3D micro-fluidic chip and 3D mixing system for producing nano-biopharmaceutical, method for preparing product by using chip and system, and use of product

Abstract

Disclosed are a 3D micro-fluidic chip for producing a nano-biopharmaceutical, a method for preparing a product by using the chip, and a use of the product. Mixing channels of a 3D mixing unit include a first channel and a second channel, the first channel and the second channel among the mixing channels are located in the same plane, and any two adjacent mixing channels are located in different planes. In the 3D mixing unit, any two adjacent mixing channels are located in different planes, that is, the plurality of mixing channels form a 3D layout, so that when fluid in a mixing channel enters the next mixing channel, the fluid enters another space plane, and thus the probability that a turbulence component and a fluid component collide with each other is increased, the fluid mixing efficiency is improved, and the mixing effect is improved, thereby solving the problems in various applications such as low mixing efficiency and poor mixing effect of a lipid nanoparticle aqueous phase and an organic phase, thus accurately controlling the nanoparticle production, improving the drug embedding efficiency, and improving the drug delivery and treatment effects.

Description

3D microfluidic chip for producing nanobiomedicine, 3D mixing system and method and application of preparation product thereof TECHNICAL FIELD

[0001] The present application relates to the technical field of biopharmaceuticals, in particular to a 3D microfluidic chip for producing nanobiomedicine and a method and application of preparation product thereof. BACKGROUND

[0002] Nanobiomedicine combines nanotechnology and biopharmaceutical technology to develop effective drugs. Nanoparticles are widely used for the delivery of biomolecules to prepare vaccines and biopharmaceuticals for the prevention and treatment of diseases. Biological macromolecules such as proteins and nucleic acid drugs have more clear disease mechanisms and therapeutic effects, and have developed faster in recent years. However, protein and nucleic acid drugs, especially nucleic acid drugs, have poor stability and require effective delivery technology to protect the drugs and increase the therapeutic effect. Lipid nanoparticles (LNP) have been developed in recent years for the delivery of nucleic acid drugs including mRNA, siRNA, ASO, etc. mRNA-LNP technology has been widely used for the development of vaccines, protein replacement drugs, gene editing and cell therapy.

[0003] Lipid nanoparticles need to mix aqueous phase and organic phase to form stable high-quality nanoparticles, so there are certain challenges in production. In recent years, various microfluidic mixers have been widely used in the production of lipid nanoparticles, but different designs have a big gap in mixing efficiency and nanoparticle quality, production scale-up and cost.

[0004] SUMMARY

[0005] The present application provides a 3D microfluidic chip for producing nanobiomedicine and a method and application of preparation product thereof, which solves the problems of low mixing efficiency and poor mixing effect of aqueous phase and organic phase of lipid nanoparticles.

[0006] In one embodiment, a 3D mixing unit for producing nanobiomedicine is provided, comprising a plurality of mixing channels connected in sequence:

[0007] The mixing channel comprises a first channel and a second channel, the inlet of the first channel is connected with the inlet of the second channel to form a common inlet, and the outlet of the first channel and the outlet of the second channel are connected to form a common outlet;

[0008] The common inlet of the mixing channel located at the front end is used at least for injecting a first raw material and a second raw material, the common outlet of the mixing channel located at the rear end is used for discharging a mixture formed by mixing the first raw material and the second raw material, the common inlet of the mixing channel located at the middle part is communicated with the common outlet of the adjacent mixing channel, and the common outlet of the mixing channel located at the middle part is communicated with the common inlet of the adjacent mixing channel;

[0009] The first channel and the second channel of the mixing channel are located in the same plane, and any two adjacent mixing channels are located in different planes.

[0010] In one embodiment, the angle between the planes in which the two adjacent mixing channels are located is 15°-165°.

[0011] In one embodiment, the angle between the planes in which the two adjacent mixing channels are located is 45°-135°.

[0012] In one embodiment, the angle between the planes in which the two adjacent mixing channels are located is 90°.

[0013] In one embodiment, a plurality of the mixing channels are spirally arranged along a straight line.

[0014] In one embodiment, the first channel and / or the second channel is / are bent, the angle between the inlet of the first channel and the inlet of the second channel is 15°-165°, and the angle between the outlet of the first channel and the outlet of the second channel is 15°-165°.

[0015] In one embodiment, the angle between the inlet of the first channel and the inlet of the second channel is 45°-135°, and the angle between the outlet of the first channel and the outlet of the second channel is 45°-135°.

[0016] In one embodiment, the angle between the inlet of the first channel and the inlet of the second channel is 90°, and the angle between the outlet of the first channel and the outlet of the second channel is 90°.

[0017] In one embodiment, the inner diameter of the first channel and the second channel is 5.0mm-0.05mm;

[0018] Preferably, the inner diameter of the first channel and the second channel is 2.0mm-0.1mm;

[0019] More preferably, the inner diameter of the first channel and the second channel is 1.5mm-0.2mm.

[0020] In one embodiment, the inner diameter of the first channel and the second channel and the particle size of the mixture satisfy the following regression formula requirements: y = -17.612x1 2 +72.706x1+47.987

[0021] Wherein, x1 is the inner diameter of the first channel and the second channel, and y is the particle size of the mixture.

[0022] In one embodiment, a 3D microfluidic chip for producing nanobiomedicine is provided, comprising: a first inlet channel, a second inlet channel, an outlet channel and the 3D mixing unit as described above, wherein the first inlet channel and the second inlet channel are in communication with a common inlet of the mixing channel at the front end, and the outlet channel is in communication with a common outlet of the mixing channel at the rear end.

[0023] In one embodiment, a third inlet channel is further included, wherein the outlet of the first inlet channel and the outlet of the second inlet channel are in communication with the inlet of the third inlet channel, and the outlet of the third inlet channel is in communication with the common inlet of the mixing channel at the front end.

[0024] In one embodiment, the first inlet channel and / or the second inlet channel and the third inlet channel are distributed on different axes.

[0025] In one embodiment, the included angle between the first inlet channel and / or the second inlet channel and the third inlet channel is 15°-165°.

[0026] In one embodiment, the included angle between the first inlet channel and / or the second inlet channel and the third inlet channel is 90°.

[0027] In one embodiment, a split channel and a convergence channel are further included, wherein a plurality of the 3D mixing units are arranged between the split channel and the convergence channel; the split channel comprises one or two inlets and a plurality of outlets, the first inlet channel and the second inlet channel are in communication with the inlets of the split channel, and the plurality of outlets of the split channel are in one-to-one correspondence with the plurality of 3D mixing units; the convergence channel comprises a plurality of inlets and an outlet, the plurality of inlets of the convergence channel are in one-to-one correspondence with the plurality of 3D mixing units, and the outlet of the convergence channel is in communication with the outlet channel.

[0028] In one embodiment, the plurality of 3D mixing units are arranged in parallel and at intervals.

[0029] In one embodiment, the 3D microfluidic chip further comprises a substrate, and the first inlet channel, the second inlet channel, the outlet channel and the 3D mixing unit are located in the substrate.

[0030] In one embodiment, the substrate is a flat plate structure, and the inlet of the first inlet channel, the inlet of the second inlet channel and the outlet of the outlet channel are located on the same side of the substrate.

[0031] In one embodiment, the first fluid joint, the second fluid joint and the third fluid joint are installed on the same side of the substrate, the first fluid joint is in communication with the inlet of the first inlet channel, the second fluid joint is in communication with the inlet of the second inlet channel, and the third fluid joint is in communication with the outlet of the outlet channel.

[0032] In one embodiment, the first fluid joint, the second fluid joint and the third fluid joint are female luer type fluid joints.

[0033] In one embodiment, a 3D mixing system for producing nanobiomedicine is provided, characterized in that it comprises:

[0034] The 3D microfluidic chip described above;

[0035] A first raw material supply device for storing a first raw material, the first raw material supply device being in communication with the first inlet channel, and the first raw material supply device being configured to inject the first raw material into the first inlet channel;

[0036] A second raw material supply device for storing a second raw material, the second raw material supply device being in communication with the second inlet channel, and the second raw material supply device being configured to inject the second raw material into the second inlet channel; and

[0037] A collection device in communication with the outlet channel, the collection device being configured to collect a mixture produced by mixing the first raw material and the second raw material.

[0038] In one embodiment, the system further comprises a driving device connected to the first raw material supply device and the second raw material supply device, the driving device being configured to drive the first raw material to be injected into the first inlet channel and the second raw material to be injected into the second inlet channel.

[0039] In one embodiment, the system further comprises a cleaning device in communication with the first inlet channel and the second inlet channel, the cleaning device being configured to inject a cleaning liquid; and a waste liquid device in communication with the outlet channel, the waste liquid device being configured to collect waste cleaning liquid.

[0040] In one embodiment, a method for mixing and preparing a product using the 3D microfluidic chip described above or the 3D mixing system described above is provided, characterized in that it comprises the following steps:

[0041] Injecting the first raw material and the second raw material into the 3D microfluidic chip, respectively;

[0042] Mixing the first raw material and the second raw material in the 3D mixing unit to form a particle size of the mixture.

[0043] In one embodiment, the flow rate of the first raw material and the second raw material in the 3D mixing unit and the particle size of the mixture satisfy the following regression equation: y = 0.0718x2 2 -5.4452x2+185.47

[0044] wherein x2 is the flow rate of the first raw material and the second raw material in the 3D mixing unit, and y is the particle size of the mixture.

[0045] In one embodiment, a use of the above-mentioned 3D microfluidic chip or the above-mentioned 3D mixing system in the preparation of a product is provided.

[0046] Preferably, the product is a particle carrying a drug molecule or a cell or a pharmaceutical composition.

[0047] More preferably, the particle is a lipid nanoparticle for delivering a drug.

[0048] The 3D microfluidic chip for producing nanobiomedicine according to the above-mentioned embodiment and the method and use thereof in the preparation of a product can increase the probability of turbulent flow and collision of fluid components when the fluid in the mixing channel enters the next mixing channel, improve the fluid mixing efficiency and the mixing effect, and thus solve the problems of low mixing efficiency and poor mixing effect of the water phase and the organic phase of the lipid nanoparticle. BRIEF DESCRIPTION OF DRAWINGS

[0049] FIG. 1 is a structural schematic diagram of a 3D mixing unit in one embodiment;

[0050] FIG. 2 is a structural schematic diagram of a 3D microfluidic chip in one embodiment;

[0051] FIG. 3 is a structural schematic diagram of a 3D microfluidic chip in one embodiment;

[0052] FIG. 4 is a diagram of the relationship between the chip structure and LNP particles and PDI in one embodiment;

[0053] FIG. 5 is a diagram of the relationship between the inner diameter of the chip and LNP particles and PDI in one embodiment;

[0054] FIG. 6 is a diagram of the relationship between the inner diameter of the chip and LNP particles in one embodiment;

[0055] FIG. 7 is a structural schematic diagram of a 3D microfluidic chip in one embodiment;

[0056] FIG. 8 is a structural block diagram of a 3D mixing system in one embodiment;

[0057] Fig. 9 is a structural block diagram of a 3D mixing system in an embodiment;

[0058] Fig. 10 is a structural block diagram of a 3D mixing system in an embodiment;

[0059] Fig. 11 is a graph of the relationship between preparation flow rate and LNP particle, PDI in an embodiment;

[0060] Fig. 12 is a graph of the relationship between preparation flow rate and LNP particle in an embodiment;

[0061] Wherein the reference signs are as follows:

[0062] 10-3D mixing unit, 11-mixing channel, 111-first channel, 112-second channel, 20-first inlet channel, 30-second inlet channel, 40-outlet channel, 50-third inlet channel, 61-first fluid joint, 62-second fluid joint, 63-third fluid joint, 70-splitting channel, 80-converging channel;

[0063] 100-3D microfluidic chip, 101-substrate, 200-first raw material supply device, 300-second raw material supply device, 400-collecting device, 500-driving device, 600-cleaning device, 700-waste liquid device;

[0064] P1-plane where the previous mixing channel is located, P2-plane where the next mixing channel is located. DETAILED DESCRIPTION

[0065] The application will be further described in detail below with specific embodiments and accompanying drawings. In different embodiments, similar elements are associated with similar element reference signs. In the following embodiments, many details are described in order to make the application better understood. However, those skilled in the art can easily recognize that some features can be omitted in different cases, or can be replaced by other elements, materials, methods. In some cases, some operations related to the application are not shown or described in the specification in order to avoid the core part of the application being overwhelmed by too much description, and it is not necessary to describe these related operations in detail for those skilled in the art according to the description in the specification and general technical knowledge in the art.

[0066] In addition, features described in the specification, operations or characteristics can be combined in any appropriate manner in various embodiments. Meanwhile, the steps or actions in the method description can also be sequentially changed or adjusted in a manner apparent to those skilled in the art. Therefore, the various sequences in the specification and the drawings are only for the purpose of clearly describing a certain embodiment, and do not mean that the sequence is necessary, unless otherwise stated that a certain sequence must be followed.

[0067] The serial numbers of components in this paper, such as "first", "second", etc., are only used to distinguish the described objects, and do not have any order or technical meaning. The "connection" and "coupling" in this application include direct and indirect connection (coupling) unless otherwise specified.

[0068] Embodiment one:

[0069] The embodiment provides a 3D mixing unit for producing nanobiological medicine, which is used for mixing a plurality of raw materials to obtain a mixture. The first raw material and the second raw material can be mixed to obtain the mixture. A larger number of different raw materials can be injected into the 3D mixing unit to obtain a mixed product, hereinafter referred to as a mixture, for example, mixing an aqueous phase and an organic phase to obtain a lipid nanoparticle (LNP). The 3D mixing unit can also mix other substances to achieve the preparation of vaccines or other biological drugs.

[0070] Referring to FIG. 1, the 3D mixing unit 10 of the embodiment mainly comprises a plurality of mixing channels 11 connected in sequence. The plurality of mixing channels 11 are connected in sequence in a single chain structure.

[0071] The mixing channel 11 comprises a first channel 111 and a second channel 112. The first channel 111 and the second channel 112 are combined into a structure in which the head and the tail are connected and the middle is separated. The first channel 111 and the second channel 112 are combined into a mouth-shaped structure, a circular structure or an elliptical structure.

[0072] The first channel 111 and the second channel 112 have an inlet and an outlet at the head and the tail, respectively. The inlet of the first channel 111 is connected with the inlet of the second channel 112 to form a common inlet. The outlet of the first channel 111 is connected with the outlet of the second channel 112 to form a common outlet.

[0073] The plurality of mixing channels 11 are connected in sequence end to end. The common inlet of the mixing channel 11 at the front end is used to inject the first raw material and the second raw material, and the common outlet of the mixing channel at the rear end is used to discharge the mixture formed by mixing the first raw material and the second raw material. The common inlet of the mixing channel 11 between the front end and the rear end is in communication with the common outlet of the previous mixing channel 11, and the common outlet of the mixing channel 11 between the front end and the rear end is in communication with the common inlet of the next mixing channel 11. The first raw material and the second raw material will be mixed in the plurality of mixing channels 11 in sequence.

[0074] In the embodiment, the first channel 111 and the second channel 112 of each mixing channel 11 are located in the same plane, and the central axis of the first channel 111 and the central axis of the second channel 112 are located in the same plane. Adjacent two mixing channels 11 are located in different planes, for example, the previous mixing channel 11 is located in plane P1, and the next mixing channel 11 is located in plane P2. The plane P1 and the plane P2 have an included angle, and the two planes are not coplanar and not parallel.

[0075] The included angle between the plane P1 and the plane P2 is 15°-165°, for example, the included angle between the plane P1 and the plane P2 is 15°, 25°, 30°, 45°, 60°, 70°, 80°, 90°, 100°, 110°, 120°, 135°, 145°, 155° and 165°, etc., that is, the plane P1 and the plane P2 are at least ±15°, so that the first raw material and the second raw material mixed by the previous mixing channel 11 will at least collide after a 15° turn when entering the next mixing channel 11, thereby improving the efficiency and uniformity of mixing the first raw material and the second raw material.

[0076] In a preferred embodiment, the included angle between the plane P1 and the plane P2 is 45°-135°, for example, the included angle between the plane P1 and the plane P2 is 45°, 60°, 70°, 80°, 90°, 100°, 110°, 120° and 135°, etc., that is, the plane P1 and the plane P2 are at least ±45°, so that the first raw material and the second raw material mixed by the previous mixing channel 11 will at least collide after a 45° turn when entering the next mixing channel 11, thereby significantly improving the efficiency and uniformity of mixing the first raw material and the second raw material.

[0077] In a more preferred embodiment, the included angle between the plane P1 and the plane P2 is 90°, so that the first raw material and the second raw material mixed by the previous mixing channel 11 will collide after a 90° turn when entering the next mixing channel 11, thereby significantly improving the efficiency and uniformity of mixing the first raw material and the second raw material.

[0078] In the embodiment, the plurality of mixing channels 11 are helically arranged along a straight line, which is conducive to the layout of the 3D mixing unit, and the included angles between the plurality of mixing channels 11 are sequentially increased or decreased. For example, the included angles of the planes in which the plurality of mixing channels 11 are located are 0°, 90°, 180°, 270°, 0°, and so on from the front end to the rear end, and are sequentially helically arranged. In this way, the 3D mixing unit is distributed along a straight line as a whole, which is conducive to the parallel arrangement and distribution of the plurality of 3D mixing units 10, and the planes in which the adjacent two mixing channels 11 are located have the same included angle, which is conducive to the production of the 3D mixing unit with different numbers of mixing channels 11.

[0079] In other embodiments, the plurality of mixing channels 11 can also be helically arranged along a curve, and the planes in which the adjacent mixing channels 11 are located can also have different included angles to meet the needs of different use scenarios.

[0080] In the embodiment, one or both of the first channel 111 and the second channel 112 of the mixing channel 11 can be arranged in a bent manner, and the first channel 111 and the second channel 112 can be bent into a broken line or a curve. At least one of the first channel 111 and the second channel 112 is arranged in a bent manner, so that when the first channel 111 and the second channel 112 converge, collision mixing can be formed to improve the mixing effect.

[0081] The included angle between the inlet of the first channel 111 and the inlet of the second channel 112 is 15°-165°, for example, the included angle is 15°, 25°, 30°, 45°, 60°, 70°, 80°, 90°, 100°, 110°, 120°, 135°, 145°, 155°, and 165°, etc., which ensures that the inlet of the first channel 111 and the inlet of the second channel 112 are at least different by ±15°, so that the first raw material and the second raw material can realize collision mixing when they converge, and at least collide after a 15° turn, thereby improving the efficiency and uniformity of the mixing of the first raw material and the second raw material.

[0082] In a preferred embodiment, the included angle between the inlet of the first channel 111 and the inlet of the second channel 112 is 45°-135°, for example, the included angle is 45°, 60°, 70°, 80°, 90°, 100°, 110°, 120°, and 135°, etc., which ensures that the inlet of the first channel 111 and the inlet of the second channel 112 are at least different by ±45°, so that the first raw material and the second raw material can realize collision mixing when they converge, and at least collide after a 45° turn, thereby significantly improving the efficiency and uniformity of the mixing of the first raw material and the second raw material.

[0083] More preferably, in the embodiment, the included angle between the inlet of the first channel 111 and the inlet of the second channel 112 is 90°, which ensures that the inlet of the first channel 111 and the inlet of the second channel 112 are 90° apart, so that the first raw material and the second raw material can be mixed by collision when they meet, and the collision after a 90° turn can significantly improve the efficiency and uniformity of the mixing of the first raw material and the second raw material.

[0084] In the embodiment, the inner diameters of the first channel 111 and the second channel 112 are related to the particle size of the prepared mixture, and the inner diameters of the first channel 111 and the second channel 112 can be designed according to the desired production of the mixture, for example, the inner diameters of the first channel 111 and the second channel 112 are 1.5mm-0.2mm, wherein the inner diameters of the first channel 111 and the second channel 112 can be the same or different.

[0085] The 3D mixing unit 10 of the embodiment, since in the 3D mixing unit 10, any two adjacent mixing channels 11 are located in different planes, i.e., the plurality of mixing channels 11 form a 3D layout, so that when the fluid in the mixing channel 11 enters the next mixing channel 11, it will enter another spatial plane, increasing the probability of collision between the fluids, improving the efficiency of fluid mixing, and improving the mixing effect, thereby solving the problems of low mixing efficiency and poor mixing effect of lipid nanoparticle water phase and organic phase.

[0086] Embodiment two:

[0087] Please refer to FIG. 2, in the embodiment, a 3D microfluidic chip 100 for producing nanobiological medicine is provided, the 3D microfluidic chip 100 includes a first inlet channel 20, a second inlet channel 30, an outlet channel 40 and the 3D mixing unit 10 in the above embodiment one.

[0088] The first inlet channel 20 and the second inlet channel 30 are arranged at the front end of the 3D mixing unit 10, the outlet of the first inlet channel 20 and the outlet of the second inlet channel 30 are in communication with the common inlet of the frontmost mixing channel 11, the first inlet channel 20 is used for injecting the first raw material into the mixing channel 11, and the second inlet channel 30 is used for injecting the second raw material into the mixing channel 11. The outlet channel 40 is arranged at the rear end of the mixing channel 11, the inlet of the outlet channel 40 is in communication with the common outlet of the last mixing channel 11, and the outlet channel 40 is used for discharging the mixture formed by mixing.

[0089] The 3D microfluidic chip 100 further comprises a substrate 101, which can be a flat plate structure, and the 3D mixing unit 10, the first inlet channel 20, the second inlet channel 30 and the outlet channel 40 are located in the substrate 101. The inlet of the first inlet channel 20, the inlet of the second inlet channel 30 and the outlet of the outlet channel 40 can be exposed on the same side of the substrate 101 to realize the injection of the first raw material and the second raw material and the discharge of the mixture.

[0090] The substrate 101 can comprise an upper layer plate and a lower layer plate, and the upper layer plate and the lower layer plate combine to form the 3D mixing unit 10, the first inlet channel 20, the second inlet channel 30 and the outlet channel 40. The substrate 101 can also be an integrated structure, and the 3D mixing unit 10, the first inlet channel 20, the second inlet channel 30 and the outlet channel 40 can be made on the substrate 101 by injection molding or 3D printing.

[0091] Referring to FIG. 2, in the present embodiment, the 3D microfluidic chip 100 can further comprise a third inlet channel 50, which is a transition channel. The first inlet channel 20 and the second inlet channel 30 communicate with the 3D mixing unit 10 through the third inlet channel 50. The inlet of the third inlet channel 50 communicates with the outlet of the first inlet channel 20 and the outlet of the second inlet channel 30, and the outlet of the third inlet channel 50 communicates with the common inlet of the mixing channel 11 located at the front end of the 3D mixing unit 10. The first raw material and the second raw material can be pre-mixed in the third inlet channel 50. The arrangement of the third inlet channel 50 can improve the uniformity of the mixture of the first raw material and the second raw material.

[0092] The third inlet channel 50 is distributed on a different axial direction from the first inlet channel 20 and the second inlet channel 30, so that the connection between the first inlet channel 20, the second inlet channel 30 and the third inlet channel 50 forms a turn, so that the first raw material and the second raw material can collide when they meet, thereby achieving a certain mixing effect.

[0093] The angle between the first inlet channel 20 and the third inlet channel 50 can be 15°-165°, and the angle between the second inlet channel 30 and the third inlet channel 50 can be 15°-165°.

[0094] For example, the angle between the first inlet channel 20 and the third inlet channel 50 can be 90°, and the angle between the second inlet channel 30 and the third inlet channel 50 can be 90°. The third inlet channel 50 forms a T-shaped distribution with the first inlet channel 20 and the second inlet channel 30, which can significantly improve the mixing effect of the first raw material and the second raw material.

[0095] In other embodiments, the third inlet channel 50 has an included angle with one of the first inlet channel 20 and the second inlet channel 30, and is parallel or collinear with the other, which can also play a certain mixing effect of the first raw material and the second raw material.

[0096] In other embodiments, the 3D microfluidic chip 100 can also not include the third inlet channel 50, and the first inlet channel 20 and the second inlet channel 30 are directly communicated with the 3D mixing unit 10, and the mixing of the 3D mixing unit 10 can also realize the mixing of multiple raw materials to obtain a mixture.

[0097] Please refer to FIG. 3, in the embodiment, the same side of the substrate 101 of the 3D microfluidic chip 100 is provided with the first fluid connector 61, the second fluid connector 62 and the third fluid connector 63. The first fluid connector 61, the second fluid connector 62 and the third fluid connector 63 can be female luer type fluid connectors or other connectors with good sealing connection effect.

[0098] The first fluid connector 61, the second fluid connector 62 and the third fluid connector 63 are respectively vertically installed on the same side of the substrate 101. The first fluid connector 61 is in communication with the inlet of the first inlet channel 20, the second fluid connector 62 is in communication with the inlet of the second inlet channel 30, and the third fluid connector 63 is in communication with the outlet of the third inlet channel 50. The first fluid connector 61, the second fluid connector 62 and the third fluid connector 63 play a role of adapter. The first fluid connector 61 is used to connect a device containing the first raw material, the second fluid connector 62 is used to connect a device containing the second raw material, and the third fluid connector 63 is used to connect a device for collecting the mixture.

[0099] In other embodiments, the first fluid connector 61, the second fluid connector 62 and the third fluid connector 63 can also be arranged on different sides of the substrate 101, for example, the first fluid connector 61 and the second fluid connector 62 are arranged on the same side of the substrate 101, and the third fluid connector 63 is arranged on the other side of the substrate 101. Correspondingly, the inlet of the first inlet channel 20 and the inlet of the second inlet channel 30 are located on the same side of the substrate 101, and the outlet of the outlet channel 40 is located on the other side of the substrate 101.

[0100] In the embodiment, the 3D microfluidic chip includes a 3D mixing unit 10 with a 3D structure.

[0101] The differences between the 3D microfluidic chip of the present embodiment and the traditional 2D ring-shaped microfluidic mixer in preparing LNP are that four chip scales are set in the experiment: 2D-0.2mm-9 rings, 2D-0.5mm-9 rings, 3D-0.2mm-8 rings, and 3D-0.2mm-16 rings. The effects of the four chips on the preparation of empty LNP at 4ml / min, 8ml / min, 12ml / min, and 16ml / min are compared, and the effects on the preparation of mRNA-LNP are further compared on this basis. Among them, the number of rings refers to the number of mixing channels 11, for example, 9 rings, which means that the 3D mixing unit 10 includes 9 mixing channels 11.

[0102] Referring to FIG. 4, it can be seen from FIG. 4 that for the same chip, as the flow rate increases, the LNP particle size gradually decreases, and the particle size of the 2D chip 0.2mm is smaller than that of the 0.5mm. Similarly, for the 0.2mm, the particle size of the 3D is smaller than that of the 2D. Similarly, for the 3D-0.2mm 16 rings, the particle size is slightly smaller than that of the 8 rings.

[0103] To further compare the effects of 3D and 2D, 3D-0.2mm and 2D-0.2mm are used to prepare mRNA-LNP under the same mRNA, lipid and process conditions, and the results are shown in Table 1. After purification, the particle size of the 3D is smaller than that of the 2D, and the encapsulation efficiency of the 3D is significantly improved compared with the 2D.

[0104] Table 1. Comparison of 3D-0.2mm and 2D-0.2mm mRNA-LNP preparation results

[0105] Referring to Table 1 again, it can be seen that compared with the traditional 2D ring-shaped microfluidic mixer, the 3D microfluidic chip can obtain a mixture with smaller particle size and better mixing effect.

[0106] The 3D microfluidic chip 100 of the present embodiment, wherein the inner diameter of the chip can be set to 5.0mm-0.05mm, i.e., the inner diameter of the first channel 111 and the second channel 112 is 5.0mm-0.05mm, wherein the preferred scheme is that the inner diameter of the first channel 111 and the second channel 112 is 2.0mm-0.1mm, and the more preferred scheme is that the inner diameter of the first channel 111 and the second channel 112 is 1.5mm-0.2mm. For example, the inner diameter of the first channel 111 and the second channel 112 is 0.2mm, 0.5mm, 0.75mm, 1.0mm, 1.25mm, and 1.5mm, so as to more accurately produce LNP of different sizes for matching the delivery requirements of different tissues and organs.

[0107] The targeted delivery of tissue organs is the goal of drug delivery, and passive tissue localization mainly depends on the LNP particle size and charge, which is affected by 3D microfluidic chip, lipid composition and process conditions. Referring to FIG. 5, the influence of the chip inner diameter on the LNP particle size and PDI is shown. Under the same lipid and process conditions, as the chip inner diameter decreases, the produced LNP particle size also decreases, from 1.5 mm inner diameter to 0.2 mm inner diameter, the LNP particle size decreases from ~120 nm to ~60 nm, and the PDI is controlled within 0.1, which reflects the high uniformity of the particles. Referring to FIG. 6, the data trend line of LNP particle size y and chip inner diameter x satisfies the following regression equation: y = -17.612x1 2 +72.706x1+47.987

[0108] wherein x1 is the inner diameter of the 3D microfluidic chip 100, i.e. the inner diameter of the first channel 111 and the second channel 112, and y is the mixture particle size (LNP particle size).

[0109] The regression coefficient R 2 = 0.9965 in the above regression equation is very high. Therefore, according to the chip inner diameter and process control, LNP of different sizes can be accurately produced, as shown in Table 2.

[0110] Table 2. Comparison table of LNP particle size theoretical value and experimental value

[0111] The 3D microfluidic chip 100 in the embodiment belongs to a 3D structure, and the 3D microfluidic chip 100 is provided with a 3D mixing unit 10. Any two adjacent mixing channels 11 are located in different planes, i.e. the plurality of mixing channels 11 form a 3D layout, so that when the fluid in the mixing channel 11 enters the next mixing channel 11, it will enter another spatial plane, increasing the probability of collision between the fluids, improving the fluid mixing efficiency and improving the mixing effect.

[0112] Example Three

[0113] The embodiment provides a 3D microfluidic chip 100 for producing nanobiological medicines. The difference between the 3D microfluidic chip 100 of the embodiment and the above-mentioned embodiment two is that the 3D microfluidic chip 100 of the embodiment comprises a plurality of 3D mixing units 10, which can improve the throughput of the 3D microfluidic chip 100 and improve the production efficiency.

[0114] Please refer to FIG. 7, in the embodiment, the 3D microfluidic chip 100 further comprises a split channel 70 and a merging channel 80, and a plurality of 3D mixing units 10 are arranged between the split channel 70 and the merging channel 80. The front end of the split channel 70 is connected with the first inlet channel 20 and the second inlet channel 30, and the rear end of the merging channel 80 is connected with the outlet channel 40.

[0115] The split channel 70 comprises one or two inlets and a plurality of outlets, the first inlet channel 20 and the second inlet channel 30 are communicated with one inlet of the split channel 70, or the first inlet channel 20 and the second inlet channel 30 are respectively communicated with two inlets of the split channel 70. The number of outlets of the split channel 70 is the same as the number of 3D mixing units 10, and the plurality of outlets of the split channel 70 are one-to-one corresponding communication with the plurality of 3D mixing units 10. The merging channel 80 comprises a plurality of inlets and one outlet, the number of inlets of the merging channel 80 is the same as the number of 3D mixing units 10, and the plurality of inlets of the merging channel 80 are one-to-one corresponding communication with the plurality of 3D mixing units 10, and the outlet of the merging channel 80 is communicated with the outlet channel 40.

[0116] For example, the 3D microfluidic chip 100 comprises five 3D mixing units 10, the split channel 70 is provided with five outlets and the merging channel 80 is provided with five outlets corresponding connection, which can realize the simultaneous mixing of five 3D mixing units 10 to produce mixed substances.

[0117] The plurality of 3D mixing units 10 in the 3D microfluidic chip 100 can be arranged in parallel and spaced apart, and the central axes of the 3D mixing units 10 are distributed in the same plane, so that the layout of the plurality of 3D mixing units 10 is more compact, and the large flux spacing miniaturization of the 3D microfluidic chip 100 can be realized. Of course, the plurality of 3D mixing units 10 can also be arranged in a plurality of parallel planes.

[0118] The 3D microfluidic chip 100 of the present embodiment uses the same mechanism of chip to solve the problem of production amplification, ensuring the repeatability of process and quality before and after amplification, and avoiding the adverse effects on the quality of the product after amplification. For most applications, the ideal LNP particle size is in the range of about 80-120 nm, and the initial size is usually about 80 nm, which gradually increases with the production process and storage and transportation. As shown in Table 3, the ideal preparation flow rate should be 40 ml / min, which is equivalent to 2.4 L / hour, and 9.6 L / batch can be produced per 4 hours / batch, which is equivalent to 96,000 doses / batch according to 0.1 ml / dose vaccine, which is enough to meet the clinical demand. For commercial large-scale production, we designed a unique parallel chip, and Fig. 7 takes a 5-parallel 3D mixing unit 10 as an example. Since the structure and function of each channel are consistent, the stability of the product quality can be ensured when the production is amplified. Using a 5-parallel chip, 96 L can be produced per 10 hours / batch, which is equivalent to 1.2 million doses / batch according to 0.1 ml / dose, and 3.96 million doses can be produced per year according to 1 batch per day and 330 batches per year. The 5-parallel chip can further increase the number of parallel channels to increase the production capacity. Another amplification scheme is to simply repeat the 5-parallel chip production, and three 5-parallel chips can produce 1.188 billion doses, which can meet the demand for large-scale vaccination during the epidemic.

[0119] Table 3. Comparison table of amplification routes of 3D microfluidic chip 100 for laboratory preparation, clinical production and commercial production

[0120] As shown in Table 3 above, the present 3D microfluidic chip 100 with multiple channels can greatly improve the yield to meet market demand.

[0121] Embodiment Four:

[0122] Referring to Fig. 8, the present embodiment provides a 3D mixing system for producing nanobiological medicines, which comprises the 3D microfluidic chip 100 in the above-mentioned embodiment three or embodiment four, further comprises a first raw material supply device 200, a second raw material supply device 300 and a collection device 400, and the 3D mixing system can further be provided with more raw material supply devices when the 3D microfluidic chip 100 needs more raw material mixing.

[0123] The first raw material supply device 200 is used for storing the first raw material, and the first raw material supply device 200 is connected with the first fluid joint 61 through a pipeline. The first raw material supply device 200 is used for injecting the first raw material into the 3D microfluidic chip 100.

[0124] The second raw material supply device 300 is used for storing the second raw material, and the second raw material supply device 300 is connected with the second fluid joint 62 through a pipeline, and the second raw material supply device 300 is used for injecting the second raw material into the 3D micro-fluidic chip 100.

[0125] The collecting device 400 is connected with the third fluid joint 63 through a pipeline, and the collecting device 400 is used for collecting the mixture prepared by the 3D micro-fluidic chip 100.

[0126] Please refer to FIG. 9, in the embodiment, the 3D mixing system can further include a driving device 500, the driving device 500 can include two, one driving device 500 is connected with the first raw material supply device 200, and is used for driving the first raw material to be injected into the 3D micro-fluidic chip 100, and the other driving device 500 is connected with the second raw material supply device 300, and is used for driving the second raw material to be injected into the 3D micro-fluidic chip 100. The setting of the driving device 500 can realize the automatic injection of the first raw material and the second raw material, and control the injection ratio of the first raw material and the second raw material, and can also control the flow rate of the first raw material and the second raw material.

[0127] In other embodiments, the first raw material supply device 200 and the second raw material supply device 300 are provided with driving components, and the automatic injection of the first raw material and the second raw material can also be realized.

[0128] Please refer to FIG. 10, in other embodiments, the 3D mixing system can further include a cleaning device 600 and a waste liquid device 700, the cleaning device 600 is used for storing cleaning liquid, the cleaning device 600 is connected with the first fluid joint 61 and the second fluid joint 62 of the 3D micro-fluidic chip 100 through a pipeline, and the cleaning device 600 is used for injecting the cleaning liquid into the 3D micro-fluidic chip 100, and cleaning the flow channel in the 3D micro-fluidic chip 100. The waste liquid device 700 is connected with the third fluid joint 63 through a pipeline, and the waste liquid device 700 is used for collecting the waste liquid after cleaning.

[0129] Among them, the pipeline connected with the first fluid joint 61, the second fluid joint 62 and the third fluid joint 63 is provided with a multi-way valve, which can be used for switching the injection of raw materials or cleaning liquid, and switching the discharge of mixed liquid or waste liquid into different devices.

[0130] In the embodiment, the 3D mixing system includes the 3D micro-fluidic chip 100 in the above-mentioned embodiments. Since any two adjacent mixing channels 11 in the 3D mixing unit 10 are located in different planes, that is, the plurality of mixing channels 11 form a 3D layout, when the fluid in the mixing channel 11 enters the next mixing channel 11, it will enter another spatial plane, increasing the probability of collision between the fluids, and improving the fluid mixing efficiency and the mixing effect.

[0131] Embodiment five:

[0132] A method for mixing product is provided in this embodiment, which is realized by using the 3D microfluidic chip 100 or 3D mixing system in the above embodiments.

[0133] The method mainly includes the following two steps:

[0134] The first raw material and the second raw material are respectively injected into the 3D microfluidic chip 100;

[0135] The first raw material and the second raw material are mixed in the 3D mixing unit 10 to form a mixture.

[0136] The above two steps can be realized by a controller to automatically mix, so as to control the injection ratio of the first raw material and the second raw material, the injection speed of the first raw material and the second raw material, and further control the flow rate of the mixed first raw material and second raw material.

[0137] Among them, the particle size of the product can be controlled by controlling the flow rate in the 3D microfluidic chip 100.

[0138] Please refer to FIG. 11 and FIG. 12, under the condition of fixed chip inner diameter, process flow rate affects LNP particle size (product), and increasing flow rate can significantly reduce LNP particle size. Under the condition of same chip inner diameter and flow rate, increasing the number of rings (the number of mixing channels) can slightly reduce LNP particle size.

[0139] Raw material parameters such as N / P ratio (lipid RNA ratio) and PEG lipid ratio can affect LNP particle size, but changes in these raw material parameters will significantly change the efficacy and safety of the product, so they need to be strictly controlled within a small change range. Under the condition that the raw material parameters are unchanged, the chip inner diameter and flow rate become the key control factors of LNP particle size.

[0140] The flow rate and LNP particle size data of 3D-0.5MM-12 rings are linearly regressed to obtain Table 4 and the following regression equation: y = 0.0718x2 2 -5.4452x2+185.47

[0141] Among them, x2 is the flow rate of the first raw material and the second raw material in the 3D mixing unit 10, y is the particle size of the mixture (LNP particle size), and the regression coefficient R 2 of the above equation is 0.9955.

[0142] Table 4. Calculated values of LNP particle size corresponding to different flow rates

[0143] It can be seen that the faster the flow rate, the smaller the LNP particle size, and different particle sizes of LNP can be controlled by controlling the flow rate to meet different production needs.

[0144] Example 6

[0145] The present example provides an application of the 3D microfluidic chip 100 in the preparation of products using the 3D microfluidic chip 100 or the 3D hybrid system in the above examples.

[0146] The 3D microfluidic chip 100 can be used to produce particles carrying drug molecules or cells or pharmaceutical compositions, for example, the particles are lipid nanoparticles. The 3D microfluidic chip 100 can also be used to produce other forms of nano or microparticles for application in different fields.

[0147] As shown in Table 5, the 3D microfluidic chip 100 can be used to produce nano or microparticles based on lipids or polymers, etc. for delivery of small molecules, biological macromolecules and cells, for various medical, therapeutic, diagnostic and detection fields.

[0148] Table 5. Application fields of 3D microfluidic chip

[0149] The above application of specific examples is used to illustrate the present application, which is only used to help understand the present application and does not limit the present application. For those skilled in the art to which the present application belongs, according to the idea of the present application, a number of simple deductions, deformations or substitutions can be made.

Claims

1. A 3D mixing unit for producing nanobiopharmaceuticals, characterized by, The mixing channel comprises a plurality of mixing channels connected in sequence: The mixing channel comprises a first channel and a second channel, the inlet of the first channel is connected with the inlet of the second channel to form a common inlet, and the outlet of the first channel and the outlet of the second channel are connected to form a common outlet; The common inlet of the mixing channel at the front end is used at least for injecting a first raw material and a second raw material, the common outlet of the mixing channel at the rear end is used for discharging a mixture formed by mixing the first raw material and the second raw material, the common inlet of the mixing channel at the middle part is communicated with the common outlet of the adjacent mixing channel, and the common outlet of the mixing channel at the middle part is communicated with the common inlet of the adjacent mixing channel; The first channel and the second channel of the mixing channel are located in the same plane, and any two adjacent mixing channels are located in different planes.

2. The 3D mixing unit of claim 1, wherein, The included angle between the planes where the two adjacent mixing channels are located is 15°-165°.

3. The 3D mixing unit of claim 2, wherein, The included angle between the planes where the two adjacent mixing channels are located is 45°-135°.

4. The 3D mixing unit of claim 3, wherein, The included angle between the planes where the two adjacent mixing channels are located is 90°.

5. The 3D mixing unit of claim 1, wherein, The plurality of mixing channels are spirally arranged along a straight line.

6. The 3D mixing unit of claim 1, wherein, The first channel and / or the second channel are arranged in a bent manner, the included angle between the inlet of the first channel and the inlet of the second channel is 15°-165°, and the included angle between the outlet of the first channel and the outlet of the second channel is 15°-165°.

7. The 3D mixing unit of claim 6, wherein, The included angle between the inlet of the first channel and the inlet of the second channel is 45°-135°, and the included angle between the outlet of the first channel and the outlet of the second channel is 45°-135°.

8. The 3D mixing unit of claim 7, wherein, The included angle between the inlet of the first channel and the inlet of the second channel is 90°, and the included angle between the outlet of the first channel and the outlet of the second channel is 90°.

9. The 3D mixing unit of claim 1, wherein, The inner diameter of the first channel and the second channel is 5.0mm-0.05mm; Preferably, the inner diameter of the first channel and the second channel is 2.0mm-0.1mm; More preferably, the inner diameter of the first channel and the second channel is 1.5mm-0.2mm.

10. The 3D mixing unit of claim 9, wherein, The inner diameters of the first and second channels and the particle size of the mixture satisfy the following regression formula requirements: y = -17.612x1 2 + 72.706x1 + 47.987 Wherein, x1 is the inner diameter of the first channel and the second channel, and y is the particle size of the mixture.

11. A 3D microfluidic chip for producing nanobiopharmaceuticals, characterized by, Comprise: The first inlet channel, the second inlet channel, the outlet channel and the 3D mixing unit according to any one of claims 1-10, the first inlet channel and the second inlet channel are communicated with the common inlet of the mixing channel at the front end, and the outlet channel is communicated with the common outlet of the mixing channel at the rear end.

12. The 3D microfluidic chip of claim 11, wherein, Further comprising a third inlet channel, the outlet of the first inlet channel and the outlet of the second inlet channel are communicated with the inlet of the third inlet channel, and the outlet of the third inlet channel is communicated with the common inlet of the mixing channel at the front end.

13. The 3D microfluidic chip of claim 12, wherein, The first inlet channel and / or the second inlet channel and the third inlet channel are distributed on different axes.

14. The 3D microfluidic chip of claim 13, wherein, The included angle between the first inlet channel and / or the second inlet channel and the third inlet channel is 15°-165°.

15. The 3D microfluidic chip of claim 14, wherein, The included angle between the first inlet channel and / or the second inlet channel and the third inlet channel is 90°.

16. The 3D microfluidic chip of claim 11, wherein, Further comprising a split channel and a converging channel, a plurality of 3D mixing units are arranged between the split channel and the converging channel; the split channel comprises one or two inlets and a plurality of outlets, the first inlet channel and the second inlet channel are in communication with the inlets of the split channel, and the plurality of outlets of the split channel are in one-to-one correspondence with the plurality of 3D mixing units; the converging channel comprises a plurality of inlets and an outlet, the plurality of inlets of the converging channel are in one-to-one correspondence with the plurality of 3D mixing units, and the outlet of the converging channel is in communication with the outlet channel.

17. The 3D microfluidic chip of claim 16, wherein, The plurality of 3D mixing units are arranged in parallel and at intervals.

18. The 3D microfluidic chip of any one of claims 11 to 17, wherein, The 3D microfluidic chip further comprises a substrate, and the first inlet channel, the second inlet channel, the outlet channel and the 3D mixing units are located in the substrate.

19. The 3D microfluidic chip of claim 18, wherein, The substrate is a flat plate structure, and the inlet of the first inlet channel, the inlet of the second inlet channel and the outlet of the outlet channel are located on the same side of the substrate.

20. The 3D microfluidic chip of claim 19, wherein, The same side of the substrate is provided with a first fluid connector, a second fluid connector and a third fluid connector, the first fluid connector is in communication with the inlet of the first inlet channel, the second fluid connector is in communication with the inlet of the second inlet channel, and the third fluid connector is in communication with the outlet of the outlet channel.

21. The 3D microfluidic chip of claim 20, wherein, The first fluid connector, the second fluid connector and the third fluid connector are female luer type fluid connectors.

22. A 3D mixing system for producing nanobiopharmaceuticals, characterized by, Comprising: The 3D microfluidic chip according to any one of claims 11 to 21; A first raw material supply device for storing a first raw material, the first raw material supply device being in communication with the first inlet channel, and the first raw material supply device being used for injecting the first raw material into the first inlet channel; A second raw material supply device for storing a second raw material, the second raw material supply device being in communication with the second inlet channel, and the second raw material supply device being used for injecting the second raw material into the second inlet channel; And A collection device in communication with the outlet channel, the collection device being used for collecting a mixture generated by mixing the first raw material and the second raw material.

23. The 3D mixing system of claim 22, wherein, Further comprising a driving device connected with the first raw material supply device and the second raw material supply device, the driving device being used for driving the first raw material to be injected into the first inlet channel and driving the second raw material to be injected into the second inlet channel.

24. The 3D mixing system of claim 22, wherein, Further comprising a cleaning device and a waste liquid device, the cleaning device being in communication with the first inlet channel and the second inlet channel, and the cleaning device being used for injecting cleaning liquid; the waste liquid device being in communication with the outlet channel, and the waste liquid device being used for collecting waste cleaning liquid.

25. A method for mixing a product by using the 3D microfluidic chip according to any one of claims 11 to 21 or the 3D mixing system according to any one of claims 22 to 24, characterized in that, Comprising the following steps: Injecting the first raw material and the second raw material into the 3D microfluidic chip respectively; Mixing the first raw material and the second raw material in the 3D mixing unit to form a mixture particle size of nanobiological medicine.

26. The method of claim 25, wherein, The flow rate of the first raw material and the second raw material in the 3D mixing unit and the particle size of the mixture satisfy the following regression equation: y = 0.0718x2 2 -5.4452x2+185.47 Wherein, x2 is the flow rate of the first raw material and the second raw material in the 3D mixing unit, and y is the mixture particle size.

27. Use of a 3D microfluidic chip according to any one of claims 11 to 21 or a 3D hybrid system according to any one of claims 22 to 24 for the preparation of a product; Preferably, the product is a particle or a pharmaceutical composition loaded with a drug molecule or a cell; More preferably, the particle is a lipid nanoparticle for the delivery of a drug.

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