SOD (superoxide dismutase) compound preparation and preparation process thereof

Abstract

The invention discloses an SOD (superoxide dismutase) compound preparation and a preparation process thereof, and relates to the technical field of biochemical engineering. The preparation is composed of an SOD-trehalose eutectic nucleus, a lecithin / cholesterol bilayer and a hyaluronic acid targeting shell. The composition comprises the following components in percentage by mass: 0.5 to 1.2 percent of SOD (superoxide dismutase), 8 to 12 percent of trehalose, 4.5 to 6.0 percent of lecithin / cholesterol and 0.8 to 1.5 percent of hyaluronic acid. The preparation process comprises the following steps: 1) micro-fluidic low-temperature eutectic: emulsifying SOD and trehalose at 4 + / -1 DEG C through a micro-fluidic chip, and freezing at-30 + / -2 DEG C to form eutectic particles of 2.5 + / -0.3 mu m; 2) supercritical in-situ embedding: the particles react with lecithin / cholesterol for 40 + / -5 min in supercritical CO at 8 + / -0.5 MPa / 15 + / -2 DEG C, and the encapsulation efficiency is greater than or equal to 87%; and 3) cold spray drying: adding hyaluronic acid for homogenizing, and drying under the conditions that the air inlet temperature is less than or equal to 35 DEG C and the atomization pressure is 0.15 + / -0.02 MPa. The process realizes that the SOD activity recovery rate is greater than or equal to 92%, the transdermal permeation amount is increased by 3.2 times, the storage half-life period at 4 DEG C is predicted to reach 18 months through an acceleration test, and the problems of large activity loss, low encapsulation efficiency and thermal damage in the traditional process are solved.

Description

Technical Field

[0001] This invention belongs to the field of biochemical technology and relates to a superoxide dismutase (SOD) complex preparation with high stability and targeting and its low-temperature integrated preparation process. Specifically, it relates to an SOD complex preparation and its preparation process, which is particularly suitable for the fields of pharmaceuticals, cosmetics and functional foods. Background Technology

[0002] Superoxide dismutase (SOD), a core antioxidant enzyme in organisms, can efficiently catalyze superoxide anion free radicals (O2). - SOD is converted into hydrogen peroxide and oxygen, playing a crucial role in combating oxidative stress damage, delaying cell aging, and treating inflammatory diseases. According to a Global Market Insights 2023 report, the global SOD formulation market will exceed US$4.8 billion in 2027, with its application demand continuing to rise in the pharmaceutical (such as the treatment of radiation dermatitis), high-end cosmetics (anti-aging serums), and functional foods (oral antioxidants).

[0003] Current industrial production technologies primarily rely on two technical routes: chemical modification and carrier encapsulation. Chemical modification, exemplified by polyethylene glycol (PEG) covalent coupling, extends the plasma half-life of SOD by modifying the amino groups on its surface. Carrier encapsulation, mainly using liposomes and nanoemulsions, utilizes phospholipid bilayers or oil-water interfaces to encapsulate enzyme molecules and isolate them from degradation. However, existing processes suffer from drawbacks such as significant activity loss, low delivery efficiency, and cumulative losses due to multi-stage process chains. Firstly, significant activity loss occurs. PEGylation requires an alkaline environment (pH 8.5-9.0), leading to the degradation of Cu in the SOD active site. 2+ / Zn 2+First, the coordination structure is disrupted, leading to enzyme activity loss. Although liposome encapsulation avoids chemical modification, the emulsification process requires temperatures above 45°C to achieve the phospholipid phase transition. High temperatures induce a decrease in the SOD α-helix content from 42% to 28%, resulting in a specific activity reduction of over 30%. Second, delivery efficiency is low. The encapsulation rate of hydrophilic SOD by liposomes is generally below 65%, mainly because the phospholipid bilayer and the hydrophilic core are difficult to form a stable inclusion. Although nanoemulsions can increase drug loading to 8%, surfactants (such as Tween 80) disrupt the hydrophobic core of the enzyme protein, accelerating inactivation during storage. After 7 days at 37°C, the residual activity is less than 50%. Third, the multi-stage process chain causes cumulative losses. The traditional process requires four separate steps: "encapsulation → pre-freezing → sublimation drying → targeted modification". High temperatures during the encapsulation stage damage the initial activity; ice crystal growth during freeze-drying generates shear stress, causing secondary inactivation; and the highly acidic environment (pH 4.5-5.5) during chemical coupling of the target molecule induces protein aggregation. Statistics show that the cumulative activity loss in the three-step process is as high as 40%, and the final product particle size distribution (PDI>0.3) leads to fluctuations in bioavailability.

[0004] The aforementioned shortcomings have limited the clinical application and economic value of SOD formulations: insufficient activity necessitates doubling the dosage, increasing the burden on patients; poor transdermal absorption limits the efficacy of topical formulations; and intermittent production models result in high costs. Therefore, developing a low-temperature integrated process that avoids thermal / chemical damage while achieving high activity retention, high encapsulation efficiency, and enhanced targeted delivery has become an urgent need to overcome industry bottlenecks. Summary of the Invention

[0005] The purpose of this invention is to address the problems of low activity retention, poor transdermal delivery efficiency, and large cumulative losses in multi-stage processes in SOD formulation production, and to provide a preparation process and dedicated integrated system based on a three-stage low-temperature coupling of microfluidic eutectic-supercritical in-situ embedding-cold spray drying.

[0006] To achieve the above objectives, the present invention provides the following technical solution: an SOD complex formulation, comprising an SOD-trehalose co-crystal nucleus, a lecithin-cholesterol bilayer, and a hyaluronic acid targeting shell, wherein the formulation contains the following components by mass percentage: Superoxide dismutase 0.5%–1.2%; Trehalose eutectic protectant 8%–12%; Lecithin / cholesterol ratio: 4.5%–6.0%; Hyaluronic acid 0.8%–1.5%; The remainder is a phosphate buffer system, pH 6.8±0.2.

[0007] A preparation process for an SOD complex formulation includes the following steps: Step 1: Mix SOD enzyme solution with a specific activity of not less than 4000 U / mg with trehalose at a mass ratio of 1:8 to 1:12, and pump the mixture into a microfluidic chip with a channel width of 180 to 220 μm. Under the conditions of temperature 2 to 5℃ and water phase to oil phase flow rate ratio of 1:5, a W / O emulsion is formed. Freeze the mixture at -28 to -32℃ for 30 minutes to form eutectic nuclei with a particle size of 2.2 to 2.8 μm. Step 2, In-situ Phospholipid Encapsulation: The eutectic nuclei obtained in Step 1 are pumped into a supercritical deposition reactor and dispersed in a supercritical CO2 fluid containing lecithin and cholesterol, wherein the molar ratio of lecithin to cholesterol is 3:1, the operating pressure is 7.5–8.5 MPa, the operating temperature is 13–17℃, and the reaction is stirred at a rate of 250–350 rpm for 35–45 minutes using a magnetic stirring system to form a bilayer lipid coating using supercritical antisolvent deposition technology; Step 3, Targeted Modification and Cold Spray Drying: Add a hyaluronic acid solution with a final concentration of 0.8% to 1.5% to the product from Step 2. After three cycles of high-pressure homogenization at 50 MPa, pump the solution into a cold spray drying tower and dry it under conditions where the inlet air temperature does not exceed 35°C and the atomization pressure is 0.13 to 0.17 MPa, controlling the moisture content to not exceed 3.0%, to obtain a free-flowing powder, i.e., the SOD complex preparation.

[0008] As a further embodiment of the present invention: the oil phase of the microfluidic chip in step one is a mixture of perfluoropolyether and cyclopentane, with a volume ratio of 7:3; the supercritical CO2 fluid in step two contains 0.05% to 0.1% by mass of vitamin E; the operating pressure in step two is 8.0 MPa, the operating temperature is 15°C, the magnetic stirring speed is 300 rpm, and the reaction time is 40 minutes; the dried powder obtained in step three is mixed with 0.1% by mass of nano-silica flow aid in a double-cone vacuum mixer, based on the total mass of the powder.

[0009] As a further embodiment of the present invention: the supercritical deposition reactor includes a reactor body, a reactor lid at the top of the reactor body, a feed pipe and a vent pipe fixedly connected to both sides of the top of the reactor lid, a supersonic particle nozzle installed at the bottom of the feed pipe, an enamel lining on the inner wall of the reactor body, a discharge pipe fixedly connected to the bottom of the reactor body, a valve installed on the outer wall of the discharge pipe, a temperature control coil fixedly connected to the outer wall of the reactor body, a fixing seat fixedly connected to the top of the reactor lid, and a mounting base fixedly connected to the inner wall of the fixing seat. A sealing shell is installed at the top of the mounting base. A stirring shaft is rotatably connected to the mounting base. A stirring head is installed on the outer wall of the stirring shaft. An inner magnetic rotor is installed at the top of the stirring shaft. A protective shell is installed at the top of the fixed base. A motor is installed at the top of the protective shell. An outer magnetic rotor is connected to the output end of the motor. A sealing ring is fixedly connected at the junction of the top of the mounting base and the sealing shell. The inner and outer magnetic rotors can be quickly installed and removed through a replacement mechanism. The stirring head is installed through an installation mechanism.

[0010] As a further embodiment of the present invention: the replacement mechanism includes through holes symmetrically opened on both sides of the protective shell; mounting holes symmetrically opened at the top of the fixing base; bolts threadedly connected to the inner wall of the mounting holes; connecting grooves symmetrically opened on both sides of the sealing shell; connecting blocks symmetrically fixedly connected to the top of the mounting base; a locking block extending from the connecting block is slidably connected to the inner wall of the connecting block; a first spring is connected between the locking block and the connecting block; a push plate is slidably connected inside the fixing base and the mounting base at the bottom of the mounting holes; and a connection is made between the push plate and the fixing base. The second spring, the top of the push plate is fixedly connected to a T-shaped frame, the T-shaped frame passes through the locking block, the top of the locking block is provided with a positioning groove, the inner wall of the inner magnetic rotor is symmetrically provided with fixing grooves, the top of the stirring shaft is rotatably connected to a rotating disk, the bottom of the rotating disk is fixedly connected to a threaded rod, the outer wall of the threaded rod is slidably connected to a displacement frame, the displacement frame is slidably connected to the inside of the stirring shaft, the inside of the stirring shaft is slidably connected to both sides of the displacement frame, the bottom of the displacement frame is fixedly connected to a sliding rod, the outer wall of the fixing block is provided with a sliding groove for the sliding rod to slide.

[0011] As a further embodiment of the present invention: the mounting mechanism includes a groove, the groove being formed at the top of the mounting base; a positioning plate is fixedly connected to the outer wall of the stirring shaft within the inner cavity of the groove; a positioning ring is slidably connected to the inner wall of the groove at the top of the positioning plate; a limiting seat is fixedly connected to the outer wall of the stirring shaft within the inner cavity of the vessel body; a threaded seat is fixedly connected to the top of the limiting seat; a rotating ring is threadedly connected to the outer wall of the threaded seat; transverse grooves are symmetrically formed on the outer wall of the threaded seat; a crossbar is fixedly connected to the outer wall of the stirring head; a slot is formed on the outer wall of the crossbar; a lower pressure frame is slidably connected inside the stirring shaft; the positioning plate extends from the top of the lower pressure frame; a third spring connects the lower pressure frame and the positioning plate; and an insertion rod is fixedly connected to the outer wall of the lower pressure frame, the insertion rod being located above the transverse groove.

[0012] As a further embodiment of the present invention: the inner wall of the connecting groove is in contact with the outer wall of the connecting block, the end of the card block extending out of the connecting block is provided with an inclined surface, and the inner wall of the positioning groove is in contact with the top outer wall of the T-shaped frame.

[0013] As a further embodiment of the present invention: the top outer wall of the stirring shaft is in contact with the inner wall of the inner magnetic rotor, and the inner wall of the fixing groove is in contact with the outer wall of the fixing block.

[0014] As a further embodiment of the present invention: the top end of the displacement frame is provided with a threaded hole, the threaded hole is matched with the threaded rod, and the outer wall of the slide rod is in contact with the inner wall of the fixing block.

[0015] As a further embodiment of the present invention: the outer wall of the crossbar is fitted with the inner wall of the cross groove, and the inner wall of the slot is fitted with the outer wall of the insertion rod.

[0016] Compared with the prior art, the beneficial effects of the present invention are: 1. The preparation process of the SOD complex formulation of this invention integrates a three-stage low-temperature process of microfluidic co-crystallization, supercritical in-situ embedding, and cold spray drying, which improves production efficiency. Microfluidic low-temperature co-crystallization technology forms SOD-trehalose co-crystallization nuclei, effectively avoiding ice crystal damage. Supercritical CO2 embedding technology achieves high encapsulation efficiency at low temperatures, avoiding phospholipid membrane instability and enzyme inactivation caused by traditional high-temperature emulsification. The integrated process avoids intermediate transfers in traditional multi-stage processes, reducing cumulative activity loss and shortening the production cycle by 40%. Supercritical CO2 embedding coupled with cold spray drying completely eliminates high-temperature / chemical damage, reducing energy consumption by 35% compared to traditional spray drying. 2. The SOD complex formulation prepared in this invention improves activity and stability through a three-level structural design of co-crystal nucleus-lipid layer-targeting shell. Hyaluronic acid forms a hydrogen bond network with the phosphate groups of the lipid layer through its carboxyl groups, densifying the membrane structure and increasing transdermal penetration. The trehalose co-crystal nucleus locks the conformation of the SOD active site, resulting in an enzyme activity recovery rate ≥92% and a transdermal penetration of 82.5 μg / cm³ over 24 hours. 2 Hyaluronic acid-modified bilayer structure, formulation with a half-life of 18 months under accelerated conditions of 40℃ / RH75% (conventional liposomes ≤9 months), and a reconstitution time ≤28 seconds; 3. By setting up a replacement mechanism, the bolts are removed from the mounting holes, and the protective shell is taken off. At this time, the outer magnetic rotor can be replaced. Simultaneously, the bolts separate from the push plate, and the push plate is reset by the elastic force of the second spring. The reset of the push plate drives the T-shaped frame to move out of the positioning groove to cancel the fixation of the locking block. At this time, the locking block is pushed to separate from the sealing shell, so that the sealing shell can be disassembled. Rotating the rotating disk drives the fixing block to move out of the fixing groove, and the inner magnetic rotor is removed from the stirring shaft, completing the disassembly operation of the inner magnetic rotor. This facilitates the quick disassembly of the inner and outer magnetic rotors, so that the replacement operation can be performed quickly when the inner and outer magnetic rotors are damaged. 4. By setting up an installation mechanism, the crossbar is inserted into the horizontal groove. When the positioning ring is placed at the top of the positioning plate, the positioning ring contacts the lower pressure frame, pushing the lower pressure frame to move and squeezing the third spring. The displacement of the lower pressure frame drives the insertion rod to move. The insertion rod moves into the slot to fix the stirring head, which facilitates the quick replacement of the stirring head. Thus, the shearing force generated when the stirring shaft rotates can be adjusted by replacing the stirring head. Attached Figure Description

[0017] Figure 1 This is a schematic diagram of the structure of the present invention; Figure 2 This is a cross-sectional view of the lid of the vessel according to the present invention; Figure 3 This is a schematic diagram of the motor installation according to the present invention; Figure 4 This is a schematic diagram of the installation of the protective shell of the present invention; Figure 5 This is a schematic diagram of the installation of the sealing shell of the present invention; Figure 6 This is a schematic diagram of the card block structure of the present invention; Figure 7 This is a schematic diagram of the installation of the internal magnetic rotor of the present invention; Figure 8 This is a schematic diagram of the displacement frame of the present invention; Figure 9 This is a schematic diagram of the installation of the stirring head of the present invention; Figure 10This is a schematic diagram of the lower pressure frame of the present invention.

[0018] In the diagram: 1. Reactor body; 2. Reactor lid; 3. Feed pipe; 4. Supersonic particle nozzle; 5. Vent pipe; 6. Discharge pipe; 7. Valve; 8. Replacement mechanism; 801. Through hole; 802. Bolt; 803. Mounting hole; 804. Connecting groove; 805. Connecting block; 806. Locking block; 807. First spring; 808. Push plate; 809. Second spring; 810. T-shaped frame; 811. Fixing groove; 812. Rotating disk; 813. Threaded rod; 814. Displacement frame; 815. Slide rod; 816. Fixing block; 817. Slide groove; 818. Positioning groove; 9. Mounting mechanism; 901. Groove; 902. Positioning plate; 903. Positioning ring; 904. Limiting seat; 905. Threaded seat; 906. Rotating ring; 907. Crossbar; 908. Slot; 909. Horizontal groove; 910. Lower pressure frame; 911. Third spring; 912. Insert rod; 10. Temperature control coil; 11. Fixed seat; 12. Mounting seat; 13. Sealing shell; 14. Stirring shaft; 15. Stirring head; 16. Inner magnetic rotor; 17. Protective shell; 18. Motor; 19. Outer magnetic rotor; 20. Sealing ring. Detailed Implementation

[0019] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0020] In the description of this invention, it should be noted that the terms "center," "upper," "lower," "left," "right," "vertical," "horizontal," "inner," and "outer," etc., indicating orientation or positional relationships, are based on the orientation or positional relationships shown in the accompanying drawings and are only for the convenience of describing this invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this invention. Furthermore, the terms "first," "second," and "third" are used for descriptive purposes only and should not be construed as indicating or implying relative importance. In the description of this invention, it should be noted that unless otherwise explicitly specified and limited, the terms "installed," "connected," "linked," and "set up" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal communication of two components. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances. The following describes embodiments of the invention based on its overall structure.

[0021] Please see Figures 1 to 10 In this embodiment of the invention, an SOD complex formulation is composed of an SOD-trehalose co-crystal core, a lecithin-cholesterol bilayer, and a hyaluronic acid targeting shell. The formulation contains the following components by mass percentage: Superoxide dismutase 0.5%–1.2%; Trehalose eutectic protectant 8%–12%; Lecithin / cholesterol ratio: 4.5%–6.0%; Hyaluronic acid 0.8%–1.5%; The remainder is a phosphate buffer system, pH 6.8±0.2.

[0022] A preparation process for an SOD complex formulation includes the following steps: Step 1: Mix SOD enzyme solution with a specific activity of not less than 4000 U / mg with trehalose at a mass ratio of 1:8 to 1:12, and pump the mixture into a microfluidic chip with a channel width of 180 to 220 μm. Under the conditions of temperature 2 to 5℃ and water phase to oil phase flow rate ratio of 1:5, a W / O emulsion is formed. Freeze the mixture at -28 to -32℃ for 30 minutes to form eutectic nuclei with a particle size of 2.2 to 2.8 μm. Step 2, In-situ Phospholipid Encapsulation: The eutectic nuclei obtained in Step 1 are pumped into a supercritical deposition reactor and dispersed in a supercritical CO2 fluid containing lecithin and cholesterol, wherein the molar ratio of lecithin to cholesterol is 3:1, the operating pressure is 7.5–8.5 MPa, the operating temperature is 13–17℃, and the reaction is stirred at a rate of 250–350 rpm for 35–45 minutes using a magnetic stirring system to form a bilayer lipid coating using supercritical antisolvent deposition technology; Step 3, Targeted Modification and Cold Spray Drying: Add a hyaluronic acid solution with a final concentration of 0.8% to 1.5% to the product from Step 2. After three cycles of high-pressure homogenization at 50 MPa, pump the solution into a cold spray drying tower and dry it under conditions where the inlet air temperature does not exceed 35°C and the atomization pressure is 0.13 to 0.17 MPa, controlling the moisture content to not exceed 3.0%, to obtain a free-flowing powder, i.e., the SOD complex preparation.

[0023] Step 1: The oil phase of the microfluidic chip is a mixture of perfluoropolyether and cyclopentane in a volume ratio of 7:3. Step 2: The supercritical CO2 fluid contains 0.05% to 0.1% by mass of vitamin E. Step 2: The operating pressure is 8.0 MPa, the operating temperature is 15℃, the magnetic stirring speed is 300 rpm, and the reaction time is 40 minutes. Step 3: The resulting dried powder is mixed with 0.1% by mass of nano-silica flow aid in a double-cone vacuum mixer, based on the total mass of the powder.

[0024] In this embodiment: The preparation process of the SOD complex formulation of the present invention integrates a three-stage low-temperature process of microfluidic eutectic crystallization, supercritical in-situ embedding, and cold spray drying, which improves production efficiency. The integrated process avoids intermediate transfers in traditional multi-stage processes, reducing the cumulative activity loss from 40% to ≤8% and shortening the production cycle by 40%; supercritical CO2 embedding coupled with cold spray drying completely eliminates high-temperature / chemical damage, reducing energy consumption by 35% compared to traditional spray drying; The SOD complex formulation prepared in this invention improves activity and stability through a three-level structural design of co-crystal nucleus-lipid layer-targeting shell. Hyaluronic acid forms a hydrogen bond network with the phosphate groups of the lipid layer through its carboxyl groups, densifying the membrane structure and increasing transdermal penetration. The trehalose co-crystal nucleus locks in the conformation of the SOD active site, resulting in an enzyme activity recovery rate ≥92% and a transdermal penetration of 82.5 μg / cm³ after 24 hours. 2 Hyaluronic acid modifies the bilayer structure, enabling the formulation to have a half-life of 18 months (compared to ≤9 months for traditional liposomes) and a reconstitution time of ≤28 seconds under accelerated conditions of 40℃ / RH75%.

[0025] Please refer to this carefully. Figures 1 to 3A supercritical sedimentation reactor includes a reactor body 1, a reactor cover 2 at the top of the reactor body 1, a feed pipe 3 and a vent pipe 5 fixedly connected to the two sides of the top of the reactor cover 2, a supersonic particle nozzle 4 installed at the bottom of the feed pipe 3, an enamel lining on the inner wall of the reactor body 1, a discharge pipe 6 fixedly connected to the bottom of the reactor body 1, a valve 7 installed on the outer wall of the discharge pipe 6, a temperature control coil 10 fixedly connected to the outer wall of the reactor body 1, a fixing seat 11 fixedly connected to the top of the reactor cover 2, a mounting seat 12 fixedly connected to the inner wall of the fixing seat 11, and a sealing shell 1 installed on the top of the mounting seat 12. 3. A stirring shaft 14 is rotatably connected to the mounting base 12. A stirring head 15 is installed on the outer wall of the stirring shaft 14. An inner magnetic rotor 16 is installed at the top of the stirring shaft 14. A protective shell 17 is installed at the top of the fixed base 11. A motor 18 is installed at the top of the protective shell 17. An outer magnetic rotor 19 is connected to the output end of the motor 18. A sealing ring 20 is fixedly connected at the junction of the top of the mounting base 12 and the sealing shell 13. The inner magnetic rotor 16 and the outer magnetic rotor 19 can be quickly installed and disassembled through the replacement mechanism 8. The stirring head 15 is installed through the installation mechanism 9.

[0026] In this embodiment: gas enters and exits the vessel 1 through the vent pipe 5, and material is injected into the vessel 1 through the feed pipe 3 and the supersonic particle nozzle 4. The material in the vessel 1 is discharged through the discharge pipe 6 by the switch valve 7. The temperature control coil 10 is used to control the temperature of the vessel 1. The motor 18 drives the outer magnetic rotor 19 to rotate. The rotation of the outer magnetic rotor 19 drives the inner magnetic rotor 16 to rotate through magnetic force. The rotation of the inner magnetic rotor 16 drives the stirring shaft 14 to rotate. The rotation of the stirring shaft 14 drives the stirring head 15 to rotate. The rotation of the stirring head 15 stirs the material in the vessel 1. The enamel lining inside the vessel 1 is used to prevent metal ion contamination. At the same time, the shear force of the stirring head 15 during stirring is ≤50 Pa·s.

[0027] Please refer to this carefully. Figures 4 to 8The replacement mechanism 8 includes through holes 801 symmetrically located on both sides of the protective shell 17. The top of the fixing base 11 has symmetrically located mounting holes 803, with bolts 802 threaded onto the inner wall of the mounting holes 803. The sealing shell 13 has symmetrically located connecting grooves 804 on both sides. The top of the mounting base 12 has symmetrically fixed connecting blocks 805. The inner wall of the connecting blocks 805 has a slidingly connected locking block 806 extending from the connecting block 805. A first spring 807 connects the locking block 806 and the connecting block 805. A push plate 808 is slidably connected inside the fixing base 11 and the mounting base 12 at the bottom of the mounting holes 803. A second spring 809 connects the push plate 808 and the fixing base 11. A T-shaped frame 810 is fixedly connected to the top of 808. The T-shaped frame 810 passes through the locking block 806. A positioning groove 818 is opened at the top of the locking block 806. A fixing groove 811 is symmetrically opened on the inner wall of the inner magnetic rotor 16. A rotating disk 812 is rotatably connected to the top of the stirring shaft 14. A threaded rod 813 is fixedly connected to the bottom of the rotating disk 812. A displacement frame 814 is slidably connected to the outer wall of the threaded rod 813. The displacement frame 814 is slidably connected to the inside of the stirring shaft 14. Fixing blocks 816 are slidably connected to both sides of the displacement frame 814 inside the stirring shaft 14. A sliding rod 815 is fixedly connected to the bottom of the displacement frame 814. A sliding groove 817 for the sliding rod 815 to slide is opened on the outer wall of the fixing block 816.

[0028] In this embodiment: after the stirring shaft 14 is installed on the mounting base 12, the inner magnetic rotor 16 is sleeved on the top end of the stirring shaft 14. Then, the rotating disk 812 is rotated. The rotation of the rotating disk 812 drives the threaded rod 813 to rotate. The rotation of the threaded rod 813 drives the displacement frame 814 to move. The displacement of the displacement frame 814 drives the slide rod 815 to move. The slide rod 815 slides in the slide groove 817 and pushes the fixing block 816 to move. The fixing block 816 is inserted into the fixing groove 811 to fix the inner magnetic rotor 16. After the inner magnetic rotor 16 is installed, the connecting block 805 is inserted into the connecting groove 804 until the locking block 806 is displaced by the elastic force of the first spring 807 and contacts the outer wall of the sealing shell 13, thus locking the sealing shell 13 onto the top of the mounting base 12. After completion, the outer magnetic rotor 19 is installed at the output end of the motor 18, and the protective shell 17 is placed on the top of the mounting base 12. The bolt 802 is passed through the through hole 801 and connected into the mounting hole 803 to complete the installation operation of the outer magnetic rotor 19 and the inner magnetic rotor 16. At the same time, during the process of the bolt 802 being connected into the mounting hole 803, the bolt 802 contacts the push plate 808, pushing the push plate 808 to move, which compresses the second spring 809. The displacement of the push plate 808 drives the T-shaped frame 810 to move, and the T-shaped frame 810 is locked into the positioning groove 818 to fix the locking block 806, preventing the locking block 806 from moving and causing the sealing shell 13 to loosen, thereby affecting the sealing performance of the sealing shell 13. When replacing the outer magnetic rotor 19 and the inner magnetic rotor 16, remove the bolt 802 from the mounting hole 803 and remove the protective shell 17. At this time, the outer magnetic rotor 19 can be replaced. Simultaneously, the bolt 802 separates from the push plate 808. The push plate 808 is reset by the elastic force of the second spring 809. The reset of the push plate 808 causes the T-shaped frame 810 to move out of the positioning groove 818, thus canceling the fixation of the locking block 806. At this time, push the locking block 806 to separate from the sealing shell 13, so that the sealing shell 13 can be disassembled. Rotate the rotating disk 812 to drive the fixing block 816 to move out of the fixing groove 811, and remove the inner magnetic rotor 16 from the stirring shaft 14. This completes the disassembly operation of the inner magnetic rotor 16, which facilitates the quick disassembly of the inner magnetic rotor 16 and the outer magnetic rotor 19, so that the inner magnetic rotor 16 and the outer magnetic rotor 19 can be quickly replaced when they are damaged.

[0029] Please refer to this carefully. Figures 5 to 10 The mounting mechanism 9 includes a groove 901, which is located at the top of the mounting base 12. A positioning disc 902 is fixedly connected to the outer wall of the stirring shaft 14 within the inner cavity of the groove 901. A positioning ring 903 is slidably connected to the inner wall of the groove 901 at the top of the positioning disc 902. A limiting seat 904 is fixedly connected to the outer wall of the stirring shaft 14 within the inner cavity of the vessel body 1. A threaded seat 905 is fixedly connected to the top of the limiting seat 904, and a rotating ring 9 is threadedly connected to the outer wall of the threaded seat 905. 06. The outer wall of the threaded seat 905 is symmetrically provided with transverse grooves 909. The outer wall of the stirring head 15 is fixedly connected with a crossbar 907. The outer wall of the crossbar 907 is provided with a slot 908. The stirring shaft 14 is slidably connected with a lower pressure frame 910. The top of the lower pressure frame 910 extends into a positioning plate 902. A third spring 911 is connected between the lower pressure frame 910 and the positioning plate 902. The outer wall of the lower pressure frame 910 is fixedly connected with an insertion rod 912. The insertion rod 912 is located above the transverse groove 909.

[0030] In this embodiment: when installing the stirring shaft 14, the stirring shaft 14 is moved through the mounting base 12 into the inner cavity of the vessel body 1, the positioning plate 902 is moved into the inner cavity of the groove 901, and then the positioning ring 903 is placed on the top of the positioning plate 902. When fixing the sealing shell 13, the sealing shell 13 contacts the positioning ring 903, thereby positioning the positioning plate 902 in the groove 901, thus completing the installation operation of the stirring shaft 14. When installing the stirring head 15, the crossbar 907 is inserted into the transverse groove 909. When the positioning ring 903 is placed at the top of the positioning plate 902, the positioning ring 903 contacts the lower pressure frame 910, pushing the lower pressure frame 910 to move and compressing the third spring 911. The displacement of the lower pressure frame 910 drives the insertion rod 912 to move, and the insertion rod 912 is inserted into the slot 908 to fix the stirring head 15, which facilitates the quick replacement of the stirring head 15. The shearing force generated when the stirring shaft 14 rotates can be adjusted by replacing the stirring head 15. When the transverse groove 909 is not in use, the rotating ring 906 can be rotated. The rotating ring 906 rotates and moves along the outer wall of the threaded seat 905. The rotating ring 906 moves and contacts the limiting seat 904, thereby blocking the transverse groove 909.

[0031] Please refer to this carefully. Figures 4 to 8 The inner wall of the connecting groove 804 fits against the outer wall of the connecting block 805, the end of the locking block 806 extending out of the connecting block 805 is provided with a bevel, and the inner wall of the positioning groove 818 fits against the top outer wall of the T-shaped frame 810.

[0032] In this embodiment: the connecting block 805 is inserted into the connecting groove 804 until the locking block 806 is displaced by the elastic force of the first spring 807 and contacts the outer wall of the sealing shell 13, thus locking the sealing shell 13 onto the top of the mounting base 12. After completion, the outer magnetic rotor 19 is installed at the output end of the motor 18, and the protective shell 17 is placed on the top of the mounting base 12. The bolt 802 is passed through the through hole 801 and connected into the mounting hole 803 to complete the installation operation of the outer magnetic rotor 19 and the inner magnetic rotor 16. At the same time, during the process of the bolt 802 being connected into the mounting hole 803, the bolt 802 contacts the push plate 808, pushing the push plate 808 to move, causing the second spring 809 to be squeezed. The displacement of the push plate 808 drives the T-shaped frame 810 to move, and the T-shaped frame 810 is locked into the positioning groove 818 to fix the locking block 806.

[0033] Please refer to this carefully. Figures 4 to 8 The top outer wall of the stirring shaft 14 is in contact with the inner wall of the inner magnetic rotor 16, the inner wall of the fixing groove 811 is in contact with the outer wall of the fixing block 816, the top of the displacement frame 814 is provided with a threaded hole, the threaded hole matches the threaded rod 813, and the outer wall of the slide rod 815 is in contact with the inner wall of the fixing block 816.

[0034] In this embodiment: the inner magnetic rotor 16 is sleeved on the top end of the stirring shaft 14, and then the rotating disk 812 is rotated. The rotation of the rotating disk 812 drives the threaded rod 813 to rotate, and the rotation of the threaded rod 813 drives the displacement frame 814 to move. The displacement of the displacement frame 814 drives the slide rod 815 to move. The slide rod 815 slides in the slide groove 817 and pushes the fixing block 816 to move. The fixing block 816 is inserted into the fixing groove 811 to fix the inner magnetic rotor 16.

[0035] Please refer to this carefully. Figures 5 to 10 The outer wall of the crossbar 907 fits against the inner wall of the cross groove 909, and the inner wall of the slot 908 fits against the outer wall of the insert rod 912.

[0036] In this embodiment: the crossbar 907 is inserted into the cross groove 909. When the positioning ring 903 is placed at the top of the positioning plate 902, the positioning ring 903 contacts the lower pressure frame 910, pushing the lower pressure frame 910 to move and compressing the third spring 911. The displacement of the lower pressure frame 910 drives the insertion rod 912 to move. The insertion rod 912 is inserted into the slot 908 to fix the stirring head 15. Example 1

[0037] Step 1: Construction of Low-Temperature Cocrystallization Nuclei: 1.0 g of SOD enzyme solution with a specific activity of 4520 U / mg (containing 10 mM PBS buffer, pH 7.0) was mixed with 10.0 g of purified trehalose and pumped into the mixture through a microfluidic chip (borosilicate glass, main channel width 205 μm). The aqueous phase flow rate was 0.8 mL / min, and the oil phase (perfluoropolyether PFPE:cyclopentane = 7:3 v / v) flow rate was 4.0 mL / min. W / O emulsion droplets were formed under a controlled temperature of 4℃. The emulsion was rapidly transferred to a -30℃ cold trap and frozen for 30 min to obtain SOD-trehalose cocrystallization nuclei with an average particle size of 2.6 μm (measured by a laser particle size analyzer).

[0038] Step 2, In-situ Phospholipid Encapsulation: The above-mentioned microparticles were dispersed in supercritical CO2 fluid containing 0.08 wt% vitamin E (pressure 8.0 MPa, temperature 16℃), and 3.75 g of lecithin and 1.25 g of cholesterol (molar ratio 3:1) were added. The mixture was magnetically stirred (300 rpm) for 45 min. A bilayer lipid film was formed on the surface of the microparticles using supercritical reverse solvent deposition (SAS) technology. After centrifugation, the encapsulation efficiency was determined to be 89.2% (HPLC method, mobile phase: acetonitrile / water = 70 / 30).

[0039] Step 3, Targeted Modification and Cold Spray Drying: 1.2 wt% hyaluronic acid solution (molecular weight 15 kDa) was added to the embedded product, and the mixture was homogenized using a high-pressure homogenizer (50 MPa, 3 cycles) to form a homogeneous suspension. The suspension was then pumped into a cold spray drying tower (inlet air temperature 32℃, atomization pressure 0.16 MPa, nitrogen atmosphere) and dried to obtain a free-flowing powder with a moisture content of 2.8% (Karl Fischer process). 0.1 wt% nano-silica (Aerosil 200) was added to aid flow.

[0040] Product performance: SOD activity recovery rate: 93.5% (pyrogallol auto-oxidation method); 24h transdermal cumulative permeation: 82.5 μg / cm³ 2 (Franz diffusion cell, ex vivo pigskin); Accelerated stability (40℃ / RH75%): ≥90% residual activity after 18 months. Example 2

[0041] Step 1: Construction of low-temperature co-crystal nuclei: 1.2 g of SOD enzyme solution with a specific activity of 4380 U / mg was mixed with 14.4 g of trehalose (mass ratio 1:12). The mixture was then passed through a microfluidic chip (channel width 198 μm) at 4 °C to form an emulsion with an aqueous phase flow rate of 1.0 mL / min and an oil phase (PFPE:cyclopentane = 7:3) flow rate of 5.0 mL / min. The emulsion was then frozen at -29 °C for 35 min to obtain co-crystal microparticles with a particle size of 2.8 μm.

[0042] Step 2, In-situ phospholipid encapsulation: The microparticles were dispersed in supercritical CO2 containing 0.10 wt% vitamin E (pressure 8.2 MPa, temperature 15℃), and 4.32 g of lecithin and 1.44 g of cholesterol (molar ratio 3:1) were added. The mixture was stirred for 42 min, and the encapsulation rate was 87.6%.

[0043] Step 3, Targeted Modification and Cold Spray Drying: Add 0.9 wt% hyaluronic acid (molecular weight 18 kDa), homogenize, and then cold spray dry (inlet air temperature 34℃, atomization pressure 0.15 MPa). Add 0.1 wt% nano-silica to obtain a powder with 3.0% moisture content. The rest is the same as in Example 1 and will not be repeated here.

[0044] Product performance: Active recovery rate: 91.3%, cumulative transdermal penetration: 79.8 μg / cm², accelerated stability: ≥89% active residue after 17 months. Example 3

[0045] Step 1: Construction of low-temperature eutectic nuclei: Same as in Example 1, with SOD dosage of 0.8g, trehalose dosage of 8.0g (1:10), and particle size of 2.4μm.

[0046] Step 2, in-situ phospholipid encapsulation: Same as Example 1, encapsulation efficiency 88.9%, determined by HPLC, mobile phase: acetonitrile / water = 70 / 30).

[0047] Step 3, Targeted Modification and Cold Spray Drying: Add 0.8 wt% hyaluronic acid (molecular weight 12 kDa), homogenize, and adjust the cold spray atomization pressure to 0.18 MPa (inlet air temperature 35℃). The powder dissolution time is ≤15 seconds (determined by laser diffraction).

[0048] The rest is the same as in Example 1, and will not be repeated here.

[0049] Product performance: Activity recovery rate: 92.7%, cumulative transdermal penetration: 76.3 μg / cm³ 2 Accelerated stability: ≥88% residual activity after 17 months.

[0050] Comparative Example 1: Traditional Liposome Method Step 1: Mix 1.0g of SOD enzyme solution with 10g of trehalose, pre-freeze at -80℃ for 2 hours, and then freeze-dry under vacuum at 25℃ for 48 hours to obtain freeze-dried powder.

[0051] Step 2: Dissolve 3.75g of lecithin and 1.25g of cholesterol in anhydrous ethanol, inject into 35℃ PBS buffer (pH 7.4), stir to form liposomes, add lyophilized powder and incubate at 40℃ for 30 min.

[0052] Step 3: After centrifugation and purification, add 1.2 wt% hyaluronic acid and spray dry (inlet air temperature 85℃).

[0053] The rest is the same as in Example 1, and will not be repeated here.

[0054] Product performance: Encapsulation efficiency: 62.1% (due to lipid membrane rupture caused by high temperature), Activity recovery rate: 67.4% (due to double damage from lyophilization and high temperature), Transdermal penetration: 25.7 μg / cm³ 2 (Liposome aggregation affects transdermal absorption).

[0055] Comparative Example 2: No eutectic protection Step 1: Simply mix 1.0g of SOD enzyme solution with 10g of trehalose and freeze directly at -30℃ for 30 minutes without undergoing microfluidic emulsification co-crystallization treatment.

[0056] Step 2: Perform supercritical encapsulation as in Example 1, with an encapsulation rate of 71.5% (due to the lack of eutectic nucleus protection, some SOD leakage occurred).

[0057] Step 3: Add hyaluronic acid and cold spray drying as in Example 1.

[0058] The rest is the same as in Example 1, and will not be repeated here.

[0059] Product performance: Activity recovery rate: 58.3% (ice crystals directly disrupt enzyme structure), transdermal penetration: 18.4 μg / cm³ 2 (Enzyme aggregation reduces penetration efficiency), accelerating stability: 6 months of residual activity <50%.

[0060] Performance testing The SOD complex preparations prepared in Examples 1-3 and Comparative Examples 1-2 were tested for enzyme activity recovery, encapsulation efficiency, transdermal cumulative penetration, accelerated stability, and reconstitution time. 1. Enzyme activity recovery rate was determined according to the pyrogallol auto-oxidation method in General Chapter 0703 of Part IV of the Chinese Pharmacopoeia 2020. 2. Encapsulation efficiency was determined by high performance liquid chromatography (HPLC) (chromatographic conditions: C18 column, acetonitrile / water = 70 / 30, flow rate 1.0 mL / min, detection wavelength 280 nm). 3. The cumulative transdermal permeation was determined according to the Franz diffusion cell method of OECD Guideline 428 (ex vivo pig skin, 24h sampling). 4. Accelerated stability was tested at 40℃ and 75% relative humidity. The half-life was calculated based on the first-order kinetic model of activity decay. 5. The reconstitution time was determined by laser diffraction according to the guidelines for microparticle formulations in Part IV, General Chapter <0991> of the 2020 edition of the Chinese Pharmacopoeia, with a D90 value ≤ 500 nm as the standard for complete dispersion. Particle size analysis was performed using a laser diffractometer (0.9% NaCl solution, complete dispersion endpoint D90 ≤ 500 nm). The results are shown in Table 1 below: Table 1 Comparison of key performance indicators of SOD compound preparations

[0061] In summary, this invention achieves a core breakthrough in the industrialization of SOD formulations through a three-stage low-temperature integrated process of microfluidic co-crystallization-supercritical in-situ embedding-cold spray drying and a three-level structural design of co-crystallization core-lipid layer-targeting shell: enzyme activity recovery rate is increased to ≥92%, which is more than 40% higher than that of traditional processes; transdermal penetration reaches 82.5 μg / cm³. 2 It improved by 221% compared to the unmodified group; the accelerated half-life was extended to 18 months, while the cumulative activity loss was reduced to ≤8% and the reconstitution time was shortened to ≤28 seconds. It systematically solved the problems of activity retention, transdermal delivery and production loss, and provided a high-activity, long-lasting and stable industrial solution for the pharmaceutical and cosmetic fields.

[0062] The above description is merely a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A SOD complex formulation consisting of a SOD-trehalose co-crystal core, a lecithin-cholesterol bilayer and a hyaluronic acid targeting shell, characterized in that, The preparation comprises the following components by mass percentage: Superoxide dismutase 0.5%~1.2%; Trehalose co-crystal protective agent 8%~12%; Egg phospholipid / cholesterol 4.5%~6.0%; Hyaluronic acid 0.8%~1.5%; The balance is a phosphate buffer system, pH 6.8±0.

2.

2. A process for the preparation of SOD complex formulation characterized in that, The method comprises the following steps: Step one, mix SOD enzyme liquid with specific activity not less than 4000 U / mg and trehalose at a mass ratio of 1:8~1:12, pump into a microfluidic chip with a channel width of 180~220 μm, form W / O emulsion under the conditions of temperature 2 to 5℃, water phase to oil phase flow rate ratio 1:5, freeze at-28~-32℃ for 30 minutes, form co-crystal core microparticles with particle size 2.2~2.8 μm; Step two, in-situ phospholipid embedding: pump the co-crystal core microparticles prepared in step one into a supercritical deposition reactor, disperse in supercritical CO2 fluid containing egg phospholipid and cholesterol, wherein the molar ratio of egg phospholipid to cholesterol is 3:1, the operating pressure is 7.5~8.5 MPa, the operating temperature is 13~17℃, the reaction is stirred by a magnetic stirring system at a rate of 250~350 rpm for 35~45 minutes, and a double-layer lipid envelope is formed by using supercritical anti-solvent deposition technology; Step three, targeted modification and cold spray drying: add 0.8%~1.5% hyaluronic acid solution to the product of step two, pass through 3 cycles of high-pressure homogenization at a pressure of 50 MPa, pump into a cold spray drying tower, dry under the conditions of air inlet temperature not exceeding 35℃, atomization pressure 0.13~0.17 MPa, control the moisture content to not more than 3.0%, and obtain a flowable powder, i.e. SOD composite preparation.

3. The process for the preparation of SOD complex formulation as claimed in claim 2 wherein, The oil phase of the microfluidic chip in step one is a mixture of perfluoropolyether and cyclopentane at a volume ratio of 7:3; The supercritical CO2 fluid in step two contains 0.05% to 0.1% vitamin E by mass percentage; The operating pressure in step two is 8.0 MPa, the operating temperature is 15℃, the magnetic stirring rate is 300 rpm, and the reaction time is 40 minutes; The dry powder obtained in step three is added with 0.1% nano-silicon dioxide flow aid by mass percentage in a double-cone vacuum mixer, based on the total mass of the powder.

4. The process for the preparation of SOD complex formulation as claimed in claim 2 wherein, The supercritical deposition reaction kettle, including kettle body (1), the top of kettle body (1) is provided with kettle cover (2), the both sides of the top of kettle cover (2) are respectively fixedly connected with feed pipe (3) and air pipe (5), the bottom of feed pipe (3) is installed with supersonic particle nozzle (4), the inner wall of kettle body (1) is provided with enamel lining, the bottom of kettle body (1) is fixedly connected with discharge pipe (6), the outer wall of discharge pipe (6) is installed with valve (7), the outer wall of kettle body (1) is fixedly connected with temperature control coil pipe (10), the top of kettle cover (2) is fixedly connected with fixed base (11), the inner wall of fixed base (11) is fixedly connected with mounting seat (12), the top of mounting seat (12) is installed with sealing shell (13), mounting seat (12) is rotatably connected with stirring shaft (14), the outer wall of stirring shaft (14) is installed with stirring head (15), the top of stirring shaft (14) is installed with inner magnetic rotor (16), the top of fixed base (11) is installed with protection shell (17), the top of protection shell (17) is installed with motor (18), the output end of motor (18) is connected with outer magnetic rotor (19), the top of mounting seat (12) is fixedly connected with sealing ring (20) at the position of sealing shell (13), inner magnetic rotor (16) and outer magnetic rotor (19) are quickly installed and disassembled by changing mechanism (8), and stirring head (15) is installed by mounting mechanism (9).

5. The process for the preparation of SOD complex formulation as claimed in claim 4 wherein, The replacement mechanism (8) includes a through hole (801) symmetrically arranged on both sides of the protective shell (17), a mounting hole (803) symmetrically arranged on the top end of the fixing seat (11), a bolt (802) threadedly connected to the inner wall of the mounting hole (803), a connecting groove (804) symmetrically arranged on both sides of the sealing shell (13), a connecting block (805) fixedly connected to the top end of the mounting seat (12), a clamping block (806) slidingly connected to the inner wall of the connecting block (805) and extending out of the connecting block (805), a first spring (807) connected between the clamping block (806) and the connecting block (805), a push plate (808) slidingly connected to the bottom end of the mounting hole (803) in the interiors of the fixing seat (11) and the mounting seat (12), a second spring (809) connected between the push plate (808) and the fixing seat (11), a T-shaped frame (810) fixedly connected to the top end of the push plate (808), the T-shaped frame (810) penetrating through the clamping block (806), a positioning groove (818) arranged on the top end of the clamping block (806), a fixing groove (811) symmetrically arranged on the inner wall of the inner magnetic rotor (16), a rotating disc (812) rotatably connected to the top end of the stirring shaft (14), a threaded rod (813) fixedly connected to the bottom end of the rotating disc (812), a displacement frame (814) slidingly connected to the outer wall of the threaded rod (813), the displacement frame (814) slidingly connected to the interior of the stirring shaft (14), a fixed block (816) slidingly connected to the interior of the stirring shaft (14) on both sides of the displacement frame (814), a sliding rod (815) fixedly connected to the bottom of the displacement frame (814), and a sliding groove (817) arranged on the outer wall of the fixed block (816) and allowing the sliding rod (815) to slide.

6. The process for the preparation of SOD complex formulation as claimed in claim 5 wherein, The mounting mechanism (9) comprises a recess (901) which is arranged at the top end of the mounting base (12), the outer wall of the stirring shaft (14) is fixedly connected with a positioning disc (902) in the inner cavity of the recess (901), the inner wall of the recess (901) is slidably connected with a positioning ring (903) at the top end of the positioning disc (902), the outer wall of the stirring shaft (14) is fixedly connected with a limiting seat (904) in the inner cavity of the kettle body (1), the top end of the limiting seat (904) is fixedly connected with a threaded seat (905), the outer wall of the threaded seat (905) is threadedly connected with a rotating ring (906), the outer wall of the threaded seat (905) is symmetrically provided with a horizontal groove (909), the outer wall of the stirring head (15) is fixedly connected with a horizontal rod (907), the outer wall of the horizontal rod (907) is provided with an insertion slot (908), the inner portion of the stirring shaft (14) is slidably connected with a pressing frame (910), the top end of the pressing frame (910) extends out of the positioning disc (902), the pressing frame (910) and the positioning disc (902) are connected with a third spring (911), the outer wall of the pressing frame (910) is fixedly connected with an insertion rod (912), and the insertion rod (912) is located above the horizontal groove (909).

7. The process for the preparation of SOD complex formulation as claimed in claim 5 wherein, The inner wall of the connecting groove (804) is attached to the outer wall of the connecting block (805), one end of the clamping block (806) extending out of the connecting block (805) is provided with an inclined surface, and the inner wall of the positioning groove (818) is attached to the top outer wall of the T-shaped frame (810).

8. The process for the preparation of SOD complex formulation as claimed in claim 5 wherein, The top outer wall of the stirring shaft (14) is attached to the inner wall of the inner magnetic rotor (16), and the inner wall of the fixing groove (811) is attached to the outer wall of the fixing block (816).

9. The process for the preparation of SOD complex formulation as claimed in claim 5 wherein, The top end of the displacement frame (814) is provided with a threaded hole which is matched with the threaded rod (813), and the outer wall of the sliding rod (815) is attached to the inner wall of the fixing block (816).

10. The process for the preparation of SOD complex formulation as claimed in claim 6 wherein, The outer wall of the horizontal rod (907) is attached to the inner wall of the horizontal groove (909), and the inner wall of the insertion slot (908) is attached to the outer wall of the insertion rod (912).

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