Method for manufacturing nanomedicine by using multi-channel microfluidic mixing device and use thereof

Abstract

The application belongs to the field of biological medicine, and discloses a method for manufacturing nano drugs by using a multi-channel micro-fluidic mixing device and application. The method for manufacturing nano drugs by using the multi-channel micro-fluidic mixing device comprises the following steps: introducing a to-be-mixed solution A into a to-be-mixed solution B in a multi-channel mode to mix, and then obtaining nano drugs through secondary mixing. The application also discloses application of the multi-channel micro-fluidic mixing device in preparation of nano drugs. The method for manufacturing nano drugs by using the multi-channel micro-fluidic mixing device disclosed by the application is more beneficial to obtaining products with more excellent particle size, potential and encapsulation rate data under a large flow rate compared with a micro-fluidic preparation method in the prior art, and has a very good application prospect in scale-up production.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine and relates to a method for manufacturing nanomedicines using a multi-channel microfluidic mixing device and its application. Background Technology

[0002] Microfluidics refers to the science and technology involved in processing or manipulating tiny fluids using microchannels. It is an emerging interdisciplinary field involving chemistry, fluid physics, microelectronics, biology, and biomedical engineering. The core structure of existing microfluidic mixing devices typically employs two inlet channels to deliver two solutions in a specific ratio to the core structure of the microfluidic mixing unit for mixing. This core structure can be designed with different flow channel configurations, allowing the flowing liquids to mix through turbulent, laminar, or collisional processes. This enables chemical micro-reactions or the preparation of nanoparticles, such as liposomes, lipid nanoparticles, core-shell nanoparticles, PLGA nanoparticles, micelles, and nanoemulsions, with particle sizes at the nanometer and micrometer levels.

[0003] In existing technologies, the mixing of different fluids within the core structure of a microfluidic mixing component is generally achieved through free diffusion or turbulent mixing as the liquid flows through channels with varying cross-sections or multiple tortuous angles. The efficiency of both free diffusion and turbulent mixing is positively correlated with the contact surface area between the mixed solutions. Due to the small size and limited length of microfluidic mixing structures, two-phase or even multi-phase liquids need to be uniformly mixed within a short time to obtain nanoparticles and their composites with uniform particle size and high encapsulation efficiency. Therefore, in the design of microfluidic mixing structures, to increase the contact surface area between different fluids, the channel diameter is typically at the micrometer level. Good mixing is achieved through optimized design of the mixing channel structure. This method can be used for small-scale sample preparation, but due to its small channel diameter and limited throughput, it cannot meet the needs of large-scale sample preparation or industrial-scale sample production. When the mixing structure remains unchanged, simply increasing the channel diameter results in a low contact surface area between liquids, leading to low mixing efficiency and making complete mixing of two-phase or multiphase solutions impossible. Therefore, to improve mixing efficiency, the structure and length of the mixing channel must be adjusted. However, these changes often result in the adjusted mixing structure failing to reproduce the mixing effect of the original. This is why current microfluidic mixing equipment cannot be linearly scaled up from small-scale to pilot-scale or even industrial-scale equipment. Therefore, current microfluidic mixing structures and their optimization are only suitable for laboratory-scale micro-reaction synthesis or nanoparticle preparation, and still cannot meet the needs of large-scale production. Existing microfluidic equipment inevitably encounters problems such as mixing structure blockage, increased fluid pressure, and decreased mixing efficiency during large-scale production, leading to damage to the mixing structure, increased particle size and distribution of the prepared nanoparticles, decreased encapsulation efficiency or drug loading, and even nanoparticle sedimentation and aggregation, failing to meet the requirements of industrial-scale production. Summary of the Invention

[0004] The existing microfluidic mixing devices have the following technical shortcomings: they are only suitable for laboratory-scale micro-reaction synthesis or nano-formulation preparation, and cannot achieve linear scale-up, which is not conducive to industrial-scale production and application.

[0005] The first aspect of the present invention provides a method for manufacturing nanomedicines using a multi-channel microfluidic mixing device; comprising mixing a solution A to be mixed by introducing it into a solution B to be mixed in a multi-channel manner, and then obtaining nanomedicines by secondary mixing;

[0006] Furthermore, the nanomedicine is a lipid nanoparticle encapsulating active ingredients such as nucleic acid drugs;

[0007] Furthermore, the multi-channel configuration includes 2 to 100 channels;

[0008] Furthermore, the inner diameter of the multi-channel is 0.01–5 mm;

[0009] Furthermore, the secondary mixing involves passing the mixed liquid through a channel of a certain length;

[0010] Furthermore, the solutions to be mixed, A and B, are respectively selected from lipid solutions and aqueous solutions;

[0011] Furthermore, the lipid solution comprises ionizable lipid compounds and / or positively charged lipid compounds;

[0012] Furthermore, the lipid solution also includes one or more of the following ethanol solutions: CHOL and its analogues or derivatives, DSPC and its analogues or derivatives, DOPE and its analogues or derivatives, DMG-PEG2000 and its analogues or derivatives.

[0013] Furthermore, the aqueous solution is an aqueous solution containing water-soluble active pharmaceutical ingredients with a pH value of 3.0 to 7.0;

[0014] Furthermore, the method for manufacturing nanomedicines using a multi-channel microfluidic mixing device comprises the following steps:

[0015] Step 1: The solution A to be mixed is pumped into multiple parallel channels in the mixing channel at a certain flow rate using a power pump;

[0016] Step 2: The solution B to be mixed is introduced into the mixing channel through the injection channel at a certain flow rate using a power pump;

[0017] Step 3: Solution A to be mixed flows out through multiple channels and enters the liquid flow of solution B to be mixed in the same direction as the solution A to be mixed in multiple parallel flows. After a certain distance of mixing flow, preliminary microfluidic mixing is carried out.

[0018] Step 4: The liquid obtained from the initial microfluidic mixing in Step 3 is mixed a second time in a channel of a certain length to obtain the nanomedicine of the present invention.

[0019] Further, the flow rate ratio (mL / min) between the flow rate of solution A to be mixed in step 1 and the flow rate of solution B to be mixed in step 2 is 0.1:1 to 1:10;

[0020] Furthermore, the flow rate ratio (mL / min) between the flow rate of solution A to be mixed in step 1 and the flow rate of solution B to be mixed in step 2 is 0.5:1 to 1:10;

[0021] Furthermore, the flow rate ratio (mL / min) between the flow rate of solution A to be mixed in step 1 and the flow rate of solution B to be mixed in step 2 is 1:1 to 1:5;

[0022] Furthermore, the sum of the flow rates of the solution A to be mixed in step 1 and the solution B to be mixed in step 2 is 20 to 2000 mL / min.

[0023] Furthermore, in steps 1 and 2, the power pump, which provides power to the liquid in the pipeline, is preferably a stainless steel pump.

[0024] Furthermore, in step 2, the diameter of the mixing channel is 1–100 mm; the length is 20–1000 mm.

[0025] Furthermore, in step 3, the mixing flow distance is 10–800 mm;

[0026] Furthermore, in step 4, the channel of a certain length is either a straight channel or a curved channel;

[0027] Furthermore, in step 4, the length of the channel of a certain length is 1cm to 100m;

[0028] Furthermore, the curved channel is one or more of the following: a spiral curved channel, a figure-eight curved channel, an S-shaped curved channel, a Z-shaped curved channel, a channel with a Tesla hybrid structure, and a channel with an Ω hybrid structure;

[0029] Furthermore, the method for manufacturing nanomedicines using a multi-channel microfluidic mixing device includes a multi-channel mixing component, a secondary mixing channel, a stainless steel pump, a delivery pipeline, and connectors; the delivery pipeline connects the multi-channel mixing component, the secondary mixing channel, and the stainless steel pump via connectors; the stainless steel pump provides power to make the liquid flow in the multi-channel microfluidic mixing device to achieve the mixing operation;

[0030] Furthermore, the multi-channel hybrid component includes a capillary array assembly plate and a multi-channel base;

[0031] Furthermore, the capillary array assembly plate includes an assembly plate and capillaries connected to the assembly plate; wherein one end of the capillary is connected to the assembly plate, and the inner lumen of the capillary passes through the assembly plate;

[0032] Furthermore, the multi-channel base includes a hybrid channel that matches the capillary array assembly plate;

[0033] Furthermore, the multi-channel base includes a sample inlet channel communicating with the mixing channel;

[0034] Furthermore, the injection channel includes a left injection channel and a right injection channel;

[0035] Furthermore, the multi-channel base has a mixing channel that matches the capillary array assembly plate, wherein the matching is the matching of the capillary extending into the mixing channel;

[0036] Furthermore, the bending diameter of the secondary mixing channel is 0.5–20 cm;

[0037] Furthermore, the secondary mixing channel includes one or more combinations of spiral curved channels, figure-eight curved channels, S-shaped curved channels, Z-shaped curved channels, Tesla hybrid structure channels, and Ω hybrid structure channels;

[0038] Furthermore, the capillary array assembly plate has capillary arrays arranged in one or more of the following patterns: circular, square, elliptical, triangular, hexagonal, pentagonal, trapezoidal, star-shaped, or random.

[0039] Furthermore, the number of capillaries is 2 to 100;

[0040] Furthermore, the outer diameter of the capillary is 0.05 mm to 10 mm;

[0041] Furthermore, the inner diameter of the capillary is 0.01–5 mm;

[0042] Furthermore, the assembly plate is one of the following shapes: cylindrical, elliptical, square, trapezoidal, hexagonal, pentagonal, triangular, star-shaped, or irregular.

[0043] Furthermore, the assembly plate is cylindrical; the diameter of the circle is 1–100 mm;

[0044] Furthermore, the capillaries are preferably 7 to 50 in number; most preferably 7 to 31 in number.

[0045] Furthermore, the length of the capillary is 10–100 mm; preferably 15–21 mm.

[0046] Furthermore, the multi-channel mixing component is connected to the delivery pipe via a groove;

[0047] Furthermore, the multi-channel mixing component is connected to the delivery pipe via a connector through a groove on the multi-channel base;

[0048] Furthermore, the conveying pipeline includes one or more of stainless steel pipelines and polymer material pipelines; the conveying pipeline is a circular pipeline;

[0049] Furthermore, the connector includes one or more of the following: compression fitting, threaded fitting, and straight-insertion fitting;

[0050] In a specific embodiment of the present invention, there are 7 capillaries;

[0051] In another specific embodiment of the present invention, there are 13 capillaries;

[0052] In another specific embodiment of the present invention, there are 19 capillaries;

[0053] In another specific embodiment of the present invention, there are 31 capillaries;

[0054] In a specific embodiment of the present invention, the length of the capillary is 15 mm;

[0055] In another specific embodiment of the present invention, the length of the capillary is 21 mm;

[0056] In a specific embodiment of the present invention, the outer diameter of the capillary is 0.3 mm;

[0057] In another specific embodiment of the present invention, the outer diameter of the capillary is 0.5 mm;

[0058] In a specific embodiment of the present invention, the inner diameter of the capillary is 0.1 mm;

[0059] In another specific embodiment of the present invention, the inner diameter of the capillary is 0.2 mm;

[0060] In another specific embodiment of the present invention, the inner diameter of the capillary is 0.3 mm;

[0061] In a specific embodiment of the present invention, the collecting plate is cylindrical; the diameter of the circle is 8mm;

[0062] In a specific embodiment of the present invention, the stainless steel pump is a three-plunger stainless steel pump;

[0063] In a specific embodiment of the present invention, the upper side of the multi-channel base includes an upper sample inlet groove, the left and right sides of the multi-channel base respectively include a left sample inlet groove and a right sample inlet groove, and the lower side of the multi-channel base includes a mixing and discharging groove; the upper sample inlet groove and the mixing and discharging groove are connected by a mixing channel; the left sample inlet groove and the right sample inlet groove are connected to the mixing channel through the left sample inlet channel and the right sample inlet channel respectively.

[0064] Furthermore, the size of the upper sample inlet groove is not less than the size of the capillary array collection plate;

[0065] Furthermore, the mixing channel is square, polygonal, or cylindrical; preferably cylindrical.

[0066] Furthermore, the length of the mixing channel is 20–1000 mm;

[0067] Furthermore, the diameter of the mixing channel is 1–100 mm;

[0068] Furthermore, the left and right injection channels are cylindrical;

[0069] Furthermore, the diameters of the left and right injection channels are 1–20 mm;

[0070] The distance between the left and right injection channels and the upper injection groove is less than the length of the capillary tube;

[0071] In a specific embodiment of the present invention, the upper sample inlet groove is circular;

[0072] In a specific embodiment of the present invention, the left sample inlet groove, the right sample inlet groove, and the mixing and discharging groove are circular;

[0073] In a specific embodiment of the present invention, the water-soluble active pharmaceutical ingredient is a nucleic acid drug; preferably an mRNA drug.

[0074] Furthermore, the multi-channel microfluidic mixing device also includes a control system; the control system is a terminal control system.

[0075] Furthermore, the terminal is an intelligent interactive terminal;

[0076] Furthermore, the intelligent interactive terminal includes a computer terminal, a mobile phone terminal, and a PC terminal;

[0077] In a specific embodiment of the present invention, the multi-channel microfluidic mixing device includes a multi-channel microfluidic mixing system for mixing and preparing two liquids;

[0078] Furthermore, the two liquids include any two of the following: aqueous phase liquid, lipid phase liquid, and water-lipid mixture;

[0079] Furthermore, the two liquids include an aqueous phase liquid and a lipid phase liquid, a lipid phase liquid and a lipid phase liquid, and an aqueous phase liquid and an aqueous phase liquid;

[0080] Furthermore, the mixing equipment includes mixing equipment in the biological field and mixing equipment in the chemical field;

[0081] Furthermore, the mixing equipment in the biological field includes mixing equipment for the preparation of lipid nanoparticles, mixing equipment for the preparation of liposome nanomedicines, and mixing equipment for the preparation of lipid polymers;

[0082] Furthermore, the mixing equipment in the chemical field includes mixing equipment for the preparation of chemical nanoparticles, mixing equipment for the preparation of chemical molecules, and mixing equipment for chemical post-processing;

[0083] In a specific embodiment of this invention, the terminal refers to a device or program used to interact with a computer system or network. It acts as a bridge between the user and the computer, enabling the user to communicate and operate the computer through input and output.

[0084] In a specific embodiment of the present invention, the PC terminal is a device used to connect to a computer system and provide input and output functions.

[0085] In a specific embodiment of the present invention, the mixing device in the chemical field is a continuous flow microchannel reactor.

[0086] The second aspect of this invention provides the use of the method for manufacturing nanomedicines using the multi-channel microfluidic mixing device described in the first aspect of this invention in the preparation of nanomedicines;

[0087] The multi-channel microfluidic mixing device and method are as defined above;

[0088] Furthermore, the nanomedicine is a lipid nanoparticle, liposome, PLGA nanoparticle, micelle, nanoemulsion, microsphere, etc., encapsulating active ingredients such as nucleic acid drugs;

[0089] Furthermore, the nanomedicine is a lipid nanoparticle encapsulating active ingredients such as nucleic acid drugs;

[0090] Furthermore, the nucleic acid drug is one or more of the following: antisense oligonucleotide (ASO), small interfering RNA (siRNA), microRNA (miRNA), small activating RNA (saRNA), messenger RNA (mRNA), and RNA aptamer;

[0091] Furthermore, the nucleic acid drug also includes one or more of antisense oligonucleotides (ASO), small interfering RNA (siRNA), microRNA (miRNA), small activating RNA (saRNA), messenger RNA (mRNA), and RNA aptamers, used in combination with one or more other active pharmaceutical ingredients;

[0092] In a specific embodiment of the present invention, the nanomedicine is a lipid nanoparticle encapsulating mRNA.

[0093] The beneficial effects of this invention are as follows:

[0094] This invention employs a multi-channel microfluidic mixing device. Solution A is introduced into solution B via multiple channels for mixing, followed by secondary mixing to produce nanomedicines. This improves mixing efficiency. The secondary mixing channels of this invention ensure thorough mixing of the solutions. By optimizing the dimensions of the multi-channel system and the matching base, the microfluidic mixing flow rate is optimized. Using this method, stable microfluidic mixing can be achieved even at a total flow rate greater than 120 mL / min. The core mixing components can be linearly scaled up, resulting in lipid nanoparticles with excellent performance parameters in particle size, PDI, and encapsulation efficiency. Compared to existing microfluidic technologies, this invention demonstrates significantly superior performance in high-flow-rate sample preparation and linear scale-up. Attached Figure Description

[0095] Figure 1 A schematic front view of a capillary array assembly plate for a multi-channel hybrid component;

[0096] Figure 2 A top view of the capillary array assembly plate structure for a multi-channel hybrid component;

[0097] Figure 3 A bottom view of the capillary array assembly plate structure of the multi-channel hybrid component;

[0098] Figure 4 A partial sectional view of the main view of the capillary array assembly plate structure of the multi-channel hybrid component;

[0099] Figure 5 A front sectional view of the multi-channel base of the multi-channel hybrid component;

[0100] Figure 6 A left-side schematic diagram of the multi-channel base for a multi-channel hybrid component;

[0101] Figure 7 A top view of the multi-channel base of the multi-channel hybrid component;

[0102] Figure 8 A front sectional view of the capillary array assembly plate and the multi-channel base in conjunction.

[0103] Figure 9 A schematic diagram of liquid flow when a capillary array assembly plate is used in conjunction with a multi-channel base.

[0104] Figure 10 A schematic diagram of a multichannel microfluidic mixing device that includes a multichannel microfluidic mixing system;

[0105] Figure 11 Another specific implementation of the capillary array assembly plate for multi-channel hybrid components;

[0106] Figure 12 Another specific implementation of the capillary array assembly plate for multi-channel hybrid components;

[0107] Figure 13 Another specific implementation of the capillary array assembly plate for multi-channel hybrid components;

[0108] Figure 14 Another specific implementation of the capillary array assembly plate for multi-channel hybrid components;

[0109] Figure 15 Another specific implementation of the capillary array assembly plate for multi-channel hybrid components;

[0110] Figure 16 A schematic diagram of a commercially available Myanna chip;

[0111] In the diagram: 1. Collector plate; 2. Capillary tube; 3. Inner cavity; 4. Multi-channel base; 5. Mixing channel; 6. Upper injection groove; 7. Left injection groove; 8. Right injection groove; 9. Mixing and exiting groove; 10. Left injection channel; 11. Right injection channel; 12. Direction of the capillary array collector plate placed in the multi-channel base; 13. Schematic direction of liquid inlet for solution A to be mixed; 14. Schematic direction of liquid outlet for mixing; 15. Schematic direction of liquid inlet for solution B to be mixed; 16. Schematic direction of liquid inlet for solution B or C to be mixed; 17. Stainless steel pipe; 18. T-joint; 19. Spiral tube; 20. Pump for delivery of solution A to be mixed; 21. Pump for delivery of solution B to be mixed; 22. Compression fitting; 23. Threaded fitting; 24. Straight fitting; 25. Solution B to be mixed; 26. Solution A to be mixed; 27. Mixed solution. Detailed Implementation

[0112] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are for illustrative purposes only and are not intended to limit the scope of the invention. Furthermore, it should be noted that, for ease of description, only the parts relevant to the present invention are shown in the accompanying drawings.

[0113] It should be noted that, unless otherwise specified, the embodiments and features described in this invention can be combined with each other. The technical solution of this invention will now be described in detail with reference to the accompanying drawings and embodiments.

[0114] Unless otherwise stated, the exemplary embodiments / exemplifications shown are to be understood as providing exemplary features of various details that provide ways in which the technical concept of the invention can be implemented in practice. Therefore, unless otherwise stated, the features of the various embodiments / exemplifications may be additionally combined, separated, interchanged and / or rearranged without departing from the technical concept of the invention.

[0115] The use of crosshairs and / or shading in the accompanying drawings is generally used to clarify the boundaries between adjacent components. Thus, unless otherwise stated, the presence or absence of crosshairs or shading does not convey or indicate any preference or requirement for the specific material, material properties, dimensions, proportions, commonalities between the illustrated components, or any other characteristics, properties, etc., of the components. Furthermore, in the accompanying drawings, the dimensions and relative dimensions of components may be exaggerated for clarity and / or descriptive purposes. When exemplary embodiments can be implemented differently, a specific process sequence may be performed in a different order than that described. For example, two consecutively described processes may be performed substantially simultaneously or in the reverse order of their description. Furthermore, the same reference numerals denote the same components.

[0116] When a component is referred to as being "on" or "above" another component, "connected to" or "joined to" another component, the component may be directly on, directly connected to, or directly joined to the other component, or there may be intermediate components. However, when a component is referred to as being "directly on" another component, "directly connected to," or "directly joined to" another component, there are no intermediate components. Therefore, the term "connection" can refer to a physical connection, an electrical connection, etc., and may or may not have intermediate components.

[0117] For descriptive purposes, the present invention may use spatial relative terms such as “below,” “under,” “below,” “down,” “above,” “above,” “higher,” and “side (e.g., in a “sidewall”)” to describe the relationship between one component and another component as shown in the accompanying drawings. In addition to the orientations depicted in the drawings, the spatial relative terms are also intended to encompass different orientations of the device during use, operation, and / or manufacture. For example, if the device in the drawings is flipped, a component described as “below” or “under” another component or feature would subsequently be positioned “above” said other component or feature. Thus, the exemplary term “below” can encompass both “above” and “below” orientations. Furthermore, the device may be otherwise positioned (e.g., rotated 90 degrees or in other orientations), thus interpreting the spatial relative descriptive terms used herein accordingly.

[0118] The terminology used herein is for the purpose of describing particular embodiments and is not intended to be restrictive. As used herein, unless the context clearly indicates otherwise, the singular forms “a” and “the” are intended to include the plural forms as well. Furthermore, when the terms “comprising” and / or “including” and variations thereof are used in this specification, it indicates the presence of the stated features, integrals, steps, operations, parts, components, and / or groups thereof, but does not exclude the presence or addition of one or more other features, integrals, steps, operations, parts, components, and / or groups thereof. It should also be noted that, as used herein, the terms “substantially,” “about,” and other similar terms are used as approximate terms rather than as terms of degree, thus allowing them to account for inherent deviations in measurements, calculated values, and / or provided values ​​that would be recognized by one of ordinary skill in the art.

[0119] Unless otherwise specified, the detection methods used in this invention are based on common methods disclosed in the prior art.

[0120] Example 1: Multi-channel microfluidic mixing device

[0121] The multi-channel microfluidic mixing device of this embodiment includes a capillary array assembly plate and a multi-channel base, such as... Figures 1-4 As shown, the capillary array collection plate includes a cylindrical collection plate 1 and a capillary 2 vertically connected to one side of the collection plate 1; the inner cavity 3 of the capillary passes through the collection plate 1 to the other side of the collection plate.

[0122] In this embodiment, the capillary array collecting plate is a cylinder with a diameter of 8 mm and a thickness of 1.2 mm. There are 7 capillaries arranged in a circle. The capillary length is 15 mm, the outer diameter is 0.3 mm, and the capillary wall thickness is 0.1 mm. The inner cavity 3 of the capillary passes through the collecting plate 1 to the other side of the collecting plate.

[0123] like Figures 5-7 As shown, the multi-channel base of this embodiment has a total length of 53mm. The upper side includes an upper sample inlet groove with a diameter of 8.5mm. The left and right sides of the multi-channel base respectively include a left sample inlet groove and a right sample inlet groove with a diameter of 6.3mm. The lower side of the multi-channel base includes a mixing and exiting groove with a diameter of 8.5mm. The upper sample inlet groove and the mixing and exiting groove are connected by a mixing channel with a diameter of 2.1mm and a length of 43mm. The left sample inlet groove and the right sample inlet groove are connected to the mixing channel by left sample inlet channel and right sample inlet channel with a diameter of 2mm, respectively. In this embodiment, the upper sample inlet groove, the left sample inlet groove, the right sample inlet groove, and the mixing and exiting groove are circular.

[0124] like Figures 8-9As indicated by the directions shown, the capillary tubes of the capillary array assembly plate in this embodiment are placed downwards into the upper sample inlet groove of the matching multi-channel base. The capillary tubes enter the mixing channel and are located at the upper part of the mixing channel. The schematic direction 13 for the injection of solution A is that solution A is introduced from the upper sample inlet groove including the capillary array assembly plate. The schematic directions 15 and 16 for the injection of solution B or C are shown, where solution B is introduced from the left and right sample inlet pipes of the multi-channel base. Contact mixing occurs in the mixing channel 5 of the multi-channel base. While solution A diffuses laterally, it moves downwards together with solution B, flowing through the entire mixing channel. During this flow, nanomedicine is formed, and the mixture containing the nanomedicine flows out from the outlet of the multi-channel base. Figure 9 The mixed sample output direction shown is 14;

[0125] like Figure 10 In this embodiment of the multi-channel microfluidic mixing device, the solution to be mixed, A 26, is powered by pump 20 and transported to the upper injection groove via stainless steel pipe 17. The solution to be mixed, A, passes through the upper end of the capillary array assembly plate in the upper injection groove and then enters the mixing channel 5 through the capillary cavity; the solution to be mixed, B... Powered by pump 21, the sample enters the mixing channel from the left and right sides of the multi-channel base via stainless steel pipe and tee 18, respectively, through the left and right inlet channels. Solution A begins to contact solution B at the capillary outlet. As solution A diffuses laterally into solution B, it moves downwards along with solution B, flowing through the entire mixing channel. During this flow, the lipid components in solution A encapsulate the active ingredients in solution B, forming nanomedicines. The nanomedicines enter a 2cm diameter spiral pipe 19 through a stainless steel pipe connected to the mixing outlet groove for secondary mixing, yielding the nanomedicines described in this invention. In this mixing device, the upper inlet groove, the mixing outlet groove, and the stainless steel pipe are connected via compression fittings 22; the left and right inlet grooves are connected to the stainless steel pipe via threaded fittings 23; and the tee is connected to the stainless steel pipe via a straight-insertion fitting 24.

[0126] When the prepared nanomedicine is a lipid nanoparticle encapsulating mRNA, solution A to be mixed is a lipid solution, and solution B to be mixed is an aqueous solution. The specific manufacturing steps include:

[0127] Prepare lipid and aqueous solutions. The lipid solution is an ethanol solution containing SM102, CHOL, DSPC, and DMG-PEG2000, with a concentration of 18 mg / mL for SM102, 7.55 mg / mL for CHOL, 4.01 mg / mL for DSPC, and 1.91 mg / mL for DMG-PEG2000. The aqueous solution is a 500 mmol / L PBS buffer containing 0.5 mg / mL mRNA.

[0128] Step 1: The lipid solution is powered by a three-plunger stainless steel pump and transported through a stainless steel pipeline to the upper injection groove of the multi-channel base. It then enters the mixing channel through the inner cavities of multiple capillaries in the capillary array assembly plate.

[0129] Step 2: The aqueous solution is powered by a three-plunger stainless steel pump and transported through stainless steel pipes to the left and right injection grooves of the multi-channel base, and then enters the mixing channel through the left and right injection channels.

[0130] Step 3: The lipid solution flows out through the inner lumen of multiple capillaries and begins to contact the aqueous solution at the capillary outlet. Multiple parallel lipid liquid flows enter the aqueous liquid flow in the same direction, diffusing laterally and moving downward together with the aqueous solution, flowing through the entire mixing channel for preliminary microfluidic mixing.

[0131] Step 4: The mixed liquid obtained from the initial microfluidic mixing in Step 3 is mixed a second time in a spiral bend tube to obtain the lipid nanoparticles carrying mRNA of the present invention.

[0132] Throughout the process, the total flow rates of the lipid solution and the aqueous solution were 50 mL / min, 80 mL / min, 100 mL / min, 120 mL / min, 240 mL / min, 300 mL / min, 360 mL / min, and 600 mL / min (the flow rate ratio of the aqueous solution to the lipid solution was 3:1).

[0133] The particle size, PDI, and encapsulation efficiency of lipid nanoparticles obtained at different flow rates were measured.

[0134] Example 2

[0135] A multi-channel microfluidic mixing device was obtained by using different numbers of capillaries, as well as capillaries of different lengths and diameters; the specific capillary parameters are as follows:

[0136] Figure 11 The capillary array assembly plate has 13 capillaries arranged in a circular pattern; the length of the capillary 2 is 15mm, the outer diameter is 0.3mm, and the wall thickness of the capillary is 0.1mm.

[0137] Figure 12 The capillary array assembly plate has 19 capillaries arranged in a circular pattern; the capillary 2 has a length of 15mm, an outer diameter of 0.3mm, and a wall thickness of 0.1mm.

[0138] Figure 13 The capillary array assembly plate has 31 capillaries arranged in a circular pattern; the capillary 2 has a length of 15mm, an outer diameter of 0.3mm, and a wall thickness of 0.1mm.

[0139] Figure 14 The capillary array assembly plate has 13 capillaries arranged in a circle; the length of capillary 2 is 15mm, the outer diameter is 0.5mm, and the wall thickness of capillary 2 is 0.1mm.

[0140] Figure 15 The capillary array assembly plate has 13 capillaries arranged in a circle; the length of capillary 2 is 21mm, the outer diameter is 0.3mm, and the wall thickness of capillary 2 is 0.1mm.

[0141] Test Example 3

[0142] The multi-channel microfluidic mixing device of Example 2 was used to manufacture lipid nanoparticles loaded with mRNA using the method for manufacturing nanomedicines using the multi-channel microfluidic mixing device of the present invention of Example 1. The particle size, PDI and encapsulation efficiency of the manufactured lipid nanoparticles loaded with mRNA were detected and compared with those prepared by commercially available microfluidic devices in the prior art.

[0143] Example 1 is numbered 1; Example 2 is numbered sequentially as follows: Figure 11 (Serial No. 2) Figure 12 (Serial No. 3) Figure 13 (Serial No. 4) Figure 14 (Serial No. 5) Figure 15 (Serial Number 6)

[0144] Compared to commercially available Myanna chips (serial number 7), the hybrid core structure of commercially available Myanna chips is as follows: Figure 16 As shown, the structure is a typical multi-angle tortuous path mixing channel structure in the prior art; the lipid phase solution and the aqueous phase solution enter from the left and right inlets respectively, mix and contact in the multi-angle tortuous path mixing channel, and the lipid nanoparticle liquid is collected from the mixing channel outlet, with other conditions being the same as above.

[0145] The collected lipid nanoparticles were analyzed for particle size, PDI, and encapsulation efficiency. The results are shown in Tables 1 to 3 below.

[0146] Table 1 Particle size (nm) detection data

[0147] Serial Number 1 65.48 60.51 60.23 58.8 58.49 56.13 55.71 57.56 Serial Number 2 76.12 76.33 73.76 72.68 72.55 68.01 67.92 67.67 Serial Number 3 68.72 69.01 69.95 68.86 68.70 67.32 67.33 66.98 Serial Number 4 71.77 69.56 69.35 69.76 68.33 67.92 63.24 65.33 Serial Number 5 85.23 85.89 85.18 83.97 81.56 81.55 80.36 80.63 Serial Number 6 87.57 87.11 87.03 87.15 85.34 85.13 84.33 85.34 Serial Number 7 73.22 72.89 70.88 88.34 - - - -

[0148] Table 1 shows the particle size measurement data of lipid nanoparticles prepared by mixing lipid solution and aqueous solution using the multi-channel microfluidic mixing device of the present invention to manufacture nanomedicines. As can be seen from Table 1, at a flow rate of 50–600 mL / min, the lipid nanoparticles prepared by mixing using the multi-channel microfluidic mixing device of the present invention exhibit small particle size fluctuations. For items 1–4, when there are 7–13 capillaries, the particle size of the obtained lipid nanoparticles is between 55–70 nm; for items 5 and 6, when there are 19–31 capillaries, the particle size of the obtained lipid nanoparticles is between 80–90 nm.

[0149] In item 7, lipid nanoparticles were prepared by mixing with commercially available Myanna chips. At a flow rate of 50–100 mL / min, the particle size of the prepared lipid nanoparticles was between 70 and 75 nm. At a flow rate of 120 mL / min, the particle size of the prepared lipid nanoparticles increased to 88.34 nm. At a flow rate greater than 120 mL / min, the mixing preparation of lipid nanoparticles failed due to damage to the chip structure caused by high liquid pressure.

[0150] The data above shows that, compared with the existing commercially available Myanna chips, the method of manufacturing nanomedicines using the multi-channel microfluidic mixing device of the present invention can prepare lipid nanoparticles by mixing lipid solutions and aqueous solutions at a flow rate greater than 120 mL / min. The prepared lipid nanoparticles have small particle size fluctuations, which is conducive to the industrial-scale production application.

[0151] Table 2 PDI test data

[0152]

[0153]

[0154] Table 2 shows the PDI measurement data of lipid nanoparticles prepared by mixing lipid solution and aqueous solution using the multi-channel microfluidic mixing device of the present invention to manufacture nanomedicines. As can be seen from Table 2, at a flow rate of 50–600 mL / min, the PDI values ​​of lipid nanoparticles prepared by mixing using the multi-channel microfluidic mixing device of the present invention fluctuate within a reasonable range. When the number of capillaries is 7–13 (numbers 1–3), the PDI values ​​of the obtained lipid nanoparticles are between 0.1 and 0.15; when the number of capillaries is 19–31 (numbers 5–6), the PDI values ​​of the obtained lipid nanoparticles are between 0.12 and 0.17.

[0155] When the number of capillaries in sequence 4 is 13 and the inner diameter of the capillaries is 0.3 mm, the PDI value of the lipid nanoparticles obtained is significantly increased compared with that in sequence 3 (13 capillaries and an inner diameter of 0.1 mm).

[0156] In item 7, lipid nanoparticles were prepared by mixing using commercially available Myanna chips. At a flow rate of 50–100 mL / min, the PDI value of the prepared lipid nanoparticles was between 0.15 and 0.16. At a flow rate of 120 mL / min, the PDI value of the prepared lipid nanoparticles increased to 0.189. At a flow rate greater than 120 mL / min, the mixing preparation of lipid nanoparticles failed due to damage to the chip structure caused by high liquid pressure.

[0157] The data above demonstrate that, compared to commercially available Myanna chips, the multi-channel microfluidic mixing device of this invention for manufacturing nanomedicines can prepare lipid nanoparticles by mixing lipid solutions and aqueous solutions at flow rates greater than 120 mL / min. The prepared lipid nanoparticles exhibit low PDI fluctuations and excellent polymer dispersibility. In particular, the capillary array assembly plate with a capillary inner diameter of 0.1 mm achieves PDI values ​​of 0.1–0.13 for lipid nanoparticles at flow rates greater than 240 mL / min, exhibiting significantly superior polymer dispersibility. This is beneficial for large-scale industrial production applications.

[0158] Table 3 Encapsulation Efficiency (%) Test Data

[0159]

[0160]

[0161] Table 3 shows the encapsulation efficiency measurement data of lipid nanoparticles prepared by mixing lipid solution and aqueous solution using the multi-channel microfluidic mixing device of the present invention to manufacture nanomedicines. As can be seen from Table 3, at a flow rate of 50–600 mL / min, the encapsulation efficiency of lipid nanoparticles prepared by mixing using the multi-channel microfluidic mixing device of the present invention is greater than 89%; especially for items 1–4, with 7–13 capillaries, the encapsulation efficiency of lipid nanoparticles obtained at a high flow rate of 300–600 mL / min is greater than 95%; for items 5–6, with 19–31 capillaries, the encapsulation efficiency of lipid nanoparticles obtained is between 89% and 92%.

[0162] In item 7, lipid nanoparticles were prepared by mixing with commercially available Myanna chips. At a flow rate of 50–100 mL / min, the encapsulation efficiency of the prepared lipid nanoparticles was between 81% and 92%. At a flow rate of 120 mL / min, the PDI encapsulation efficiency of the prepared lipid nanoparticles decreased to 81.65%. At a flow rate greater than 120 mL / min, the mixing preparation of lipid nanoparticles failed due to damage to the chip structure caused by high liquid pressure.

[0163] The above data shows that, compared to existing commercially available Myanna chips, the multi-channel microfluidic mixing device of this invention can prepare lipid nanoparticles by mixing lipid solutions and aqueous solutions at flow rates greater than 120 mL / min. The prepared lipid nanoparticles have excellent encapsulation efficiency, which is beneficial for the encapsulation and delivery of mRNA active components. In particular, at flow rates greater than 240 mL / min, the encapsulation efficiency of the lipid nanoparticles is greater than 95%, showing a significantly superior encapsulation effect. This is conducive to the industrial-scale production application.

[0164] In other specific embodiments of the present invention, when the total liquid flow rate is greater than 600 mL / min during microfluidic mixing, similar beneficial effects as described above are still obtained.

[0165] In other specific embodiments of the present invention, the method for manufacturing nanomedicines using the multi-channel microfluidic mixing device of the present invention is used for the mixed preparation of nucleic acid drugs encapsulated by lipid polymers, the mixed preparation of siRNA, ASO, miRNA, saRNA, or Aptamer encapsulated by LNPs, and when the total liquid flow rate is greater than 100 mL / min during microfluidic mixing, it has similar beneficial effects as described above.

[0166] In other specific embodiments of the present invention, the method of the present invention is used in a continuous flow microchannel reactor to carry out chemical micro-reactions, and has similar beneficial effects as described above.

[0167] In summary, the method for manufacturing nanomedicines using the multi-channel microfluidic mixing device of the present invention, compared with the microfluidic mixing chips or devices disclosed in the prior art, uses a capillary array assembly plate and a matching channel base to prepare lipid nanoparticles at a total flow rate greater than 120 mL / min. The prepared lipid nanoparticles are of stable and reliable quality and can be applied to large-scale production in industrial settings.

Claims

1. A method for manufacturing nanomedicines using a multi-channel microfluidic mixing device, characterized in that: The method includes introducing a solution A to be mixed into a solution B using a multi-channel approach for mixing, followed by secondary mixing to obtain nanomedicine; the multi-channel microfluidic mixing device has a multi-channel mixing component composed of a capillary array assembly plate and a multi-channel base.

2. The method according to claim 1, characterized in that: The multi-channel has 2 to 100 channels; the inner diameter of each channel is 0.01 to 5 mm.

3. The method according to claim 2, characterized in that: The solutions to be mixed, A and B, are respectively selected from lipid solutions and aqueous solutions; wherein, the lipid solution includes ionizable lipid compounds and / or positively charged lipid compounds; and the aqueous solution is an aqueous solution with a pH of 3.0 to 7.0 containing water-soluble active pharmaceutical ingredients.

4. The method according to claim 1; characterized in that: The specific steps include: Step 1: The solution A to be mixed is pumped into multiple parallel channels in the mixing channel at a certain flow rate using a power pump; Step 2: The solution B to be mixed is introduced into the mixing channel through the injection channel at a certain flow rate using a power pump; Step 3: Solution A to be mixed flows out through multiple channels and enters the liquid flow of solution B to be mixed in the same direction as the solution A to be mixed in multiple parallel flows. After a certain distance of mixing flow, preliminary microfluidic mixing is carried out. Step 4: The liquid obtained from the initial microfluidic mixing in Step 3 is mixed a second time in a channel of a certain length to obtain nanomedicine.

5. The method according to claim 4, characterized in that: The flow rate ratio of the solution A to be mixed in step 1 to the solution B to be mixed in step 2 is 0.1:1 to 1:10, and the unit of flow rate is mL / min.

6. The method according to claim 4, characterized in that: The sum of the flow rates of solution A to be mixed in step 1 and solution B to be mixed in step 2 is 20~2000 mL / min.

7. The method according to claim 4, characterized in that: In step 2, the diameter of the mixing channel is 1~100mm; the length is 20~1000mm. In step 3, the mixing flow distance is 10~800mm.

8. The method according to claim 1, characterized in that: The multi-channel microfluidic mixing device includes a multi-channel mixing component, a secondary mixing channel, a stainless steel pump, a delivery pipeline, and connectors; the delivery pipeline connects the multi-channel mixing component, the secondary mixing channel, and the stainless steel pump through the connectors; the stainless steel pump provides power to make the liquid flow in the multi-channel microfluidic mixing device to achieve the mixing operation.

9. The method according to claim 1, characterized in that: The nanomedicines are lipid nanoparticles, liposomes, PLGA nanoparticles, micelles, nanoemulsions, and microspheres that encapsulate nucleic acid drugs.

10. The method according to claim 9, characterized in that: The nanomedicine is a lipid nanoparticle encapsulating a nucleic acid drug.

11. The use of the method described in claims 1-9 in the preparation of nanomedicines; characterized in that: The nanomedicine is a drug with nucleic acid active ingredients encapsulated.

12. The use according to claim 10, characterized in that: The nucleic acid drug is one or more of ASO, siRNA, miRNA, saRNA, mRNA, and Aptamer; or one or more of ASO, siRNA, miRNA, saRNA, mRNA, and Aptamer are used in combination with one or more other drug components.

Images