recovery and AGP’s fast metabolism, the aim of current study was to obtain and characterize ethylcellulose microparticles to deliver the drug for a prolonged time.

Materials and Methods: Microparticles were prepared by simple and double emulsification/hardening method and characterized by scanning electron microscopy, thermogravimetry (TG), differential scanning calorimetry (DSC), Fourier transform infra-red (FTIR) and FT-Raman spectroscopy and in vitro release.

Results and Discussion: Spherical structures with a mean diameter between 9.30?µm and 13.19?µm were formed. TG analysis showed that the thermal stability of AGP was even more increased by encapsulation with ethylcellulose. In addition, TG, DSC, FTIR and FT-Raman analyses proved that AGP was encapsulated in a molecular way. Higher values of encapsulation efficiency were observed for the microparticles prepared by double emulsification (57.83–83.67%) than for those prepared by simple emulsification (18.37%). However, the last ones could release the peptide in a quicker and more extensive manner than those prepared by double emulsification.

Conclusion: For the first time, microparticles containing AGP were developed and exhibited prolonged in vitro release as well as protection to the drug, and it could be considered as a dosage form for patients who suffer from insulin-induced hypoglycemia and/or nocturnal hypoglycemia.