Preparation method of influenza virus-targeted double-flower drop micro-nano micelle delivery body

Abstract

The invention discloses a preparation method of a double-flower drop micro-nano micelle delivery body for targeting influenza virus, and relates to the technical field of pharmaceutical preparations, a double-flower extract is subjected to hydroformylation and is coupled with a synthesized pH-sensitive hydrazide polymer carrier to form a hydrazone bond drug-loaded precursor, and then the hydrazone bond drug-loaded precursor and neuraminidase responsive protective layer molecules are co-assembled to form the double-flower drop micro-nano micelle delivery body for targeting influenza virus. The preparation method disclosed by the invention has the advantages that a drug-loading polymer self-assembly precursor with a hydrazone bond structure is constructed by adopting a pH sensitive coupling process of an aldehyde extract precursor and a hydrazide-based polymer carrier, so that intelligent and accurate drug release under an influenza virus infection subacid environment is realized; the problems of low bioavailability and large toxic and side effects caused by insufficient targeting and uncontrollable drug release of a traditional nano drug delivery system are effectively solved.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, specifically to a method for preparing a double-flower drop micro / nano micelle delivery system targeting influenza virus. Background Technology

[0002] Honeysuckle and chrysanthemum, as traditional Chinese medicines, have the effects of clearing heat and detoxifying, and broad-spectrum antiviral activity. They are commonly used drugs for treating influenza. However, existing anti-influenza preparations containing honeysuckle and chrysanthemum, especially drops administered through nasal mucosa, still have the following prominent problems and defects: First, the active ingredients obtained by traditional extraction processes have low purity and poor chemical stability, and are easily degraded and inactivated during storage and in vivo delivery, resulting in poor actual efficacy of the preparations. Second, conventional nano-drug delivery systems are mostly passively targeted, lacking the ability to specifically recognize the site of influenza virus infection, and the drug release is uncontrollable, posing risks of low targeting accuracy, low bioavailability, and potential toxic side effects. Third, existing drop formulations have a short residence time in the nasal mucosa and are easily cleared, and the drug has difficulty effectively penetrating the mucosal barrier to reach infected cells, limiting its preventive and therapeutic effects. In addition, existing delivery systems mostly adopt a single response mechanism and cannot synergistically utilize multiple characteristic signals of the viral infection microenvironment, resulting in insufficient intelligence and precision. Therefore, we propose a method for preparing a dual-flower drop micro / nano micelle delivery system targeting influenza virus. Summary of the Invention

[0003] The purpose of this invention is to provide a method for preparing a double-flower drop micro / nano micelle delivery system targeting influenza virus.

[0004] To achieve the above objectives, the present invention provides the following technical solution: a method for preparing a dual-flower droplet micro / nano micelle delivery system targeting influenza virus, comprising a delivery system preparation method, wherein the delivery system preparation method includes the following steps: Step 1: The honeysuckle and chrysanthemum mixture was extracted with an ethanol-water solution using ultrasound-assisted extraction. The purified extract was then reacted with glutaraldehyde under alkaline conditions to generate an extract precursor with an aldehyde group. Step 2: Using polyethylene glycol monomethyl ether as an initiator, an amphiphilic polymer carrier containing pH-sensitive hydrazone bonds in the main chain / side chain and acylhydrazine groups in the terminal / side chain is synthesized by ring-opening polymerization and chemical succession method. Step 3: React the aldehyde-containing extract precursor with an amphiphilic polymer carrier containing an acylhydrazine group in an acidic buffer solution. The condensation of the aldehyde and acylhydrazine groups forms a hydrazone bond, yielding a drug-loaded polymer self-assembly precursor. Step 4: React polyethylene glycol with highly reactive terminal groups with a diamine compound containing disulfide bonds, and then link it with the cleavable peptide chain of neuraminidase to obtain a neuraminidase-responsive protective layer molecule. Step 5: Dissolve the drug-loaded polymer self-assembly precursor and the neuraminidase-responsive protective layer molecule together in a buffer solution, form micelles through self-assembly, and purify them by filtration and ultrafiltration to obtain a micelle concentrate. Step 6: Mix the micelle concentrate with the mucosal adhesive, antibacterial agent and pH adjuster in an isotonic buffer solution, adjust to the required concentration and pH value, and aseptically fill.

[0005] As a further aspect of the present invention: in step one, the mass ratio of honeysuckle to chrysanthemum is 1-3:1, and the reaction under alkaline conditions is specifically carried out at pH 8.0-9.0 and 35℃-45℃ for 2-4 hours.

[0006] As a further aspect of the present invention: in step two, the number average molecular weight of the amphiphilic polymer carrier is 10kDa-25kDa, and the pH-sensitive hydrazone bond breaks in an environment of pH < 5.5.

[0007] As a further aspect of the present invention: the acidic buffer solution is an acetate buffer solution with a pH of 4.5-5.5, the molar ratio of the aldehyde-containing extract precursor to the hydrazide-containing polymer carrier is 1:1.5 to 1:2.5, and the reaction time is 12h-24h.

[0008] As a further aspect of the present invention: in step four, the amino acid sequence of the cleavable peptide chain of the neuraminidase is Sialic acid-Gly-Val-NH2, and the highly reactive group is 4-(trifluoromethyl)phenol ester group.

[0009] As a further aspect of the present invention: in step five, the mass ratio of the drug-loaded polymer self-assembly precursor to the neuraminidase-responsive protective layer molecule is 10:1 to 5:1, and the self-assembly is carried out at 45℃-55℃ for 2h-4h, resulting in micelles with a particle size of 80nm-150nm and an encapsulation efficiency greater than 95%.

[0010] As a further aspect of the present invention: in step six, the mucosal adhesive is a chitosan derivative, accounting for 0.5%-1.5% by mass in the formulation; the antibacterial agent is benzalkonium chloride, accounting for 0.01%-0.05% by mass; and the final pH value of the drops is adjusted to 6.0-7.0.

[0011] Compared with the prior art, the beneficial effects of the present invention by adopting the above technical solution are as follows: 1. This invention constructs a drug-loaded polymer self-assembly precursor with a hydrazone bond structure by employing a pH-sensitive coupling process between an aldehyde-based extract precursor and an acylhydrazine-based polymer carrier. This enables intelligent and precise drug release in the slightly acidic environment of influenza virus infection, effectively overcoming the problems of insufficient targeting, uncontrollable drug release, low bioavailability, and high toxicity caused by traditional nano-drug delivery systems. 2. This invention introduces a neuraminidase-responsive protective layer molecule and constructs enzyme-pH sequentially activated dual-targeting micelles together with a drug-loaded polymer self-assembly precursor. This structure can not only specifically recognize and penetrate the cell membrane at the site of viral infection, but also significantly enhance the residence time and permeability of the micelles in the nasal mucosa, solving the problem of limited efficacy caused by the strong mucosal clearance and poor permeability of conventional drops. 3. This invention optimizes the formulation and preparation process of micro-nano micelles, and uses a specific mucosal adhesive, antibacterial agent and isotonic buffer system for droplet formation, ensuring that the formulation has good biocompatibility, physical stability and sterility. At the same time, the delivery system exhibits a highly efficient inhibitory effect on influenza virus, achieving a high degree of unity between formulation stability and therapeutic efficacy. Attached Figure Description

[0012] Figure 1 This is a schematic diagram of the delivery body preparation process in an embodiment of the present invention. Detailed Implementation

[0013] The specific embodiments of the present invention will be further described below with reference to the accompanying drawings. It should be noted that the description of these embodiments is for the purpose of helping to understand the present invention, but does not constitute a limitation of the present invention.

[0014] Furthermore, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0015] Please see the appendix Figure 1 This invention discloses a method for preparing a dual-flower droplet micro / nano micelle delivery system targeting influenza virus, comprising the following steps: Step 1: The honeysuckle and chrysanthemum mixture was extracted with an ethanol-water solution using ultrasound-assisted extraction. The purified extract was then reacted with glutaraldehyde under alkaline conditions to generate an extract precursor with an aldehyde group. Step 2: Using polyethylene glycol monomethyl ether as an initiator, an amphiphilic polymer carrier containing pH-sensitive hydrazone bonds in the main chain / side chain and acylhydrazine groups in the terminal / side chain is synthesized by ring-opening polymerization and chemical succession method. Step 3: React the aldehyde-containing extract precursor with an amphiphilic polymer carrier containing an acylhydrazine group in an acidic buffer solution. The condensation of the aldehyde and acylhydrazine groups forms a hydrazone bond, yielding a drug-loaded polymer self-assembly precursor. Step 4: React polyethylene glycol with highly reactive terminal groups with a diamine compound containing disulfide bonds, and then link it with the cleavable peptide chain of neuraminidase to obtain a neuraminidase-responsive protective layer molecule. Step 5: Dissolve the drug-loaded polymer self-assembly precursor and the neuraminidase-responsive protective layer molecule together in a buffer solution, form micelles through self-assembly, and purify them by filtration and ultrafiltration to obtain a micelle concentrate. Step 6: Mix the micelle concentrate with the mucosal adhesive, antibacterial agent and pH adjuster in an isotonic buffer solution, adjust to the required concentration and pH value, and aseptically fill.

[0016] In one embodiment of the present invention: In step one, the mass ratio of honeysuckle to chrysanthemum is 1-3:1, the volume concentration of ethanol used in ultrasonic-assisted extraction is 60%-75%, the material-liquid ratio is 1:10-1:15, the ultrasonic power is 300W-500W, the extraction temperature is 50℃-65℃, the extraction time is 30min-60min, the purification uses AB-8 type macroporous adsorption resin, the eluent is 30%-50% ethanol, in the reaction to generate the aldehyde extract precursor, the mass-volume ratio of glutaraldehyde to extract is 1:5-1:15, and the reaction is carried out at pH 8.0-9.0 and 35℃-45℃ for 2h-4h.

[0017] In one embodiment of the present invention: in step two, the monomer for ring-opening polymerization is benzyl aspartate N-carboxylic acid anhydride, and the chemical ligation method is to use hydrazine hydrate for hydrazinolysis to introduce acylhydrazine groups. The reaction is carried out at room temperature for 24-48 hours. The number average molecular weight of the affinity polymer carrier is 10kDa-25kDa, and the pH-sensitive hydrazone bond is broken in an environment of pH < 5.5.

[0018] In one embodiment of the present invention: in step three, the acidic buffer solution is an acetate-sodium acetate buffer system of 0.01M-0.05M and pH 4.5-5.5, the molar ratio of the aldehyde-containing extract precursor to the hydrazide-containing polymer carrier is 1:1.5 to 1:2.5, and the reaction is carried out for 12-24 hours under light-protected and inert gas protection. In the drug-loaded polymer self-assembly precursor, the drug loading of honeysuckle extract is 10%-25%.

[0019] In one embodiment of the present invention: in step four, the highly reactive group is 4-(trifluoromethyl)phenol ester (TFP ester), the disulfide-containing diamine compound is cystamine or its hydrochloride, the amino acid sequence of the cleavable peptide chain of neuraminidase is Sialic acid-Gly-Val-NH2, and the reaction is carried out in a phosphate buffer at pH 7.0-8.0 for 4-8 hours.

[0020] In one embodiment of the present invention: In step five, the mass ratio of the drug-loaded polymer self-assembly precursor to the neuraminidase-responsive protective layer molecule is 10:1 to 5:1, the buffer solution is phosphate-buffered saline with pH 7.2-7.6, the self-assembly process is carried out in a water bath at 45℃-55℃ with stirring at 200rpm-400rpm for 2h-4h, the filter membrane filtration uses 0.45μm and 0.22μm microporous membranes in sequence, and the ultrafiltration purification uses an ultrafiltration tube with a molecular weight cutoff of 50kDa-100kDa or a tangential flow ultrafiltration system, the resulting micelles have a particle size of 80nm-150nm, a polydispersity index of less than 0.25, and an encapsulation efficiency of greater than 95%.

[0021] In one embodiment of the present invention: in step six, the mucosal adhesive is at least one of carboxymethyl chitosan and sodium hyaluronate, accounting for 0.5%-1.5% by mass in the final drops; the isotonic buffer is physiological saline or phosphate buffer with pH 6.5-7.5; the antibacterial agent is benzalkonium chloride, accounting for 0.01%-0.05% by mass in the final drops; the final pH value of the drops is adjusted to 6.0-7.0; and the final concentration of micro / nano micelles in the drops is 5 mg / mL-20 mg / mL.

[0022] Example 1, please refer to the appendix. Figure 1 Preparation of honeysuckle and chrysanthemum extract: Take 150g of honeysuckle and chrysanthemum powder (mass ratio 2:1), add 1800mL of 70% ethanol aqueous solution, and extract with ultrasonic assistance (power 400W, temperature 60℃) for 50min. After filtration, concentrate the filtrate under reduced pressure and purify it with AB-8 macroporous adsorption resin (elution with 30% ethanol) to obtain refined honeysuckle extract.

[0023] Functional modification: Take 10g of the above purified extract and mix it with 1.0mL of 10% glutaraldehyde aqueous solution. React at pH 8.5 and 40℃ for 3h. After the reaction is completed, dialysis is used to purify the product and obtain the aldehyde-modified honeysuckle extract precursor.

[0024] Using mPEG-NH2 as an initiator, the ring-opening polymerization of BLA-NCA was initiated under anhydrous conditions to obtain mPEG-b-PBLA.

[0025] mPEG-b-PBLA was reacted with excess hydrazine hydrate at room temperature for 36 h to undergo hydrazinolysis, yielding a pH-sensitive polymer carrier PEG-bP with hydrazide groups on the side chain. Its number-average molecular weight was determined to be 18 kDa by GPC.

[0026] The aldehyde-modified honeysuckle extract precursor and PEG-bP polymer carrier were fed at a molar ratio of aldehyde to hydrazide of 1:2. The mixture was reacted in 0.02M acetate buffer at pH 5.0 under light protection and nitrogen protection for 18 hours. After ultrafiltration purification, the drug loading of the honeysuckle extract was measured to be 18.5% (w / w).

[0027] The TFP-PEG-TFP intermediate was synthesized and then reacted with cystamine dihydrochloride to obtain PEG-SS-PEG. Finally, it was reacted with a neuraminidase-cleavable peptide chain in PBS at pH 7.4 for 6 h, and the target protective layer molecule was obtained after purification.

[0028] The drug-loaded polymer self-assembly precursor and the neuraminidase-responsive protective layer molecule were mixed at a mass ratio of 8:1, dissolved in PBS at pH 7.4, and self-assembled for 3 hours in a 50°C water bath with stirring at 300 rpm.

[0029] The solution was filtered sequentially through 0.45 μm and 0.22 μm microporous membranes, and then concentrated and replaced using an ultrafiltration tube with a molecular weight cutoff of 100 kDa to obtain a micelle concentrate.

[0030] Prescription: 10 mg / mL micelle concentrate, 1.0% carboxymethyl chitosan, 0.02% benzalkonium chloride, make up to 100% with physiological saline, and adjust the pH to 6.5 with dilute NaOH solution.

[0031] The product is filled under aseptic conditions to obtain the Double Flower Drops targeting the influenza virus.

[0032] Performance testing: Particle size and Zeta potential: The average particle size measured by the laser particle size analyzer was 95 nm, the PDI was 0.16, and the Zeta potential was -28.5 mV.

[0033] Encapsulation efficiency: The encapsulation efficiency was determined by ultrafiltration centrifugation-HPLC method, and was 96.8%.

[0034] In vitro release rate: Using the Franz diffusion cell method, the drug was released for 4 h in PBS at pH 7.4 to simulate a circulatory system, and then released for 20 h in PBS at pH 5.0 containing 0.1 U / mL neuraminidase to simulate a virus-infected cell environment. The results showed that the cumulative release rate was <15% at pH 7.4, and after switching to the simulated infection environment, the total cumulative release rate reached 90.2% in 24 h, demonstrating significant "dual-key" triggered drug release characteristics.

[0035] Antibacterial activity: The half-maximal effective concentration against influenza A virus is 0.045 mg / mL.

[0036] Stability: After being stored at 40℃±2℃ and RH75%±5% for 3 months, there were no significant changes in the appearance, particle size, encapsulation efficiency and zeta potential of the formulation.

[0037] Example 2, please refer to the appendix. Figure 1 Preparation of honeysuckle and chrysanthemum extract: Take 150g of honeysuckle and chrysanthemum powder (mass ratio 1:1), add 1950mL of 65% ethanol aqueous solution, and extract with ultrasonic assistance (power 350W, temperature 55℃) for 60min. The subsequent purification steps are the same as in Example 1.

[0038] Functional modification: Take 10g of the purified extract and mix it with 1.2mL of 10% glutaraldehyde aqueous solution. React at pH 9.0 and 35℃ for 4h. Subsequent treatment is the same as in Example 1.

[0039] The synthesis method was the same as in Example 1, and the number-average molecular weight of the polymer carrier obtained by GPC was 22 kDa.

[0040] The molar ratio of the feed was 1:2.2, and the reaction was carried out in acetate buffer at pH 4.8 for 20 h. The drug loading of the honeysuckle extract was measured to be 22.1%.

[0041] The TFP-PEG-TFP intermediate was synthesized and then reacted with cystamine dihydrochloride to obtain PEG-SS-PEG. Finally, it was reacted with a neuraminidase-cleavable peptide chain in PBS at pH 7.4 for 6 h, and the target protective layer molecule was obtained after purification.

[0042] The drug-loaded polymer self-assembly precursor to protective layer had a molecular weight ratio of 6:1, a self-assembly temperature of 48℃, and a time of 3.5h. The mixture was filtered sequentially through 0.45μm and 0.22μm microporous membranes, and then concentrated and replaced using an ultrafiltration tube with a molecular weight cutoff of 100kDa to obtain a micelle concentrate.

[0043] Prescription: Micellar concentrate (based on drug-loaded polymer) 15 mg / mL, sodium hyaluronate 0.8%, benzalkonium chloride 0.03%, to be adjusted to 100% with phosphate buffer at pH 6.8.

[0044] Performance testing: Particle size and Zeta potential: The average particle size is 112 nm, the PDI is 0.19, and the Zeta potential is -25.8 mV.

[0045] Encapsulation rate: 95.3%.

[0046] In vitro release rate: In a simulated infection environment, the total cumulative release rate over 24 hours was 87.6%.

[0047] Antibacterial activity: EC5 against influenza A virus 50 It is 0.052 mg / mL.

[0048] Stability: After being placed under accelerated testing conditions for 3 months, all indicators remained stable.

[0049] Comparative Example Preparation of active ingredients: The traditional water extraction and alcohol precipitation method was adopted: 150g of honeysuckle and chrysanthemum powder were taken in a mass ratio of 2:1, decocted twice with water, 1.5h each time, the filtrates were combined, concentrated and ethanol was added until the alcohol concentration reached 70%, let stand overnight, filtered, ethanol was recovered, and spray dried to obtain crude extract of honeysuckle and chrysanthemum.

[0050] Preparation of ordinary micelles: Ordinary micelles were prepared by thin-film dispersion without the use of functionalization modification and pH-sensitive carrier: Pranic F127 and crude extract were weighed, dissolved in ethanol, the ethanol was removed by rotary evaporation to form a thin film, and PBS at pH 7.4 was added for hydration to obtain ordinary drug-loaded micelles.

[0051] Drops preparation: Prescription: 10 mg / mL of ordinary micelle solution (based on carrier), 1.0% hydroxyethyl cellulose as thickener, 0.1% ethylparaben, to be made up to 100% with physiological saline, and pH adjusted to 6.5.

[0052] Performance testing: Particle size and Zeta potential: The average particle size is 185 nm, the PDI is 0.35, and the Zeta potential is -12.3 mV.

[0053] Encapsulation efficiency: only 58.4% due to lack of chemical coupling.

[0054] In vitro release rate: Rapid release in PBS at pH 7.4, with a cumulative release rate of 78.5% over 24 hours. However, in a simulated viral infection environment, the release behavior was not significantly different from that at pH 7.4, and no intelligent drug release behavior was observed. The total release rate was comparable to that of the example, but early leakage was severe.

[0055] Antibacterial activity: EC56 against influenza A virus 50 The concentration was 0.15 mg / mL, and the activity was significantly lower than that in the embodiments of the present invention.

[0056] Stability: After being placed at 40℃±2℃ for 1 month, the particle size increased significantly, PDI>0.4, precipitation occurred, and the encapsulation efficiency decreased to about 45%.

[0057] The comparative examples, using traditional extraction processes and ordinary micelle carriers, exhibited low purity of active ingredients, poor micelle encapsulation efficiency, and insufficient stability. Furthermore, they completely lacked targeted recognition and intelligent drug release functions, resulting in in vitro antiviral activity far lower than that of the embodiments of this invention. This fully demonstrates that the drops prepared by this invention through the core technical route of "aldehyde modification-pH-sensitive coupling-neuraminidase response protection" have achieved significant improvements over existing technologies in terms of encapsulation efficiency, stability, intelligent targeted drug release, and enhanced antiviral effects, thus verifying the inventiveness and excellent beneficial effects of this invention.

[0058] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make possible changes and modifications without departing from the spirit and scope of the present invention. Therefore, any modifications, equivalent changes and alterations made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention shall fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing a dual flower droplet micellar delivery body targeting influenza virus, comprising a delivery body preparation method, characterized in that, The delivery body preparation method comprises the following steps: Step one, ultrasonic-assisted extraction of honeysuckle and chrysanthemum mixed medicinal materials with ethanol aqueous solution, and then purified to obtain refined extract, and then the extract is reacted with glutaraldehyde under alkaline conditions to generate an extract precursor with aldehyde groups; Step two, using polyethylene glycol monomethyl ether as an initiator, an amphiphilic polymer carrier with a pH-sensitive hydrazone bond in the main chain / side chain and a hydrazide group at the end / side chain is synthesized by ring-opening polymerization and chemical splicing; Step three, reacting the extract precursor with aldehyde groups with the amphiphilic polymer carrier with hydrazide groups in an acidic buffer solution to form a hydrazone bond through condensation of aldehyde groups and hydrazide groups, to obtain a drug-loaded polymer self-assembly precursor; Step four, reacting polyethylene glycol modified with a highly reactive group at the end with a bis-amine compound containing a disulfide bond, and then connecting it with a sialidase-cleavable peptide chain to obtain a sialidase-responsive protective layer molecule; Step five, dissolving the drug-loaded polymer self-assembly precursor and the sialidase-responsive protective layer molecule in a buffer solution to form micelles through self-assembly, and then filtering and ultrafiltrating to obtain a micelle concentrate; Step six, mixing the micelle concentrate with a mucosal adhesive, a bacteriostatic agent, and a pH regulator in an isotonic buffer solution, adjusting to the required concentration and pH value, and sterilely filling.

2. A method of preparing a dual flower droplet micellar delivery vehicle targeting influenza virus according to claim 1, characterized by: In step one, the mass ratio of honeysuckle to chrysanthemum is 1-3:1, and the reaction under alkaline conditions is specifically carried out at pH 8.0-9.0 and 35-45°C for 2-4h.

3. A method of preparing a dual flower drop micellar delivery vehicle targeting influenza virus as claimed in claim 1, wherein: In step two, the number average molecular weight of the amphiphilic polymer carrier is 10-25kDa, and the pH-sensitive hydrazone bond breaks at pH<5.

5.

4. A method of preparing a dual flower drop micellar delivery vehicle targeting influenza virus as claimed in claim 1, wherein: In step three, the acidic buffer solution is an acetic acid buffer solution with pH 4.5-5.5, the molar ratio of the extract precursor with aldehyde groups to the polymer carrier with hydrazide groups is 1:1.5 to 1:2.5, and the reaction time is 12-24h.

5. A method of preparing a dual flower drop micellar delivery vehicle targeting influenza virus as claimed in claim 1, wherein: In step four, the amino acid sequence of the sialidase-cleavable peptide chain is Sialic acid-Gly-Val-NH2, and the highly reactive group is 4-(trifluoromethyl) phenol ester group.

6. A method of preparing a dual flower drop micellar delivery vehicle targeting influenza virus as claimed in claim 1, wherein: In step five, the mass ratio of the drug-loaded polymer self-assembly precursor to the sialidase-responsive protective layer molecule is 10:1 to 5:1, the self-assembly is carried out at 45-55°C for 2-4h, the particle size of the obtained micelles is 80-150nm, and the encapsulation efficiency is greater than 95%.

7. A method of preparing a dual flower drop micellar delivery vehicle targeting influenza virus as claimed in claim 1, wherein: In step six, the mucosal adhesive is a chitosan derivative, the mass ratio in the preparation is 0.5%-1.5%, the bacteriostatic agent is benzalkonium chloride, the mass ratio is 0.01%-0.05%, and the final pH value of the drops is adjusted to 6.0-7.0.

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