A multi-compartment calcium alginate gel microcarrier, its preparation method and application

Abstract

The present invention discloses a multi-compartment calcium alginate gel microcarrier, a preparation method thereof and an application thereof. In this method, multiple droplets of Dex-Alg solution are dropped into a biphasic receiving solution. After the Dex-Alg solution enters the lower phase and stands still, a multi-compartment calcium alginate gel microcarrier is obtained. The upper phase of the biphasic receiving solution is a PEG solution, and the lower phase is a Dex-CaCl2 solution. Multiple monodisperse droplets of Dex-Alg fuse at the liquid-liquid interface of the PEG solution and the Dex-CaCl2 solution in the biphasic receiving solution. After entering the lower phase, Alg crosslinks with Ca 2+ to form a multi-compartment calcium alginate gel microcarrier. The method of the present invention is easy to operate, can control the shape of the gel microcarrier by controlling the fusion of droplets, does not require the addition of organic reagents, has high biocompatibility, and is green and environmentally friendly; the controllable multi-compartment calcium alginate gel microcarrier prepared by the method of the present invention has broad application prospects.

Description

Technical Field

[0001] The present invention belongs to the technical field of drug microcarriers, and particularly relates to a multi-compartment calcium alginate gel microcarrier and a preparation method and application thereof. Background Art

[0002] The multi-compartment calcium alginate gel microcarrier is a novel material with multiple functions. Due to its characteristics such as low density, high specific surface area, and high biocompatibility, it can be used for isolated co-encapsulation and triggered release. In particular, it can precisely control the structure of multi-compartments and the encapsulation level of each component, and has a wide range of applications in the fields of 3D cell culture, controlled release of various substances, immobilized enzymes, and structural materials. During the preparation of colloidal particles, since the interfacial tension plays a dominant role in the preparation process, the resulting colloidal particles are usually spherical with the minimum surface energy. However, compared with spherical particles with a single compartment, particles with an anisotropic shape in multi-compartments have unique properties and are more widely used. For example, non-spherical particles can be packed more densely than spherical particles, and anisotropic-shaped particles also exhibit different properties from spherical particles under the same hydrodynamic, electrical, and magnetic conditions; the manufactured non-spherical structured particles have unique mechanical properties and are of great significance in medical devices and structural materials. In addition, most molecules are non-spherical, and by customizing the geometric shape of non-spherical particles, the shape of molecules can also be simulated for self-assembly research. These unique properties of non-spherical particles make up for many deficiencies of spherical particles, creating new opportunities for fields that traditionally use spherical particles and having a wider range of application fields.

[0003] In recent years, in order to prepare structures with more complex geometries, some bottom-up and top-down methods have been developed to prepare compartmentalized heterogeneous microparticles. Strategies for fabricating such non-spherical particles include seeded polymerization, self-assembly, particle aggregation, and microfluidics. However, since the properties of non-spherical particles strongly depend on their shape, methods for controlling not only the size but also the shape and the reagents used are crucial for producing particles with high biocompatibility and reproducible and uniform behavior. There are still challenges in terms of shape and size controllability and simplicity. It is difficult to produce a large number of monodisperse non-spherical particles with adjustable geometries, and the applications are also limited. For example, the seeded polymerization method is one of the commonly used methods. Emulsion polymerization is carried out again using the prepared microspheres as seeds, and the polymerization of the second monomer is initiated inside or on the surface of the seeds to achieve purposes such as improving the surface properties of the particles, increasing the particle size, and controlling the particle morphology. During the seeded polymerization process, the cross-linked seed microspheres are swollen by the monomer and heated to initiate polymerization. At the same time, the elastic force of the cross-linked network tends to squeeze out the swollen monomer, thus forming non-spherical particles. This method requires the addition of organic reagents, and it is not easy to control the shape, and the safety and controllability need to be improved. For the method of hindered coalescence of coated particle coating droplets, the monodisperse droplets coated with colloidal particles are prevented from coalescing due to the interference of the colloidal particles at the oil-water interface before complete fusion, and non-spherical structured colloids are prepared. When the droplets coalesce at the oil-water interface in this method, it will lead to a decrease in biological activity, and the suspension containing particles is unstable as a template for preparing monodisperse droplets, so the application is also limited. Summary of the Invention

[0004] In view of the above deficiencies of the prior art, the present invention provides a multi-compartment calcium alginate gel microcarrier with high biocompatibility, controllable shape, controllable compartments, and without the introduction of organic reagents and surfactants, as well as a preparation method and application thereof.

[0005] The present invention is realized through the following technical solutions:

[0006] A preparation method of a multi-compartment calcium alginate gel microcarrier, the preparation method comprising the following steps: dropping a Dex-Alg (dextran-sodium alginate) solution into a biphasic receiving liquid with an upper phase of a PEG (polyethylene glycol) solution and a lower phase of a Dex-CaCl2 solution, and after the Dex-Alg solution enters the lower phase, standing it until the droplets of the Dex-Alg solution and the Ca 2+ in the lower phase cross-link to obtain a multi-compartment calcium alginate gel microcarrier. By adopting the above technical solution, the beneficial effects of the present invention are:

[0007] According to the required shape, the droplets of the Dex-Alg solution are dropped into the biphasic receiving liquid, and then the adjacent droplets coalesce and fuse at the liquid-liquid interface of the PEG solution and the Dex-CaCl2 solution; the fused droplets sink and contact the lower-layer Dex solution containing calcium chloride, and the Ca 2+Diffuse gradually into the interior of the droplet and exchange with Na + Exchange, crosslink with Alg. Since the crosslinking and gelation process of the droplet occurs from the outside to the inside, when multiple droplets have not fully fused to form a sphere, the outer layer of the droplet has been gelated and fixed, thus stabilizing the multi-compartment gel microcarrier structure and solidifying to form a stable multi-compartment calcium alginate gel microcarrier. By arranging the dripping positions of the Dex-Alg solution in a regular array, the fusion of the Dex-Alg droplets can be controlled, thereby controlling the shape of the microcarrier. The shape of the multi-compartment calcium alginate gel microcarrier can be controlled by controlling the droplet fusion speed, crosslinking degree, and droplet shape.

[0008] Furthermore, the above-mentioned Dex-CaCl2 solution, biphasic receiving solution, and Dex-Alg solution are prepared by the following methods:

[0009] (1) Mix equal masses of PEG solution and Dex solution with the same mass fraction, and let it stand for phase separation to obtain a PEG-Dex solution system. The upper phase of the PEG-Dex solution system is the PEG solution, and the lower phase is the Dex solution. Extract the PEG solution and Dex solution respectively.

[0010] (2) Add Alg to the Dex solution obtained in step (1) to obtain a Dex-Alg solution.

[0011] (3) Add CaCl2 to the Dex solution obtained in step (1) to obtain the lower phase of the biphasic receiving solution as the Dex-CaCl2 solution, and then add the PEG solution as the upper phase of the biphasic receiving solution to obtain the biphasic receiving solution.

[0012] Furthermore, the mass percentage of PEG in the above-mentioned PEG-Dex solution system is 8-14 wt%, and the mass percentage of Dex is 8-14 wt%.

[0013] The beneficial effects of adopting the above further technical solutions are:

[0014] By changing the mass percentage of PEG in the PEG-Dex solution system to change the mass percentage of PEG in the PEG solution, the shape and fusion speed of the Dex-Alg droplets after entering the biphasic receiving solution can be changed, thereby controlling the shape of the multi-compartment calcium alginate gel microcarrier.

[0015] Furthermore, the mass percentage of Alg in the above-mentioned Dex-Alg solution is 0.2-4.0 wt%.

[0016] The beneficial effects of adopting the above further technical solutions are:

[0017] The fusion rate and crosslinking degree of Dex-Alg droplets can be controlled by changing the mass percentage of Alg in the Dex-Alg solution, thereby controlling the shape of the multi-compartment calcium alginate gel microcarriers.

[0018] Furthermore, the mass percentage of CaCl2 in the above Dex-CaCl2 solution is 5-12 wt%.

[0019] Furthermore, the height of the PEG solution in the above biphasic receiving liquid is 1-4 mm.

[0020] The beneficial effects of adopting the above further technical solutions are:

[0021] The droplet shape can be controlled by changing the height of the PEG solution in the biphasic receiving liquid, thereby controlling the shape of the multi-compartment calcium alginate gel microcarriers.

[0022] Furthermore, after the above Dex-Alg solution enters the lower phase, it is left standing for 10-25 min.

[0023] The beneficial effects of adopting the above further technical solutions are:

[0024] It can make Alg in the Dex-Alg solution crosslink fully with Ca in the Dex-CaCl2 solution 2+ to form multi-compartment calcium alginate gel microcarriers.

[0025] Furthermore, when the Dex-Alg solution is dropped into the biphasic receiving liquid, the height of the liquid droplet from the biphasic liquid surface during dropping is 2-8 mm.

[0026] The beneficial effects of adopting the above further technical solutions are:

[0027] By controlling the height of the liquid droplet from the biphasic liquid surface when the Dex-Alg solution is dropped into the biphasic receiving liquid, the droplet shape can be controlled, thereby controlling the shape of the multi-compartment calcium alginate gel microcarriers.

[0028] The multi-compartment calcium alginate gel microcarriers prepared by using the above preparation method of the multi-compartment calcium alginate gel microcarriers.

[0029] The application of the above multi-compartment calcium alginate gel microcarriers in constructing a disease model.

[0030] In summary, the beneficial effects of the present invention are:

[0031] The method of the present invention is easy to operate, controls the fusion of Dex-Alg droplets in a biphasic receiving liquid, and can control the droplet fusion speed and crosslinking degree by changing the mass percentage of Alg in the Dex-Alg solution, control the droplet shape and fusion speed by changing the height of the Dex-Alg droplet from the liquid surface of the solution, control the droplet shape and fusion speed by changing the mass percentage of PEG in the upper PEG solution of the biphasic receiving liquid, or control the droplet shape by changing the height of the upper PEG solution of the biphasic receiving liquid, so as to control the shape of the multi-compartment calcium alginate gel microcarrier; the fusion of Dex-Alg droplets in the biphasic receiving liquid of the present invention is easy to operate and can be used for the preparation of customized non-spherical multi-compartment gel microcarriers; the whole system is an all-aqueous phase system, without the addition of organic reagents, has high biocompatibility, and all reagents are non-toxic, green and environmentally friendly; the controllable multi-compartment calcium alginate gel microcarriers prepared by the present invention have broad application prospects in the fields of constructing disease models, biomedicine, cosmetics, etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 It is a schematic diagram of the principle for preparing the multi-compartment calcium alginate gel microcarrier of the present invention.

[0033] Figure 2 It is the top view (a) and side view (b) morphology diagrams of the multi-compartment calcium alginate gel microcarrier.

[0034] Figure 3 It is the overall view diagrams of the multi-compartment calcium alginate gel microcarriers prepared when the height of the dropping needle from the liquid surface of the solution is 3 mm (a), 6 mm (b), and 12 mm (c).

[0035] Figure 4 It is the overall view diagrams of the multi-compartment calcium alginate gel microcarriers prepared when the height of the upper PEG enrichment phase solution of the biphasic receiving liquid is 1.5 mm (a), 3 mm (b), and 6 mm (c).

[0036] Figure 5 It is the overall view diagrams of the multi-compartment calcium alginate gel microcarriers prepared when the mass percentage of Alg is 0.2 wt% (a), 1.0 wt% (b), and 2.0 wt% (c).

[0037] Figure 6 It is for the multi-compartment calcium alginate gel microcarriers prepared when the mass percentage of PEG in the upper solution of the biphasic receiving liquid is 8 wt% (a), 10 wt% (b), and 14 wt%

[0038] (c).

[0039] Figure 7 It is the overall view diagrams for controlling the number of droplet fusions to be 3 (a) and 4 (b). DETAILED DESCRIPTION OF THE INVENTION

[0040] Example 1

[0041] The preparation principle of multi-compartment calcium alginate gel microcarriers is as follows Figure 1 As shown, the preparation process is as follows:

[0042] (1) Preparation of PEG-Dex solution system:

[0043] a. Weigh PEG with a molecular weight of 8 kDa and add it to deionized water to prepare a 20 wt% PEG solution;

[0044] b. Weigh Dex with a molecular weight of 500 kDa and add it to deionized water to prepare a 20 wt% Dex solution;

[0045] c. Weigh the PEG solution and Dex solution obtained in step (a) and step (b) of equal mass, mix them thoroughly on a rotary incubator, let them stand for 6 hours, and then separate them into layers to obtain a PEG-Dex solution system, wherein the upper layer of the PEG-Dex solution system is the PEG solution, and the lower layer is the Dex solution. The upper layer solution and the lower layer solution are respectively extracted for use;

[0046] (2) Preparation of Dex-Alg solution: Add Alg to the Dex solution obtained in step (1) c and fully dissolve to form a Dex-Alg solution with an Alg mass percentage of 1.0 wt %;

[0047] (3) Preparation of a two-phase receiving solution: Add CaCl2 to the Dex solution obtained in step (1) c, mix and dissolve thoroughly to form a Dex-CaCl2 solution with a CaCl2 mass percentage of 10 wt% as the lower layer of the two-phase receiving solution, pour it into a receiving dish, and then add a PEG solution of the same volume as the lower layer solution as the upper layer of the two-phase receiving solution;

[0048] (4) Preparation of multi-chamber calcium alginate gel microcarriers:

[0049] a. Use a 2 mL syringe with a 0.45 mm needle, keep the syringe 3 mm away from the upper liquid surface of the solution, suspend it vertically, and push the syringe plunger to drip the Dex-Alg solution into the two-phase receiving solution obtained in step (3) at a rate of 4 seconds per drop, and drip it into the PEG solution in the upper layer of the solution, control the number of droplets and the position of the arrangement, and drip 3 Dex-Alg solution droplets in a row, or drip 3 Dex-Alg solution droplets at the three vertices of a triangle, and wait for the Dex-Alg solution to drop to the lower layer of the two-phase receiving solution and stand for 20 minutes to form a stable multi-compartment calcium alginate gel microcarrier; drip 4 Dex-Alg solution droplets in a row, or drip 4 Dex-Alg solution droplets at the four vertices of a quadrilateral, and wait for the Dex-Alg droplets to drop to the lower layer of the two-phase receiving solution and stand for 20 minutes to form a stable multi-compartment calcium alginate gel microcarrier;

[0050] b. Take a picture of the multi-compartment calcium alginate gel microcarrier obtained in step a with a digital camera. The result is as follows Figure 2 a, as shown in 2b.

[0051] Depend on Figure 2 It can be seen that the method of the present invention can be used to prepare multi-compartment calcium alginate gel microcarriers with different shapes.

[0052] Example 2

[0053] In this embodiment, the shape of the multi-compartment calcium alginate gel microcarrier is controlled by changing the height of the Dex-Alg solution droplet (i.e., the instillation needle) from the solution surface. The preparation process is as follows:

[0054] (1) Preparation of PEG-Dex solution system:

[0055] a. Weigh PEG with a molecular weight of 8 kDa and add it to deionized water to prepare a 20 wt% PEG solution;

[0056] b. Weigh Dex with a molecular weight of 500 kDa and add it to deionized water to prepare a 20 wt% Dex solution;

[0057] c. Weigh the PEG solution and Dex solution obtained in step (a) and step (b) of equal mass, mix them thoroughly on a rotary incubator, let them stand for 6 hours, and then separate them into layers to obtain a PEG-Dex solution system, wherein the upper layer of the PEG-Dex solution system is the PEG solution, and the lower layer is the Dex solution. The upper layer solution and the lower layer solution are respectively extracted for use;

[0058] (2) Preparation of Dex-Alg aqueous solution: Add Alg to the Dex solution obtained in step (1)c and dissolve it thoroughly to form a Dex-Alg solution with 1.0 wt% of Alg by mass;

[0059] (3) Preparation of biphasic receiving solution: Add CaCl2 to the Dex solution obtained in step (1)c, mix and dissolve it thoroughly to form a Dex-CaCl2 solution with 10 wt% of CaCl2 by mass as the lower layer of the biphasic receiving solution and pour it into a receiving dish. Then add a PEG solution with the same volume as the lower layer solution as the upper layer of the biphasic receiving solution;

[0060] (4) Preparation of multi-chamber calcium alginate gel microcarriers:

[0061] a. Use a 2 mL syringe equipped with a 0.45 mm needle. Keep the syringe suspended vertically at 3 mm, 6 mm, and 12 mm above the upper liquid level of the solution respectively. Push the syringe plunger to drop the Dex-Alg solution into the biphasic receiving solution obtained in step (3) at a rate of 1 drop every 4 seconds. The drops fall into the PEG solution on the upper layer of the solution. Control the number and arrangement position of the droplets. Continuously drop 2 Dex-Alg solution droplets in a row by the dropping method. After the Dex-Alg droplets drop to the lower layer of the biphasic receiving solution and stand for 20 min, stable multi-compartment calcium alginate gel microcarriers are formed;

[0062] b. Take a photo of the multi-compartment calcium alginate gel microcarriers obtained in step a with a digital camera. The results are as Figure 3 (a, b, and c) shown.

[0063] It can be seen from Figure 3 that when the height of the dropping needle above the liquid level of the solution is different, the shapes of the obtained multi-compartment calcium alginate gel microcarriers are different; when the height of the dropping needle above the liquid level of the solution is 3 mm and 6 mm, the droplets can fuse. When two droplets fuse, peanut-like double-compartment calcium alginate gel microcarriers as shown in Figure 3 a, Figure 3 b can be obtained. However, when the height of the dropping needle above the liquid level of the solution is too high, such as 12 mm, the droplets do not fuse and single calcium alginate microspheres are obtained.

[0064] Example 3

[0065] In this example, the shape of the multi-compartment calcium alginate gel microcarriers is controlled by changing the height of the PEG solution on the upper layer of the biphasic receiving solution. The specific preparation process is as follows:

[0066] (1) Preparation of PEG-Dex solution system:

[0067] a. Weigh PEG with a molecular weight of 8 kDa and add it to deionized water to prepare a PEG solution with 20 wt% by mass;

[0068] b. Weigh Dex with a molecular weight of 500 kDa and add it to deionized water to prepare a Dex solution with a mass percentage of 20 wt%.

[0069] c. Weigh equal masses of the PEG solution and Dex solution obtained in steps (a) and (b), mix them well on a rotary shaker, let it stand for 6 h and then layer. A PEG-Dex solution system is obtained. The upper layer of the PEG-Dex solution system is the PEG solution, and the lower layer is the Dex solution. Extract the upper and lower layer solutions separately for standby.

[0070] (2) Preparation of Dex-Alg aqueous solution: Add Alg to the Dex solution obtained in step (1)c and dissolve it well to form a Dex-Alg solution with an Alg mass percentage of 1.0 wt%.

[0071] (3) Preparation of biphasic receiving solution: Divide the Dex solution obtained in step (1)c into 3 groups, add CaCl2 to each group respectively, mix and dissolve it well to form a Dex-CaCl2 solution with a CaCl2 mass percentage of 10 wt% as the lower layer of the biphasic receiving solution and pour it into a receiving dish. Then add the PEG solution as the upper layer of the biphasic receiving solution to the 3 groups respectively. The heights of the PEG solutions are 1.5 mm, 3 mm and 6 mm respectively.

[0072] (4) Preparation of multi-chamber calcium alginate gel microcarriers:

[0073] a. Use a 2 mL syringe equipped with a 0.45 mm needle. Keep the syringe 3 mm above the upper liquid surface of the solution respectively, suspend it vertically, and drop the Dex-Alg solution into the biphasic receiving solution obtained in step (3) at a speed of 1 drop every 4 seconds. The drops fall into the PEG solution on the upper layer of the solution. Control the number and arrangement position of the liquid drops, and continuously drop 2 Dex-Alg solution drops in a row by the dropwise infusion method. After the Dex-Alg drops fall to the lower layer of the biphasic receiving solution and stand for 20 min, stable multi-compartment calcium alginate gel microcarriers are formed.

[0074] b. Take pictures of the multi-compartment calcium alginate gel microcarriers obtained in step a with a digital camera, and the results are as Figure 4 (a, b and c) shown.

[0075] It can be seen from Figure 4 that when the height of the PEG solution on the upper layer of the biphasic receiving solution is different, the shapes of the obtained multi-compartment calcium alginate gel microcarriers are different; when the height of the PEG-rich phase on the upper layer of the biphasic receiving solution is 1.5 mm and 3 mm, the liquid drops can all fuse. When two liquid drops fuse, the result can be as Figure 4 a, Figure 4b shows a peanut-shaped double-compartment calcium alginate gel microcarrier. When the height of the PEG-rich phase in the upper layer of the two-phase receiving liquid is too high, such as 6 mm, the droplets do not merge and single calcium alginate microspheres are obtained.

[0076] Example 4

[0077] In this embodiment, the shape of the multi-compartment calcium alginate gel microcarrier is controlled by changing the mass percentage of Alg in the Dex-Alg solution. The preparation process is as follows:

[0078] (1) Preparation of PEG-Dex solution system:

[0079] a. Weigh PEG with a molecular weight of 8 kDa and add it to deionized water to prepare a 20 wt% PEG solution;

[0080] b. Weigh Dex with a molecular weight of 500 kDa and add it to deionized water to prepare a 20 wt% Dex solution;

[0081] c. Weigh the PEG solution and Dex solution obtained in step (a) and step (b) of equal mass, mix them thoroughly on a rotary incubator, let them stand for 6 hours, and then separate them into layers to obtain a PEG-Dex solution system, wherein the upper layer of the PEG-Dex solution system is the PEG solution, and the lower layer is the Dex solution. The upper layer solution and the lower layer solution are respectively extracted for use;

[0082] (2) Preparation of Dex-Alg aqueous solution: The Dex solution obtained in step (1) was divided into three groups, and Alg was added to each group to fully dissolve the Dex-Alg solutions with Alg mass percentages of 0.2 wt%, 1.0 wt% and 4.0 wt% respectively;

[0083] (3) Preparation of a two-phase receiving solution: Add CaCl2 to the Dex solution obtained in step (1), mix and dissolve thoroughly to form a Dex-CaCl2 solution with a CaCl2 mass percentage of 10 wt% as the lower layer of the two-phase receiving solution, and pour it into a receiving dish, then add a PEG solution of the same volume as the lower layer solution as the upper layer of the two-phase receiving solution;

[0084] (4) Preparation of multi-chamber calcium alginate gel microcarriers

[0085] a. Use a 2 mL syringe with a 0.45 mm needle. Keep the syringe 3 mm above the upper liquid level of the solution, suspended vertically. Push the syringe plunger to drop Dex-Alg solutions with different mass percentages into the biphasic receiving solution obtained in step (3) at a rate of 1 drop every 4 seconds. The drops fall into the PEG solution in the upper phase of the biphasic receiving solution. Control the number and arrangement position of the droplets. Continuously drop 2 Dex-Alg solution droplets in a row by the dropping method. After the Dex-Alg droplets drop to the lower layer of the biphasic receiving solution and stand for 20 min, stable multi-compartment calcium alginate gel microcarriers are formed;

[0086] b. Take pictures of the multi-compartment calcium alginate gel microcarriers obtained in step a with a digital camera. The results are as Figure 5 (a, b, and c) shown.

[0087] It can be seen from Figure 5 that different mass percentages of Alg in the Dex-Alg solution can result in multi-compartment calcium alginate gel microcarriers with different shapes; when the mass percentage of Alg is too low, for example, when the mass percentage of Alg is 0.2 wt%, as shown in Figure 5 a, the area of the droplet fusion part is large, and the multi-compartment calcium alginate is relatively flat; when the mass percentage of Alg is increased to 1 wt%, as shown in Figure 5 b, the area of the droplet fusion part becomes smaller; and the obtained multi-compartment calcium alginate gel microcarriers are more three-dimensional; when the mass percentage of Alg is too high, for example, when the mass percentage of Alg is 4.0 wt%, as shown in Figure 5 c, the area of the droplet fusion part becomes very small, and only a small part of the droplets are fused and connected.

[0088] Example 5

[0089] In this example, the shape of the multi-compartment calcium alginate gel microcarriers is controlled by changing the concentration of PEG in the PEG solution. The specific preparation process is as follows:

[0090] (1) Preparation of the PEG-Dex solution system:

[0091] a. Weigh 3 groups of PEG with a molecular weight of 8 kDa and add them to deionized water to prepare PEG solutions with mass percentages of 16 wt%, 20 wt%, and 28 wt% respectively;

[0092] b. Weigh 3 groups of Dex with a molecular weight of 500 kDa and add them to deionized water to prepare Dex solutions with mass percentages of 16 wt%, 20 wt%, and 28 wt% respectively;

[0093] c. Respectively, the PEG solution and Dex solution with the same mass fraction obtained in steps (a) and (b) are fully mixed in equal masses on a rotary shaker, and after standing for 6 h, they are stratified to obtain 3 groups of PEG-Dex solution systems. The upper layer of the PEG-Dex solution system is the PEG solution, and the mass percentages of PEG in the PEG-Dex solution systems are 8 wt%, 10 wt%, and 14 wt% respectively; the lower layer is the Dex solution, and the mass percentages of Dex are 8 wt%, 10 wt%, and 14 wt% respectively. The upper layer solution and the lower layer solution are respectively extracted and reserved.

[0094] (2) Preparation of Dex-Alg solution: Add Alg to the Dex solution with a mass percentage of 10 wt% obtained in step (1)c, and fully dissolve it to form a Dex-Alg solution with a mass percentage of Alg of 1.0 wt%.

[0095] (3) Preparation of biphasic receiving solution: Add CaCl2 to the Dex solution with a mass percentage of 10 wt% obtained in step (1)c, and fully mix and dissolve it to form a Dex-CaCl2 solution with a mass percentage of CaCl2 of 10 wt%. Divide it into 3 groups and pour them into receiving dishes respectively as the lower layer of the biphasic receiving solution. Then, add the PEG solutions with mass percentages of 8 wt%, 10 wt%, and 14 wt% obtained in step (1)c with the same volume as the lower layer solution as the upper layer PEG solution of the biphasic receiving solution respectively as the upper layer of the biphasic receiving solution;

[0096] (4) Preparation of multi-compartment calcium alginate gel microcarriers:

[0097] a. Use a 2 mL syringe equipped with a 0.45 mm needle. Keep the syringe 3 mm above the upper liquid level of the solution respectively, suspend it vertically, and drop the Dex-Alg solution into the biphasic receiving solution obtained in step (3) at a speed of 1 drop every 4 seconds. The drops fall into the PEG solution in the upper layer of the solution. Control the number and arrangement position of the droplets, and continuously drop 2 Dex-Alg solution droplets in a row by the dropwise method. After the Dex-Alg droplets drop to the lower layer of the biphasic receiving solution and stand for 20 min, stable multi-compartment calcium alginate gel microcarriers are formed;

[0098] b. Take pictures of the multi-compartment calcium alginate gel microcarriers obtained in step a with a digital camera, and the results are as Figure 6 (a, b, and c) shown, with controllable and stable shapes.

[0099] It can be seen from Figure 6 that the shape of the multi-compartment calcium alginate gel microcarriers can be controlled by controlling the mass percentage of the PEG solution in the upper layer of the biphasic receiving solution; when the mass percentage of the PEG solution in the upper layer of the biphasic receiving solution is relatively low, such as when the mass percentage of the PEG solution is 8 wt%, asFigure 6 As shown in a, only a small part of the Dex-Alg solution droplets fuse, and most are monodisperse calcium alginate gel microspheres; when the mass percentage of the upper-layer PEG solution in the biphasic receiving solution is increased to 10 wt%, as Figure 6 shown in b, two adjacent Dex-Alg solution droplets will fuse to form calcium alginate gel microcarriers with a peanut-like structure; when the mass percentage of the upper-layer PEG solution in the biphasic receiving solution is relatively high, such as when the mass percentage of the PEG solution is 14 wt%, as Figure 6 shown in c, a trailing phenomenon appears at the fusion site of the Dex-Alg solution droplets, and two droplets fuse to obtain calcium alginate gel microcarriers in the shape of a "heart".

[0100] Example 6

[0101] In this example, the shape of the multi-compartment calcium alginate gel microcarrier is controlled by changing the number of droplet fusions. The specific preparation process is as follows:

[0102] (1) Preparation of the PEG-Dex solution system:

[0103] a. Weigh PEG with a molecular weight of 8 kDa and add it to deionized water to prepare a PEG solution with a mass percentage of 20 wt%.

[0104] b. Weigh Dex with a molecular weight of 500 kDa and add it to deionized water to prepare a Dex solution with a mass percentage of 20 wt%.

[0105] c. Weigh equal masses of the PEG solution and Dex solution obtained in steps (a) and (b), mix them thoroughly on a rotary shaker, and let them stand for 6 h to separate into layers. A PEG-Dex solution system is obtained, where the upper layer of the PEG-Dex solution system is the PEG solution and the lower layer is the Dex solution. The upper-layer solution and the lower-layer solution are respectively extracted and reserved for use;

[0106] (2) Preparation of the Dex-Alg aqueous solution: Add Alg to the Dex solution obtained in step (1) and dissolve it thoroughly to form a Dex-Alg solution with an Alg mass percentage of 1.0 wt%.

[0107] (3) Preparation of the biphasic receiving solution: Add CaCl2 to the Dex solution obtained in step (1), mix and dissolve it thoroughly to form a Dex-CaCl2 solution with a CaCl2 mass percentage of 10 wt% as the lower layer of the biphasic receiving solution and pour it into a receiving dish. Then add a PEG solution with the same volume as the lower-layer solution as the upper layer of the biphasic receiving solution;

[0108] (4) Preparation of the multi-compartment calcium alginate gel microcarrier:

[0109] a. Use a 2 mL syringe with a 0.45 mm needle. Keep the syringe 3 mm above the upper liquid level of the solution, suspended vertically. Drop the Dex-Alg solution into the biphasic receiving solution obtained in step (3) at a rate of one drop every 4 seconds by pushing the syringe plunger. The drops fall into the PEG solution on the upper layer of the solution. Control the number and arrangement positions of 3 and 4 droplets respectively. Continuously drop the droplets in a row or drop the droplets at the vertices of a polygon. After the Dex-Alg droplets drop to the lower layer of the biphasic receiving solution and stand still for 20 min, stable multi-compartment calcium alginate gel microcarriers are formed;

[0110] b. Take pictures of the multi-compartment calcium alginate gel microcarriers obtained in step a with a digital camera. The results are as shown in Figure 7 (a, b).

[0111] It can be seen from Figure 7 that multi-compartment calcium alginate gel microcarriers with different shapes can be prepared by controlling the number and arrangement positions of the droplets; continuously drop 3 droplets in a row or drop 3 droplets at the three vertices of a triangle to obtain three multi-compartment calcium alginate gel microcarriers with different shapes as shown in Figure 7 a; or continuously drop 4 droplets of the Dex-Alg solution in a row or at the four vertex positions of a quadrilateral to obtain the multi-compartment calcium alginate gel microcarriers as shown in Figure 7 b.

Claims

1. A preparation method of a multi-compartment calcium alginate gel microcarrier, characterized in that, The preparation method includes the following steps: dropping the Dex-Alg solution into a biphasic receiving solution with a PEG solution as the upper phase and a Dex-CaCl2 solution as the lower phase. After the Dex-Alg solution enters the lower phase, it is allowed to stand until the Dex-Alg solution droplets react with Ca 2+ in the lower phase and crosslink to obtain a multi-compartment calcium alginate gel microcarrier; The Dex-CaCl2 solution, biphasic receiving solution, and Dex-Alg solution are prepared by the following methods: (1) Mix equal masses of PEG solution and Dex solution with the same mass fraction, let it stand for phase separation to obtain a PEG-Dex solution system. The upper phase of the PEG-Dex solution system is the PEG solution, and the lower phase is the Dex solution. Extract the PEG solution and Dex solution separately; (2) Add Alg to the Dex solution obtained in step (1) to obtain a Dex-Alg solution; (3) Add CaCl2 to the Dex solution obtained in step (1) to obtain the lower phase of the biphasic receiving solution as the Dex-CaCl2 solution, and then add the PEG solution as the upper phase of the biphasic receiving solution to obtain the biphasic receiving solution.

2. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, In the PEG-Dex solution system, the mass percentage of PEG is 8-14 wt%, and the mass percentage of Dex is 8-14 wt%.

3. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, In the Dex-Alg solution, the mass percentage of Alg is 0.2-4.0 wt%.

4. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, In the Dex-CaCl2 solution, the mass percentage of CaCl2 is 5-12 wt%.

5. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, In the biphasic receiving solution, the height of the PEG solution is 1-4 mm.

6. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, Wait for the Dex-Alg solution to enter the lower phase and stand for 10-25 min.

7. The preparation method of the multi-compartment calcium alginate gel microcarrier according to claim 1, characterized in that, The Dex-Alg solution is dropped into the biphasic receiving solution, and the height of the liquid drop from the biphasic liquid surface during dropping is 2-8 mm.

8. A multi-compartment calcium alginate gel microcarrier prepared by using the preparation method of the multi-compartment calcium alginate gel microcarrier according to any one of claims 1-7.

9. The application of the multi-compartment calcium alginate gel microcarrier according to claim 8 in constructing a disease model.

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