Method for driving liquid drops to move in aqueous two-phase system and application of method

Abstract

The invention discloses a method for driving liquid drops to move in an aqueous two-phase system and application of the method, and the method comprises the following steps: dissolving polyethylene glycol and glucan in deionized water, centrifuging and standing for layering to respectively obtain a PEG solution and a DEX solution; a DEX solution is dropwise added to different positions of the PEG solution containing SDBS, and two DEX liquid drops are formed; dropwise adding a DEX solution containing calcium chloride between the two DEX liquid drops to form DEX liquid drops containing calcium chloride; the two DEX liquid drops can move towards the DEX liquid drop containing calcium chloride and are finally gathered together; or dropwise adding a DEX solution containing calcium chloride to different positions of the PEG solution containing SDBS to form two DEX liquid drops containing calcium chloride; the two DEX liquid drops containing calcium chloride can move towards each other and finally gather together. The method can break through the limitation of low interfacial tension of an aqueous two-phase system, realizes controllable movement of liquid drops, has biocompatibility, and can be used for conveying biochemical reagents and initiating an enzymatic reaction.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of chemical engineering and technology, and particularly relates to a method for driving droplet movement in a double aqueous phase system and application thereof. BACKGROUND

[0002] In the fields of chemical and biological detection, drug delivery, microfluidic chip, etc., as the carrier of chemical and biological reagents, the controllable movement of droplets is of great significance for realizing accurate transportation of reagents, efficient completion of biochemical reactions and targeted delivery, and is one of the core links for promoting the development of related technologies towards miniaturization, intelligentization and high efficiency. At present, the movement control technology of droplets in fluid has become a research hotspot in the cross fields of biomedical engineering and micro-nano manufacturing, and the optimization of the technology directly affects the efficiency of biochemical analysis, the safety of drug delivery and the feasibility of industrial application.

[0003] In the prior art, the movement control of droplets depends on the organic phase-water phase binary system, which is specifically manifested in two mainstream modes of movement of organic droplets in water phase or movement of water droplets in organic phase. Such system realizes the driving and control of droplets by regulating the interfacial properties between the two phases, and has been widely used in early droplet carrier technology. However, such system generally has a core defect: it must rely on organic solvents to construct a non-aqueous phase environment, and most of the organic solvents have certain biological toxicity, which can easily damage the structure and functional integrity of bioactive reagents such as enzymes, proteins and cells, resulting in the decrease of biochemical reaction efficiency or even failure. This defect greatly limits the application of such system in the fields of biomedicine, clinical diagnosis and drug delivery which have very high requirements for biological compatibility, and cannot meet the mild transportation and reaction requirements of bioactive substances.

[0004] To solve the problem of biological toxicity caused by organic solvents, the double aqueous phase system has gradually become an optimal solution to replace the organic phase-water phase system. The double aqueous phase system is a binary aqueous phase system obtained by phase separation of a homogeneous aqueous solution formed by mixing two polymers, one polymer and one lyophile salt, or two salts at a suitable concentration ratio or under specific temperature conditions, and both phases are aqueous phase environment, without the need to introduce organic solvents, and have excellent biological compatibility, which can effectively protect the structure and function of bioactive substances, and show unique advantages in the fields of biological macromolecule separation and biochemical reaction carrier.

[0005] However, the double aqueous phase system itself has inherent technical bottlenecks: since both phases are aqueous phase, the interfacial tension is usually very low, much lower than that of the organic phase-water phase system. This low interfacial tension characteristic makes it difficult to use the traditional method of regulating droplet movement by changing the interfacial tension, and it is impossible to realize the efficient and controllable movement of double aqueous phase droplets.

[0006] Therefore, developing a droplet-driven system that can overcome the limitations of low interfacial tension in aqueous two-phase systems, achieve controllable droplet movement, and possess both biocompatibility and practical applicability has become a pressing technical problem for those skilled in the art. A breakthrough in this technology will provide novel solutions for scenarios such as the gentle transport of chemical and biological reagents, efficient biochemical reactions, and targeted drug delivery, possessing significant technological value and broad application prospects. Summary of the Invention

[0007] To address the aforementioned technical problems, this invention provides a method for driving droplet motion in an aqueous two-phase system and its application. This method overcomes the limitations of low interfacial tension in aqueous two-phase systems, enabling controllable droplet motion, and also possesses biocompatibility, allowing it to be used to transport biochemical reagents and initiate enzymatic reactions.

[0008] The technical solution adopted in this invention is as follows:

[0009] A method for driving droplet motion in an aqueous two-phase system, the method comprising the following steps:

[0010] (1) Dissolve polyethylene glycol and dextran in deionized water, centrifuge and allow to stand to separate the layers. The upper layer is the PEG solution layer and the lower layer is the DEX solution layer. Separate the upper and lower layers to obtain the PEG solution and DEX solution respectively.

[0011] (2) Sodium dodecylbenzenesulfonate was added to the PEG solution to obtain a PEG solution containing SDBS;

[0012] (3) DEX solution is added to different positions of the PEG solution containing SDBS to form two DEX droplets; DEX solution containing calcium chloride is added between the two DEX droplets to form a DEX droplet containing calcium chloride; the two DEX droplets will move toward the DEX droplet containing calcium chloride and eventually gather together.

[0013] Furthermore, step (3) can be replaced by: adding a DEX solution containing calcium chloride to different positions of the PEG solution containing SDBS to form two DEX droplets containing calcium chloride; the two DEX droplets containing calcium chloride will move toward each other and eventually gather together.

[0014] In step (1), the mass fractions of polyethylene glycol and dextran in deionized water are 2-6% and 5-10%, respectively, preferably 4% and 8%, respectively.

[0015] The polyethylene glycol has a molecular weight of 30,000 to 50,000, preferably 35,000. The glucose has a molecular weight of 300,000 to 800,000, preferably 500,000.

[0016] The settling time is 8-12 hours. After sufficient settling, the solution separates into two layers of similar volume.

[0017] In step (2), the concentration of sodium dodecylbenzenesulfonate in the PEG solution containing SDBS is 5~10 mM, preferably 7 mM.

[0018] In step (3), the DEX solution containing calcium chloride is obtained by adding calcium chloride to the DEX solution.

[0019] In step (3), the concentration of calcium chloride in the DEX solution containing calcium chloride is 0.1~0.5 mM, preferably 0.1 mM.

[0020] In step (3), the volume of the solution added is 2~5 μL, preferably 2 μL.

[0021] The present invention also provides the application of the method for driving droplet motion in a two-phase aqueous system in chemical and biological detection or drug delivery.

[0022] This invention provides two methods for driving droplet motion in an aqueous two-phase system. Both methods require first dissolving polyethylene glycol (PEG) and dextran in deionized water, followed by centrifugation and settling to separate the layers. The upper layer is a PEG solution layer, and the lower layer is a DEX solution layer. The two layers are then separated to obtain the PEG solution and the DEX solution, respectively. The PEG-DEX aqueous two-phase system exhibits excellent phase separation stability, maintaining the independence of the two phases for a long period after centrifugation, providing a stable environment for droplet formation and motion. Sodium dodecylbenzenesulfonate is then added to the PEG solution to obtain a PEG solution containing SDBS. The addition of SDBS to the PEG phase can regulate the surface tension distribution at the phase interface. Then, either method one or method two can be performed.

[0023] Method 1: Add DEX solution to different positions in a PEG solution containing SDBS to form two DEX droplets; add a DEX solution containing calcium chloride between the two DEX droplets to form a DEX droplet containing calcium chloride; the two DEX droplets will move towards the DEX droplet containing calcium chloride and eventually aggregate together. Method 2: Add a DEX solution containing calcium chloride to different positions in a PEG solution containing SDBS to form two DEX droplets containing calcium chloride; the two DEX droplets containing calcium chloride will move towards each other and eventually aggregate together.

[0024] In Method 1, the calcium chloride in the DEX droplet containing calcium chloride diffuses upwards to the solution surface, interacts with the surfactant SDBS, further reducing the interfacial tension and forming the Marangoni effect. This, in turn, creates eddies around the droplet. The DEX droplet within these eddies, hindering the flow of the eddies, is subjected to a force by the eddies, causing it to move towards the calcium chloride-containing DEX droplet and eventually accumulate around it. Figure 1 As shown.

[0025] In Method 2, the calcium chloride in two DEX droplets containing calcium chloride diffuses upwards to the solution surface, interacts with the surfactant SDBS, further reduces the interfacial tension, and forms the Marangoni effect, which in turn creates eddies around the droplets. Since both droplets contain calcium chloride, eddies can be generated around both droplets. The two droplets are propelled by each other's eddies and move towards each other simultaneously, eventually converging together.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] This invention provides a method for driving droplet motion in an aqueous two-phase system. Based on an all-aqueous system, it uses PEG and DEX, two biocompatible polymers, as the basic components of the aqueous two-phase system. Only two conventional reagents, SDBS and calcium chloride, are introduced, eliminating the need for any organic solvents. This fundamentally avoids the toxic damage to bioactive substances caused by organic solvents. PEG and DEX are both inert polymers commonly used in the biomedical field, and they do not damage the structural and functional integrity of bioactive reagents such as enzymes, proteins, and cells. SDBS, as a commonly used surfactant, is compatible with biological systems at appropriate concentrations. Calcium chloride is a naturally occurring ionic component in living organisms and will not trigger biotoxic reactions. This all-aqueous design enables the lossless carrying and transport of bioactive reagents, ensuring the efficient conduct of subsequent biochemical reactions and solving the pain point that traditional organic phase systems cannot adapt to highly biocompatible scenarios.

[0028] The preparation of the aqueous two-phase system can be completed simply by dissolving, centrifuging, and allowing the phases to separate. The phase separation effect is stable, and high-purity PEG and DEX phases can be obtained quickly. The addition of SDBS is a simple mixing operation, and the formation and aggregation of droplets can be achieved simply by precise addition. The entire process does not require extreme conditions such as high temperature, high pressure, strong acid or alkali, and is easy to standardize and scale up.

[0029] The method for driving droplet motion in an aqueous two-phase system provided by this invention does not rely on external forces or complex interface modification. It can achieve precise control of droplet motion simply by adding reagents. This breaks through the inherent limitation of droplet driving in aqueous two-phase systems due to low interfacial tension, fills the gap in existing droplet controllable motion technology in aqueous two-phase systems, and transforms the biocompatibility advantage of aqueous two-phase systems into practical application capabilities.

[0030] The method for driving droplet motion in an aqueous two-phase system provided by this invention shows promising application prospects in the fields of chemical and biological detection and drug delivery. In chemical and biological detection, droplets can serve as independent reagent carriers, enabling precise separation and directional transport of different detection reagents. Controllable aggregation facilitates precise mixing and reaction of reagents, avoiding errors caused by reagent diffusion in traditional detection methods, thus improving detection sensitivity and accuracy. Simultaneously, the all-aqueous system protects the activity of the sample, making it suitable for biological detection scenarios such as immunoassay and enzyme-catalyzed reactions. In drug delivery, drugs can be loaded into droplets, and the directional aggregation and targeted delivery of drug carriers can be achieved by controlling the droplet position, reducing drug diffusion at non-target sites and improving drug utilization. Attached Figure Description

[0031] Figure 1 This is a schematic diagram illustrating the mechanism of the method for driving droplet motion in a two-phase aqueous system provided by the present invention;

[0032] Figure 2 This is a schematic diagram of droplet motion in Example 1;

[0033] Figure 3 This is a schematic diagram of the Marangoni flow in the solution after polystyrene tracer particles were added to the solution in step (3) of Example 1.

[0034] Figure 4 The graph shows the changes in surface tension of the PEG solution containing SDBS in step (2) of Example 1 after adding different concentrations of CaCl2.

[0035] Figure 5 This is a schematic diagram of droplet motion in Example 2;

[0036] Figure 6 This is a schematic diagram of the Marangoni flow in the solution after polystyrene tracer particles were added to the solution in step (3) of Example 2.

[0037] Figure 7 This is a schematic diagram of the droplet movement and enzymatic reaction in Application Example 1. Detailed Implementation

[0038] The present invention will now be described in detail with reference to the embodiments.

[0039] The polyethylene glycol used in the examples has a molecular weight of 35,000, and the dextran has a molecular weight of 500,000.

[0040] Example 1

[0041] A method for driving droplet motion in an aqueous two-phase system, the method comprising the following steps:

[0042] (1) Dissolve polyethylene glycol and dextran in deionized water to make their mass fractions 4% and 8% respectively. Centrifuge the solution and let it stand overnight. After standing, the solution is divided into two layers of similar volume. The upper layer is the PEG solution layer and the lower layer is the DEX solution layer. Separate the upper and lower layers of solution to obtain PEG solution and DEX solution respectively. The PEG solution contains only trace amounts of DEX, and the mass fraction of DEX is <0.005%. The DEX solution contains only trace amounts of PEG, and the mass fraction of PEG is <0.01%.

[0043] (2) Sodium dodecylbenzenesulfonate was added to the PEG solution to obtain a PEG solution containing SDBS with a concentration of 7 mM.

[0044] (3) Add 2 μL of DEX solution to different positions in the PEG solution containing SDBS to form two DEX droplets; add 2 μL of DEX solution containing 0.1 mM calcium chloride between the two DEX droplets to form a DEX droplet containing calcium chloride; the two DEX droplets will move towards the DEX droplet containing calcium chloride and eventually aggregate together, the whole process is as follows. Figure 2 As shown in the figure, the position of the DEX droplet containing calcium chloride remains stationary, while both DEX droplets gradually converge towards the DEX droplet containing calcium chloride.

[0045] Adding 22-micrometer-diameter polystyrene tracer particles to the solution in step (3) allows for the visualization of Marangoni flow in the solution using particle tracking technology. Figure 3 As shown.

[0046] Different concentrations of CaCl2 were added to the PEG solution containing SDBS in step (2), and the surface tension of the solution was measured using the hanging drop method. The results are as follows: Figure 4 As shown in the figure, the surface tension of the system gradually decreases with the increase of calcium chloride concentration, and the surface tension of the system drops to the lowest level when the calcium chloride concentration reaches 10 mM.

[0047] Example 2

[0048] A method for driving droplet motion in an aqueous two-phase system, the method comprising the following steps:

[0049] (1) Same as step (1) in Example 1.

[0050] (2) Same as step (2) in Example 1.

[0051] (3) Add 2 μL of DEX solution containing 0.1 mM calcium chloride to different positions in the PEG solution containing SDBS to form two DEX droplets containing calcium chloride; the two DEX droplets containing calcium chloride will move towards each other and eventually aggregate together, the whole process is as follows. Figure 5 As shown in the figure, two DEX droplets containing calcium chloride move toward each other and eventually aggregate together.

[0052] Adding 22-micrometer-diameter polystyrene tracer particles to the solution in step (3) allows for the visualization of Marangoni flow in the solution using particle tracking technology. Figure 6 As shown.

[0053] Application Example 1

[0054] Application of the method in Example 2 in enzymatic reactions

[0055] Two DEX solutions containing 0.1 mM calcium chloride were prepared. One solution was treated with 1% (v / v) triglyceride, and the other with 1% (w / w) lipase. 2 μL of each solution was rapidly added to a PEG solution containing 7 mM SDBS, forming two droplets. The two droplets immediately moved towards each other and eventually coalesced. At the contact point, an enzymatic hydrolysis of triglyceride occurred, producing carboxylic acid, such as... Figure 7 As shown, the method for driving droplet motion in an aqueous two-phase system provided by this invention has promising applications in chemical and biological detection and drug delivery.

[0056] The above detailed description of a method for driving droplet motion in an aqueous two-phase system and its application, with reference to the embodiments, is illustrative rather than limiting. Several embodiments may be listed within the defined scope. Therefore, variations and modifications that do not depart from the overall concept of the present invention should be within the protection scope of the present invention.

Claims

1. A method for driving droplet motion in an aqueous two-phase system, characterized in that, The method includes the following steps: (1) Dissolve polyethylene glycol and dextran in deionized water, centrifuge and allow to stand to separate the layers. The upper layer is the PEG solution layer and the lower layer is the DEX solution layer. Separate the upper and lower layers to obtain the PEG solution and DEX solution respectively. (2) Sodium dodecylbenzenesulfonate was added to the PEG solution to obtain a PEG solution containing SDBS; (3) DEX solution is added to different positions of the PEG solution containing SDBS to form two DEX droplets; DEX solution containing calcium chloride is added between the two DEX droplets to form a DEX droplet containing calcium chloride; the two DEX droplets will move toward the DEX droplet containing calcium chloride and eventually gather together.

2. The method for driving droplet motion in a two-phase aqueous system according to claim 1, characterized in that, Replace step (3) with: add DEX solution containing calcium chloride to different positions of the PEG solution containing SDBS to form two DEX droplets containing calcium chloride; the two DEX droplets containing calcium chloride will move toward each other and eventually gather together.

3. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, In step (1), the mass fractions of polyethylene glycol and dextran in deionized water are 2-6% and 5-10%, respectively.

4. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, The polyethylene glycol has a molecular weight of 30,000 to 50,000; the glucose has a molecular weight of 300,000 to 800,000.

5. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, The settling time is 8-12 hours.

6. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, In step (2), the concentration of sodium dodecylbenzenesulfonate in the PEG solution containing SDBS is 5~10 mM.

7. The method for driving droplet motion in a two-phase aqueous system according to claim 1 or 2, characterized in that, In step (3), the DEX solution containing calcium chloride is obtained by adding calcium chloride to the DEX solution.

8. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, In step (3), the concentration of calcium chloride in the DEX solution containing calcium chloride is 0.1~0.5 mM.

9. The method for driving droplet motion in an aqueous two-phase system according to claim 1 or 2, characterized in that, In step (3), the volume of the solution added is 2~5 μL.

10. The application of the method for driving droplet motion in an aqueous two-phase system as described in any one of claims 1-9 in chemical and biological detection or drug delivery.

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