Nasal mucosa administration preparation and preparation method thereof

Abstract

The invention relates to a nasal mucosa administration preparation and a preparation method thereof. The nasal mucosa administration preparation comprises powder, and the powder is prepared from the following raw materials: exosome with the particle number of 20-5 billion, 2-10 wt% of a freeze-drying protective agent, 0.02-0.1 wt% of a pH regulator, 0.02-0.5 wt% of a surfactant and the balance of water; the powder is obtained after drying treatment, and the water content is smaller than or equal to 5%. According to the nasal mucosa drug delivery preparation, by adjusting ingredients of the auxiliary materials, a product capable of forming gel and resisting inflammation durably or a product capable of penetrating through nasal mucosa to achieve brain-targeted delivery through the nose can be obtained respectively. The gel preparation has anti-inflammatory, regeneration and repair functions, so that an anti-nasal allergy effect is locally generated. The osmotic preparation can achieve the effects of penetrating through the blood-brain barrier, promoting damaged tissue repair, promoting blood vessel growth, promoting proliferation and regeneration and the like, and can improve central diseases such as ischemic stroke, traumatic brain injury, spinal cord injury, Parkinson's disease, gradual freezing disease, autism and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of medical preparations, and in particular to a nasal mucosal drug delivery preparation and a preparation method thereof. Background Art

[0002] The nasal cavity is anatomically located between the skull base and the oral cavity, extending from the anterior nares to the narrow cavity of the nasopharynx. The nasal cavity is composed of bone and cartilage, lined with mucous membranes. The nasal septum divides it into two halves. Each nasal cavity is divided into the nasal vestibule and the proper nasal cavity by the nasal threshold.

[0003] The nasal cavity is a vital channel for the human body to communicate with the outside world. Inhalation of specific antigens such as dust, pollen, mites, bacteria, fungi, and animal fur can trigger an abnormal localized inflammatory response in the nasal mucosa, clinically known as allergic rhinitis. Typical symptoms include sneezing, watery nasal discharge, nasal congestion, and itching. Some patients also experience decreased sense of smell, and other complications such as asthma and sinusitis may occur, severely impacting patients' quality of life.

[0004] Allergic rhinitis currently has no complete cure. Topical or oral glucocorticoids, nasal decongestants, and antihistamines can effectively relieve symptoms. However, long-term use of these medications (for more than one week) can easily lead to atrophy of the nasal mucosa and weakened ciliary motility, which reduces the nasal cavity's ability to clear bacteria, increasing the risk of infection and even causing nosebleeds and drug-induced rhinitis. Long-term use of nasal glucocorticoids can also cause hormonal reactions and even lead to dependence.

[0005] Therefore, it is a technical problem that needs to be solved urgently in this field to prepare a local nasal mucosa repair preparation that can effectively prevent and improve the discomfort symptoms caused by nasal allergies and does not contain drugs with relatively large side effects. Summary of the Invention

[0006] The present invention aims to disclose a nasal mucosal administration preparation and a preparation method thereof, so as to solve one or more technical problems existing in the prior art and provide at least one beneficial option or create conditions.

[0007] To achieve the above object, the present invention is implemented through the following technical solutions: The first aspect of the present invention is to provide a preparation for nasal mucosal administration.

[0008] The second aspect of the present invention is to provide a method for preparing the nasal mucosal administration preparation according to the first aspect of the present invention.

[0009] The nasal mucosal administration preparation described in the first aspect of the present invention includes a powder, the raw materials of which include: 2 to 5 billion exosomes, 2 to 10 wt% of a lyoprotectant, 0.02 to 0.1 wt% of a pH adjuster, 0.02 to 0.5 wt% of a surfactant, and the balance is water; the powder is obtained after drying, and the moisture content is ≤5%.

[0010] Exosomes are extracellular vesicles with a diameter of 50-200 nm and a lipid bilayer membrane. They are released outside the cell and play a vital role in intercellular communication. The nasal mucosal formulation contains a large number of exosomes (2-5 billion particles), which can exert regulatory effects through the horizontal transfer of proteins, mRNA, and regulatory microRNAs, inhibiting inflammation and promoting local repair of the nasal mucosa. The surfactant can control the surface tension of the solution within an appropriate range. Under the protection of the lyoprotectant and surfactant, the freeze-dried exosomes can be stored for a long time and easily reconstituted without hydrolysis or other adverse reactions.

[0011] In some embodiments of the first aspect of the present invention, the exosomes are derived from umbilical cord-derived mesenchymal stem cells and / or adipose-derived mesenchymal stem cells.

[0012] In some embodiments of the first aspect of the present invention, the lyoprotectant is selected from at least one of trehalose, mannitol, or glycine.

[0013] In some embodiments of the first aspect of the present invention, the pH adjuster is phosphate buffer with a pH of 6.0 to 8.0, wherein the main components of the phosphate buffer are disodium hydrogen phosphate dihydrate and sodium dihydrogen phosphate dihydrate.

[0014] In some embodiments of the first aspect of the present invention, the surfactant is selected from poloxamer, Tween 80 or dodecyl maltoside.

[0015] In some embodiments of the first aspect of the present invention, the nasal mucosal administration preparation further includes a resolubilizing agent, wherein the resolubilizing agent includes 0-1.3 wt% of an osmotic pressure regulator, 0-6.7 wt% of a stabilizer, 0-1.0 wt% of a thickener, and the balance is water.

[0016] In some embodiments of the first aspect of the present invention, the stabilizer is xylitol.

[0017] In some embodiments of the first aspect of the present invention, the thickener is selected from at least one of sodium hyaluronate, sodium carboxymethyl cellulose, carbomer homopolymer type C, or collagen.

[0018] In some embodiments of the first aspect of the present invention, the osmotic pressure regulator is sodium chloride or glycerol.

[0019] In some embodiments of the first aspect of the present invention, the nasal mucosal administration preparation comprises a powder and a resolubilizer. The powder is composed of 2 to 5 billion exosomes, 2 to 10 wt% lyoprotectant, 0.02 to 0.1 wt% phosphate buffer, 0.02 wt% poloxamer or Tween 80, and the balance water. The powder is dried to a moisture content of ≤5%. The resolubilizer is composed of 3.3 to 6.7 wt% stabilizer, 0.05 to 1.0 wt% thickener, and the balance water. The components of this nasal mucosal administration preparation form a gel that covers the surface of the nasal mucosa, providing an insulating effect and preventing direct contact between the nasal mucosa and allergens. Furthermore, the exosomes possess anti-inflammatory and regenerative properties, thereby producing a local anti-nasal allergy effect.

[0020] In some embodiments of the first aspect of the present invention, the nasal mucosal administration preparation comprises a powder and a resolubilizer, wherein the raw materials of the powder are composed of: 2 to 5 billion particles of exosomes, 5 to 10 wt% of a lyoprotectant, 0.02 to 0.1 wt% of phosphate buffer, 0.1 to 0.5 wt% of dodecyl maltoside, and the balance is water; the powder is dried to obtain a moisture content of ≤5%; the resolubilizer is composed of: 0.3 to 1.3 wt% of sodium chloride or glycerol, and the balance is water. The nasal mucosa area is approximately 200 cm 2 , with a rich capillary plexus and close to the brain. Targeting the brain through nasal administration can rapidly increase drug concentrations in the brain and spinal cord, which helps to circumvent the first-pass effect and the limitations of the blood-brain barrier and reduce side effects; the nasal dosage is small and administration is convenient, and patient compliance is good. The nasal mucosal administration preparation of this formula component uses the non-ionic surfactant dodecyl maltoside, and the resolvent contains only sodium chloride. Combined with the characteristics of small exosome particle size and good penetration ability, the nasal mucosal administration preparation can penetrate the blood-brain barrier, inhibit inflammation, promote damaged tissue repair, promote blood vessel growth, promote growth and regeneration, etc., and improve central nervous system diseases such as ischemic stroke, traumatic brain injury, spinal cord injury, Parkinson's disease, amyotrophic lateral sclerosis, and autism.

[0021] In some embodiments of the first aspect of the present invention, the nasal mucosal administration formulation further includes an atomizer, comprising a nozzle, a delivery tube, and a chamber. The nozzle and chamber are connected via the delivery tube, and the powder and the resolvent are placed in the chamber and mixed uniformly. The mixture of the powder and the resolvent is atomized by the atomizer and then evenly sprayed into the nasal cavity, allowing the active ingredients therein to fully contact and be absorbed by the nasal mucosa. The chamber has a volume of approximately 10 to 30 mL, and the nozzle sprays approximately 0.1 mL of solution each time, which is consumed within approximately one week.

[0022] The preparation method described in the second aspect of the present invention includes the preparation of powders and the preparation of resolubilization agents.

[0023] The preparation process of the powder: 1) Mix the raw materials of the powder evenly, sterilize by filtering through a 0.22 μm pore size filter membrane, and dispense into packaging containers; 2) Pre-freeze at -40℃ for 30~600 minutes; 3) Dry at -30~0℃ for 1200~2400 minutes; 4) Dry at 0~10℃ for 30~360 minutes; 5) Inject protective gas, seal the cap, and complete the packaging; The preparation process of the resolubilization solvent: a) Mix the raw materials of the resolvent evenly and dispense into packaging containers; b) Moist heat sterilization.

[0024] In some embodiments of the second aspect of the present invention, the protective gas is nitrogen.

[0025] In some embodiments of the second aspect of the present invention, the packaging container is a vial with cecillin.

[0026] In some embodiments of the second aspect of the present invention, the moist heat sterilization is performed by heating the mixture to 121° C. for 20 to 30 minutes. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 is an electron micrograph of the exosomes. DETAILED DESCRIPTION

[0028] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0029] In the following description, many specific details are set forth to facilitate a full understanding of the present invention. However, the present invention may also be implemented in other ways different from those described herein. Those skilled in the art may make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0030] Secondly, the term "one embodiment" or "embodiment" herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in various places throughout this specification does not necessarily refer to the same embodiment, nor does it refer to a separate or selective embodiment that is mutually exclusive of other embodiments.

[0031] The raw materials used in the present invention are all commercially available medicinal raw materials.

[0032] Example 1, preparation of nasal mucosal administration preparation No. 1.

[0033] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 5 wt% trehalose, 0.02 wt% poloxamer 188, 0.06 wt% disodium hydrogen phosphate dihydrate, and 0.02 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.93; (3) Intermediate solution b: Take 2 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 600 minutes; (ii) dry at -30°C for 2400 minutes; (iii) dry at 0°C for 360 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0034] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Obtain 0.05 wt% of carbomer homopolymer (type C), add it to an appropriate container, add appropriate amount of water to swell overnight, then add 0.015 wt% of sodium hydroxide to adjust the pH to about 7.0, add 6.7 wt% of xylitol, make up the balance with water, mix well, and obtain an intermediate solution; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 20 minutes.

[0035] Example 2, preparation of nasal mucosal administration preparation No. 2.

[0036] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 6 wt% mannitol, 0.02 wt% Tween 80, 0.07 wt% disodium hydrogen phosphate dihydrate, and 0.03 wt% sodium dihydrogen phosphate dihydrate are added to a suitable container and stirred until the solution becomes clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.97; (3) Intermediate solution b: Add 4 billion particles of exosomes derived from adipose-derived mesenchymal stem cells to intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 600 minutes; (ii) dry at -25°C for 2200 minutes; (iii) dry at 0°C for 360 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0037] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Take 0.5 wt% sodium hyaluronate, add it to an appropriate container, add appropriate amount of water to swell overnight, add 6.7 wt% xylitol, make up the balance with water, mix well, and obtain an intermediate solution; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0038] Example 3, preparation of No. 3 nasal mucosal administration preparation.

[0039] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 10 wt% trehalose, 0.02 wt% poloxamer 407, 0.08 wt% disodium hydrogen phosphate dihydrate, and 0.02 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 7.42; (3) Intermediate solution b: Take 5 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 360 minutes; (ii) dry at -20°C for 2000 minutes; (iii) dry at 5°C for 300 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0040] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Obtain 0.20 wt% sodium carboxymethyl cellulose, add it to an appropriate container, add appropriate amount of water to swell overnight, add 3.3 wt% xylitol, make up the balance with water, mix well, and obtain an intermediate solution; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0041] Example 4, preparation of No. 4 nasal mucosal administration preparation.

[0042] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 2 wt% glycine, 0.02 wt% Tween 80, 0.05 wt% disodium hydrogen phosphate dihydrate, and 0.03 wt% sodium dihydrogen phosphate dihydrate are added to a suitable container and stirred until the solution becomes clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.55; (3) Intermediate solution b: Add 5 billion particles of adipose-derived mesenchymal stem cell-derived exosomes to intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 300 min; (ii) dry at -20°C for 2000 min; (iii) dry at 10°C for 300 min; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0043] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: 1.0 wt% collagen was added to an appropriate container, and an appropriate amount of water was added to swell overnight. 3.3 wt% xylitol was added, and the remaining amount of water was added. The mixture was mixed evenly to obtain an intermediate solution. (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0044] Example 5, preparation of No. 5 nasal mucosal administration preparation.

[0045] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 2 wt% glycine, 0.02 wt% poloxamer 407, 0.06 wt% disodium hydrogen phosphate dihydrate, and 0.02 wt% sodium dihydrogen phosphate dihydrate are added to a suitable container and stirred until the solution becomes clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 7.00; (3) Intermediate solution b: Take 5 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 360 minutes; (ii) dry at -20°C for 2200 minutes; (iii) dry at 5°C for 360 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0046] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: 1.0 wt% collagen was added to an appropriate container, and an appropriate amount of water was added to swell overnight. 3.3 wt% xylitol was added, and the remaining amount of water was added. The mixture was mixed evenly to obtain an intermediate solution. (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0047] Example 6, preparation of No. 6 nasal mucosal administration preparation.

[0048] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 5 wt% trehalose, 0.1 wt% dodecyl maltoside, 0.06 wt% disodium hydrogen phosphate dihydrate, and 0.02 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 7.01; (3) Intermediate solution b: Take 2 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 600 minutes; (ii) dry at -30°C for 2400 minutes; (iii) dry at 0°C for 360 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0049] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Obtain sodium chloride with a concentration of 0.6 wt%, add it to an appropriate container, add the balance of water, mix well to obtain an intermediate solution, and sterilize and filter the intermediate solution through a 0.22 μm filter membrane before packaging; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0050] The prepared No. 6 nasal mucosal administration preparation can be delivered through the nose and brain in a targeted manner.

[0051] Example 7, preparation of nasal mucosal administration preparation No. 7.

[0052] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 6 wt% mannitol, 0.25 wt% dodecyl maltoside, 0.05 wt% disodium hydrogen phosphate dihydrate, and 0.03 wt% sodium dihydrogen phosphate dihydrate are added to a suitable container and stirred until the solution becomes clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.87; (3) Intermediate solution b: Add 4 billion particles of adipose-derived mesenchymal stem cell-derived exosomes to intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 360 minutes; (ii) dry at -25°C for 2000 minutes; (iii) dry at 0°C for 300 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0053] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: obtain 1.3 wt% glycerol, add it to an appropriate container, add the balance of water, mix well to obtain an intermediate solution, sterilize the intermediate solution by filtering it through a 0.22 μm filter membrane, and then package it; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0054] The prepared No. 7 nasal mucosal administration preparation can be delivered through the nose and brain in a targeted manner.

[0055] Example 8, preparation of No. 8 nasal mucosal administration preparation.

[0056] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 8 wt% trehalose, 0.25 wt% dodecyl maltoside, 0.04 wt% disodium hydrogen phosphate dihydrate, and 0.04 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.62; (3) Intermediate solution b: Take 5 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 300 minutes; (ii) dry at -20°C for 2200 minutes; (iii) dry at 5°C for 300 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0057] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Obtain sodium chloride with a concentration of 0.45 wt%, add it to an appropriate container, add the remaining amount of water, mix well to obtain an intermediate solution, and sterilize and filter the intermediate solution through a 0.22 μm filter membrane before packaging; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 20 minutes.

[0058] The prepared No. 8 nasal mucosal administration preparation can be delivered through the nose and brain in a targeted manner.

[0059] Example 9, preparation of No. 9 nasal mucosal administration preparation.

[0060] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 8 wt% trehalose, 0.35 wt% dodecyl maltoside, 0.05 wt% disodium hydrogen phosphate dihydrate, and 0.05 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 6.24; (3) Intermediate solution b: Add 5 billion particles of adipose-derived mesenchymal stem cell-derived exosomes to intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 300 minutes; (ii) dry at -20°C for 1800 minutes; (iii) dry at 8°C for 300 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0061] The resolubilization solution was prepared as follows: (1) Preparation of intermediate solution: Obtain sodium chloride with a concentration of 0.45 wt%, add it to an appropriate container, add the balance of water, mix well to obtain an intermediate solution, and sterilize and filter the intermediate solution through a 0.22 μm filter membrane before packaging; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 20 minutes.

[0062] The prepared No. 9 nasal mucosal administration preparation can be delivered through the nose and brain in a targeted manner.

[0063] Example 10, preparation No. 10 for nasal mucosal administration.

[0064] The powder is prepared according to the following steps: (1) Preparation of intermediate solution a: 10 wt% trehalose, 0.5 wt% dodecyl maltoside, 0.06 wt% disodium hydrogen phosphate dihydrate, and 0.02 wt% sodium dihydrogen phosphate dihydrate were added to a suitable container and stirred until the solution became clear to obtain intermediate solution a. (2) Control test: Test and confirm that the pH of intermediate solution a is 7.02; (3) Intermediate solution b: Take 5 billion particles of umbilical cord-derived mesenchymal stem cell-derived exosomes and add them to the intermediate solution a, stir evenly, and set aside; (4) Filtration and filling with half-stopper: The intermediate solution b was sterilized by filtration using a 0.22 μm PES filter membrane, and 2 mL was filled into a 5 mL vial, half-stoppered, and freeze-dried; (5) Freeze-drying procedure: (i) prefreeze at -40°C for 300 minutes; (ii) dry at -20°C for 2400 minutes; (iii) dry at 10°C for 300 minutes; (6) After freeze-drying, nitrogen is filled for protection, the plug is automatically pressed, the cap is rolled, and the outer packaging is carried out.

[0065] Prepare reconstitution solution in bottle B as follows: (1) Preparation of intermediate solution: obtain 0.6 wt% glycerol, add it to an appropriate container, add the balance of water, mix well to obtain an intermediate solution, sterilize and filter the intermediate solution through a 0.22 μm filter membrane, and then package it; (2) Subpackaging, plugging and capping: Dispense 3 mL of the intermediate solution into another 5 mL vial, add a rubber stopper, seal with an aluminum-plastic cap, and sterilize with wet heat at 121°C for 30 minutes.

[0066] The prepared No. 10 nasal mucosal administration preparation can be delivered through the nose and brain in a targeted manner.

[0067] Example 11, investigation on the effect of the nasal mucosal administration preparation on anti-allergic rhinitis.

[0068] (1) Establishment of guinea pig allergic rhinitis model: 6 guinea pigs that have been adapted to the breeding environment in the laboratory were selected. A disposable nasal drug delivery atomizer was connected to a PE tube and inserted into the guinea pig nostrils about 3 to 8 mm. Olive oil was sprayed on the nasal cannula. A 10% solution of toluene diisocyanate was prepared as a sensitizer, 5 μL per side, once a day for 7 consecutive days; then once every 2 days for a total of 4 times, and the sensitization time was 15 days. The guinea pig allergic rhinitis model was established (refer to Zhao Changqing et al., Light microscopy and electron microscopy observation of the perennial allergic rhinitis model established by using toluene diisocyanate nasal drops [J]. Chinese Journal of Otorhinolaryngology and Skull Base Surgery, 1997, 3(3): 170-173.).

[0069] (2) Grouping: After 15 days, it was confirmed that the nasal mucosa of the rhinitis model group was significantly damaged. The 6 guinea pigs in the model group were randomly divided into two groups. One group was used as the sample group and numbered as sample group 1, sample group 2, and sample group 3. The other group was used as the blank group and numbered as blank 1, blank 2, and blank 3. Three guinea pigs raised normally in the laboratory were selected as the control group and numbered as control 1, control 2, and control 3.

[0070] (3) Experiment: The sample group was sprayed with the nasal mucosal preparation prepared in Example 5, and the blank group was not treated in any way. The formula of the control group test product was as follows: each 1000 mL of exosome rhinitis spray contained: mesenchymal stem cell exosomes 500 mg, poloxamer 407 20 g, poloxamer 188 5 g, glycerol 10 g, β-glucan 6 g, gelatin 0.1 g, and the balance was deionized water.

[0071] Methods: A disposable nasal atomizer with a PE tube was inserted into the nostrils about 3 to 8 mm, and 10 μL of solution was injected into each nostril. The number of guinea pigs sneezing was observed and recorded within 30 minutes, 1 hour, and 2 hours after the injection. The test results are shown in Table 1.

[0072] Table 1: Pharmaceutical effects of the nasal mucosal administration preparation of the present invention on the rat allergic rhinitis model

[0073] The experimental results show that the nasal mucosal administration preparation prepared by the method provided by the present invention has a significant improvement effect on allergic rhinitis in guinea pigs.

[0074] Example 12, In vitro transdermal release test of the nose-brain targeted delivery formulation In this study, the in vitro release test of the spray was carried out in a modified Franz diffusion cell (RYJ-12B drug transdermal diffusion tester).

[0075] Test steps: (1) Preparation of ex vivo mouse skin After killing the mouse by dislocating the neck, immediately shave the mouse's abdominal hair with a rechargeable trimmer, cut the abdominal skin immediately, and lay the removed skin flat on a clean glass plate with the stratum corneum facing down. Carefully wipe the subcutaneous adipose stem cell layer and connective tissue with a cotton ball soaked in saline. Rinse it repeatedly with saline, place it in saline, and store it in a refrigerator at 4°C until use. Experiments should be carried out within 1 week. Before the experiment, visually inspect the integrity of the mouse skin to ensure that there is no damage.

[0076] (2) Receiving liquid The receiving solution was artificially prepared nasal fluid, and the preparation method was as follows: weigh 4.385 g NaCl, 1.49 g KCl, and 0.29 g CaCl2, dissolve them in 500 mL deionized water, and set aside.

[0077] (3) Leak detection of diffusion pool Fix the skin on a Franz diffusion cell with a stirrer, with the stratum corneum facing upwards, and secure it with a spring clamp. Fill the supply cell with water and leave it for a while to see if the liquid level drops. If there is leakage, adjust the spring clamp until it stops leaking.

[0078] (4) Add receiving solution Use a syringe to slowly add the receiving solution into the receiving cell. After adding, check whether there are bubbles between the contact interface between the skin and the receiving solution. If there are bubbles, tilt the diffusion cell slightly to remove the bubbles and fill it with receiving solution.

[0079] (5) Add sample solution Use a syringe to slowly add all the sample solution into the supply cell, the amount added is about 1 mL, and after adding, seal the supply cell with plastic wrap.

[0080] Sample 1: The powder of the nasal mucosal administration preparation prepared in Example 5 and the resolvent were mixed evenly, 3 parallel samples; Sample 2: The powder of the nasal mucosal administration preparation prepared in Example 10 and the resolvent were mixed evenly, 3 parallel samples; Blank group: physiological saline, 3 parallel samples.

[0081] (6) Sampling After 8 hours of percutaneous administration, an appropriate amount of receiving solution was slowly drawn out using a curved sampling needle.

[0082] (7) Sample processing The receptor fluid was first filtered through a 0.22 μm filter membrane to remove large tissue cell residues; The filtrate was centrifuged at 2000 g for 120 min, and the supernatant was collected; The supernatant was centrifuged at 1,000,000 g for 60 min, the supernatant was discarded, and the precipitate was collected; The pellet was resuspended in PBS buffer, pH 7.4.

[0083] (8) Particle number determination The number of particles was detected using a flow nanoanalyzer to evaluate the permeability of exosomes.

[0084] Transmittance = number of exosome particles in the receiving solution / number of exosome particles in the diffusion solution * 100% (9) Test results As shown in Table 2.

[0085] Table 2: In vitro transdermal release of nasal mucosal preparations with different excipient ratios

[0086] (10) Results analysis Experimental results showed that the nasal mucosal administration preparation obtained in Example 5, because it forms a gel after mixing, retains the transdermal diffusion of exosomes, facilitating the local action of the exosomes. The nasal mucosal administration preparation obtained in Example 10, because the resolubilization agent is close to normal saline, has a significant permeability effect on the exosomes, facilitating the nasal delivery of exosomes for the targeted treatment of brain-related diseases.

[0087] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above and that the invention can be embodied in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as illustrative and non-restrictive, and the scope of the invention is defined by the appended claims rather than the foregoing description, and it is intended that all variations that come within the meaning and range of equivalents of the claims be embraced therein.

Claims

1. A preparation for nasal mucosal administration, characterized in that: The invention relates to a powder, wherein the raw materials of the powder include: 2 to 5 billion exosomes, 2 to 10 wt% of a freeze-drying protectant, 0.02 to 0.1 wt% of a pH regulator, 0.02 to 0.5 wt% of a surfactant, and the balance is water; the powder is obtained after drying, and the moisture content is ≤5%.

2. The nasal mucosal administration preparation according to claim 1, characterized in that The exosomes are derived from umbilical cord-derived mesenchymal stem cells and / or adipose-derived mesenchymal stem cells.

3. The nasal mucosal administration preparation according to claim 1, characterized in that The freeze-drying protective agent is selected from at least one of trehalose, mannitol or glycine; the pH regulator is phosphate buffer salt, pH 6.0-8.0; and the surfactant is selected from poloxamer, Tween 80 or dodecyl maltoside.

4. The nasal mucosal administration preparation according to claim 1, characterized in that The invention also includes a complex solvent, wherein the complex solvent includes 0-1.3 wt% of an osmotic pressure regulator, 0-6.7 wt% of a stabilizer, 0-1.0 wt% of a thickener, and the balance is water.

5. The nasal mucosal administration preparation according to claim 4, characterized in that: The thickener is selected from at least one of sodium hyaluronate, sodium carboxymethyl cellulose, carbomer homopolymer type C or collagen.

6. The nasal mucosal administration preparation according to claim 4, characterized in that The osmotic pressure regulator is sodium chloride or glycerol.

7. The nasal mucosal administration preparation according to claim 1, characterized in that Including powder and reconstituted solution, The raw material composition of the powder is: 2 to 5 billion exosomes, 2 to 10 wt% of a lyoprotectant, 0.02 to 0.1 wt% of phosphate buffer, 0.02 wt% of poloxamer or Tween 80, and the balance is water; the powder is dried to obtain a moisture content of ≤5%; The composition of the complex solvent is: 3.3-6.7 wt% of a stabilizer, 0.05-1.0 wt% of a thickener, and the balance is water.

8. The nasal mucosal administration preparation according to claim 1, characterized in that Including powder and reconstituted solution, The raw material composition of the powder is: 2 to 5 billion exosomes, 5 to 10 wt% of a freeze-drying protectant, 0.02 to 0.1 wt% of phosphate buffer, 0.1 to 0.5 wt% of dodecyl maltoside, and the balance is water; the powder is obtained after drying, and the moisture content is ≤5%; The composition of the resolvent is: 0.3-1.3 wt% of sodium chloride or glycerol, and the balance is water.

9. The nasal mucosal administration preparation according to any one of claims 4 to 8, characterized in that The invention also includes an atomizing device, which includes a nozzle, a delivery pipe and a cavity. The nozzle and the cavity are connected through the delivery pipe. The powder and the resolvent are placed in the cavity and mixed evenly.

10. The method for preparing the nasal mucosal administration preparation according to any one of claims 1 to 9, characterized in that: The preparation process of the powder comprises the following steps: 1) Mix the raw materials of the powder evenly, sterilize by filtering through a 0.22 μm pore size filter membrane, and dispense into packaging containers; 2) Pre-freeze at -40℃ for 30~600 minutes; 3) Dry at -30~0℃ for 1200~2400 minutes; 4) Dry at 0~10℃ for 30~360 minutes; 5) Inject protective gas, seal the cap, and complete the packaging.

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