An exosome oral spray and a preparation method thereof

Abstract

This invention relates to an exosome-based oral spray and its preparation method, belonging to the field of oral spray technology. By weight, the exosome-based oral spray comprises the following components: 0.2-0.7 parts of human umbilical cord mesenchymal stem cell exosomes, 1-5 parts of natural extract, 8-15 parts of moisturizing solubilizer, 0.1-0.5 parts of preservative, 0.1-0.5 parts of fragrance, and 78-90 parts of deionized water. The natural extract is obtained by water extraction from *Xanthoceras sorbifolium* and *Sapindus mukorossi* seeds in a weight ratio of (14-20):(5-11). This invention utilizes human umbilical cord mesenchymal stem cell exosomes and a specific composition of natural extract as active ingredients. The synergistic effect of these two components significantly improves the repair effect of the resulting exosome-based oral spray on skin damage, which is beneficial for the treatment of common oral diseases such as oral ulcers and stomatitis, and has a wide range of applications.

Description

Technical Field

[0001] This invention relates to the field of oral spray technology, and more particularly to an exosome oral spray and its preparation method. Background Technology

[0002] In the field of oral health, oral diseases such as oral ulcers and stomatitis are common and prone to recurrence, causing considerable inconvenience and suffering for patients. These diseases are often related to factors such as an imbalance in the oral environment, weakened immunity, and bacterial or viral infections. Traditional treatments often involve topical medications, such as anti-inflammatory drugs and analgesics, which can temporarily relieve symptoms but often fail to cure the underlying condition, and long-term use may produce side effects.

[0003] In recent years, with the continuous development of biotechnology, the application of exosomes in the medical field has gradually attracted attention. Exosomes are tiny vesicles secreted by cells, rich in various bioactive substances, such as growth factors and cytokines, and have multiple functions, including promoting cell proliferation, differentiation, tissue repair, and immune regulation. Therefore, applying exosomes to the treatment of oral diseases is expected to fundamentally promote the repair and regeneration of oral mucosa, improve oral immunity, and thus achieve the goal of treating oral diseases. However, there are currently relatively few exosome-based oral care products on the market, and some problems exist, such as poor repair effects on damaged skin and mucosa, which need further improvement to meet the needs of oral care products. Summary of the Invention

[0004] To address the above problems, this invention provides an exosome oral spray and its preparation method.

[0005] In a first aspect, the present invention provides an exosome oral spray, which, by weight, comprises the following components:

[0006] Human umbilical cord mesenchymal stem cell exosomes 0.2-0.7 parts, natural extracts 1-5 parts, moisturizing solubilizers 8-15 parts, preservatives 0.1-0.5 parts, fragrances 0.1-0.5 parts, and deionized water 78-90 parts;

[0007] The natural extract is obtained by water extraction using alligator crocodile flower and Sapindus mukorossi seed kernels in a weight ratio of (14~20):(5~11).

[0008] Furthermore, the exosome oral spray comprises the following components in parts by weight:

[0009] The ingredients consist of 0.4 parts human umbilical cord mesenchymal stem cell exosomes, 3.5 parts natural extracts, 12 parts moisturizing solubilizers, 0.2 parts preservatives, 0.4 parts fragrances, and 83.5 parts deionized water.

[0010] Furthermore, the weight ratio of the alligator beak flower to the kernel of the *Sapindus mukorossi* is 16:9.

[0011] Furthermore, the water extraction method is vacuum reflux extraction.

[0012] Furthermore, the working conditions for the reduced pressure reflux extraction include: a temperature of 60~68℃, a pressure of 16~23kPa, a time of 1~2 hours, and a number of extractions of 1~3 times.

[0013] Furthermore, the moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of (4~7):(1~3).

[0014] Furthermore, the preservative includes sodium benzoate, and the flavoring includes blueberry flavoring.

[0015] In a second aspect, the present invention provides a method for preparing the exosome oral spray according to any one of the first aspects, the preparation method comprising the following steps:

[0016] Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture;

[0017] Add preservatives and flavorings to the first mixture and stir to obtain the second mixture;

[0018] The second mixture is homogenized to obtain the exosome oral spray.

[0019] Furthermore, the working conditions for ultrasonic stirring and mixing include: ultrasonic power of 200~400W, stirring speed of 100~200rpm, and time of 5~15 minutes; the working conditions for homogenization include: homogenization pressure of 15~20MPa, and time of 5~10 minutes.

[0020] The technical solutions provided in the embodiments of the present invention have at least the following advantages compared with the prior art:

[0021] This invention provides an exosome-based oral spray. By using human umbilical cord mesenchymal stem cell exosomes and a specific composition of natural extracts as active ingredients, the two work synergistically to significantly improve the repair effect of the resulting exosome-based oral spray on skin damage. This is beneficial for the treatment of common oral diseases such as oral ulcers and stomatitis, and has a wide range of applications. Specifically:

[0022] 1) This invention utilizes the combination of human umbilical cord mesenchymal stem cell exosomes with extracts of *Xanthoceras sorbifolium* and *Sapindus mukorossi* seeds to create a complementary effect. On one hand, the growth factors and cytokines provided by the exosomes promote cell proliferation and differentiation, and combined with the anti-inflammatory and antioxidant components in the natural extracts, enhance the repair effect, enabling faster repair of damaged oral mucosa. On the other hand, the immunomodulatory effect of the exosomes complements the pharmacological activity of the natural extracts, enabling a more comprehensive regulation of the oral microenvironment, enhancing the resistance of the oral mucosa, improving immunomodulatory capacity, comprehensively improving oral health, and preventing the recurrence of oral ulcers, stomatitis, and other diseases.

[0023] 2) This invention combines the moisturizing and protective effects of natural extracts with the repairing effects of exosomes. The raw materials are green, natural and safe, making the oral spray more comfortable to use, reducing pain and discomfort, helping patients to persist in using it, thereby improving the treatment effect. Detailed Implementation

[0024] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. 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.

[0025] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in this invention can be purchased from the market or prepared by existing methods.

[0026] In a first aspect, the present invention provides an exosome oral spray, which, by weight, comprises the following components:

[0027] Human umbilical cord mesenchymal stem cell exosomes 0.2-0.7 parts, natural extracts 1-5 parts, moisturizing solubilizers 8-15 parts, preservatives 0.1-0.5 parts, fragrances 0.1-0.5 parts, and deionized water 78-90 parts;

[0028] The natural extract is obtained by water extraction using alligator crocodile flower and Sapindus mukorossi seed kernels in a weight ratio of (14~20):(5~11).

[0029] This invention provides an exosome-based oral spray. By using human umbilical cord mesenchymal stem cell exosomes and a specific composition of natural extracts as active ingredients, the two work synergistically to significantly improve the repair effect of the resulting exosome-based oral spray on skin damage. This is beneficial for the treatment of common oral diseases such as oral ulcers and stomatitis, and has a wide range of applications. Specifically:

[0030] 1) This invention utilizes the combination of human umbilical cord mesenchymal stem cell exosomes with extracts of *Xanthoceras sorbifolium* and *Sapindus mukorossi* seeds to create a complementary effect. On one hand, the growth factors and cytokines provided by the exosomes promote cell proliferation and differentiation, and combined with the anti-inflammatory and antioxidant components in the natural extracts, enhance the repair effect, enabling faster repair of damaged oral mucosa. On the other hand, the immunomodulatory effect of the exosomes complements the pharmacological activity of the natural extracts, enabling a more comprehensive regulation of the oral microenvironment, enhancing the resistance of the oral mucosa, improving immunomodulatory capacity, comprehensively improving oral health, and preventing the recurrence of oral ulcers, stomatitis, and other diseases.

[0031] 2) This invention combines the moisturizing and protective effects of natural extracts with the repairing effects of exosomes. The raw materials are green, natural and safe, making the oral spray more comfortable to use, reducing pain and discomfort, helping patients to persist in using it, thereby improving the treatment effect.

[0032] In some specific embodiments, the human umbilical cord mesenchymal stem cell exosomes in this invention can be directly obtained from commercially available human umbilical cord mesenchymal stem cell exosomes, such as those from Shanghai Fuyu Biotechnology Co., Ltd., catalog number WM-23GK0032 (mass concentration of 1 mg / L, BCA protein concentration of 0.5~1.0 mg / mL); alternatively, they can be prepared in-house according to existing human umbilical cord mesenchymal stem cell exosome preparation processes (such as high-speed centrifugation, density gradient centrifugation, etc.), specifically using the following methods:

[0033] 1. Umbilical cord collection and preliminary processing:

[0034] Under aseptic conditions, fresh umbilical cords were collected from healthy mothers, and Wharton's jelly tissue was carefully separated from the cord to avoid contamination. The separated Wharton's jelly tissue was cut into small pieces and evenly attached to the walls of culture flasks to ensure that the cells could adhere and grow fully. Complete culture medium was added, and the flasks were placed under suitable culture conditions to allow the umbilical cord mesenchymal stem cells to naturally emerge and proliferate.

[0035] 2. Cell expansion and collection of culture medium supernatant:

[0036] After the cells grow to a certain density, they are passaged and expanded to obtain a sufficient number of umbilical cord mesenchymal stem cells. The culture supernatant of the fifth generation cells is selected for collection, at which time the cells are in a stable state and the amount of exosomes secreted is high.

[0037] 3. Preliminary centrifugation to remove residual cells:

[0038] Aliquot the collected culture medium supernatant into centrifuge tubes and centrifuge at 300G for 10 minutes to remove any remaining live cells. Carefully collect the first supernatant after centrifugation, avoiding aspiration of cell precipitates.

[0039] 4. Further centrifugation to remove dead cells:

[0040] The first supernatant was aliquoted into centrifuge tubes and centrifuged at 3000G for 30 minutes to remove dead cells and larger cell debris; the second supernatant was collected after centrifugation to ensure the purity of exosomes.

[0041] 5. High-speed centrifugation to remove cell debris:

[0042] Transfer the second supernatant to a clean ultracentrifuge tube and centrifuge at 10,000 G for 30 minutes to thoroughly remove cell debris and other impurities; carefully collect the third supernatant after centrifugation, which mainly contains exosomes and other small molecules.

[0043] 6. Finally, exosomes are obtained by ultracentrifugation:

[0044] The third supernatant was aliquoted into clean ultracentrifuge tubes and centrifuged at 100,000 G for 90 minutes to precipitate the exosomes. After centrifugation, the supernatant was discarded, and an appropriate amount of physiological saline was added to each tube to resuspend the precipitate, thus obtaining an exosome resuspension.

[0045] 7. Filtration, sterilization, and finished product preparation:

[0046] The exosome resuspension was transferred to a new centrifuge tube and filtered with a 0.22 μm filter to ensure the sterility of the exosomes; the filtered liquid is the human umbilical cord mesenchymal stem cell exosome.

[0047] The alligator beak flower used in this invention, scientifically known as *Clinacanthus nutans* (Burm. f.) Lindau, is an herbaceous plant belonging to the genus *Clinacanthus* in the family Acanthaceae, and is considered a plant with both medicinal and edible uses. Modern scientific research has found that alligator beak flowers are rich in flavonoids, lupeol, betulin, sitosterol, polypeptides, as well as various trace elements and vitamins. These components endow alligator beak flowers with a variety of pharmacological effects, such as enhancing immunity, anti-tumor activity, anti-inflammatory and antibacterial properties, and antioxidant effects. Simultaneously, alligator beak flowers can also lower blood pressure and blood lipids, and promote intestinal peristalsis, playing a positive role in the prevention and treatment of various diseases.

[0048] This invention relates to *Xanthoceras sorbifolium* seed kernels, made from the seeds of *Xanthoceras sorbifolium* (family Sapindaceae). The kernels undergo drying, magnetic separation, dehulling, and screening processes, resulting in a natural plant product that is both food and medicine. According to traditional medical records and modern scientific research, *Xanthoceras sorbifolium* seed kernels possess various medicinal properties. For example, *Xanthoceras sorbifolium* saponins, extracted from the seed kernels, are important drugs for improving brain function. Simultaneously, various active ingredients in *Xanthoceras sorbifolium* seed kernels, such as flavonoids and coumarins, exhibit antioxidant, antitumor, and anti-inflammatory pharmacological effects. Furthermore, *Xanthoceras sorbifolium* seed kernels are rich in unsaturated fatty acids and essential oils, which help lower cholesterol, soften blood vessels, and prevent cardiovascular diseases.

[0049] In some specific embodiments, the exosome oral spray comprises the following components, by weight:

[0050] The ingredients consist of 0.4 parts human umbilical cord mesenchymal stem cell exosomes, 3.5 parts natural extracts, 12 parts moisturizing solubilizers, 0.2 parts preservatives, 0.4 parts fragrances, and 83.5 parts deionized water.

[0051] In some specific embodiments, the weight ratio of the alligator beak flower to the kernel of the *Sapindus mukorossi* is 16:9.

[0052] In some specific embodiments, the water extraction method is vacuum reflux extraction.

[0053] In some specific embodiments, the operating conditions for the reduced pressure reflux extraction include: a temperature of 60-68°C, a pressure of 16-23 kPa, a time of 1-2 hours, and 1-3 extraction cycles. Preferably, the temperature for the reduced pressure reflux extraction is 65°C, and the pressure is 20 kPa.

[0054] In some specific embodiments, the moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of (4~7):(1~3). Preferably, the moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of 5:2.

[0055] In some specific embodiments, the preservative includes sodium benzoate, and the flavoring includes blueberry flavoring.

[0056] Secondly, based on the same inventive concept, the present invention provides a method for preparing the exosome oral spray according to any one of the first aspects, the preparation method comprising the following steps:

[0057] Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture;

[0058] Add preservatives and flavorings to the first mixture and stir to obtain the second mixture;

[0059] The second mixture is homogenized to obtain the exosome oral spray.

[0060] The method for preparing the exosome oral spray provided by this invention is simple to operate, requires no additional special equipment, greatly reduces the production threshold and cost, and is suitable for industrial mass production. The resulting exosome oral spray, while maintaining its high activity, can seamlessly integrate with existing filling processes, easily achieving efficient filling into the spray bottle, ensuring that the final product is both portable and easy to use. Furthermore, this preparation method is based on the exosome oral spray described in any one of the first aspects, and therefore possesses at least the beneficial effects of the exosome oral spray described in any one of the first aspects, which will not be elaborated further here.

[0061] In some specific embodiments, the working conditions for ultrasonic stirring and mixing include: ultrasonic power of 200~400W, stirring speed of 100~200rpm, and time of 5~15 minutes; the working conditions for homogenization include: homogenization pressure of 15~20MPa and time of 5~10 minutes.

[0062] It should be noted that, unless otherwise specified or described, the raw materials involved in the exosome oral spray and its preparation method provided in the embodiments of the present invention can be directly made from commercially available products or by using existing publicly disclosed preparation methods; at the same time, unless otherwise specified or described, the steps and parameters involved can be carried out according to the existing oral spray processing technology or directly using existing equipment, and will not be described in detail in this document.

[0063] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed according to national standards. If no corresponding national standard exists, then generally accepted international standards, conventional conditions, or conditions recommended by the manufacturer are followed.

[0064] Example 1

[0065] This example provides an exosome oral spray, which, by weight, comprises the following components:

[0066] 0.4 parts human umbilical cord mesenchymal stem cell exosomes, 3.5 parts natural extracts, 12 parts moisturizing solubilizers, 0.2 parts preservatives, 0.4 parts fragrances, and 83.5 parts deionized water;

[0067] The human umbilical cord mesenchymal stem cell exosomes were selected from Shanghai Fuyu Biotechnology Co., Ltd., with the product number WM-23GK0032;

[0068] The natural extract is obtained by water extraction of *Syzygium buergerianum* and *Sapindus mukorossi* seeds in a weight ratio of 16:9. The preparation method includes the following steps: *Syzygium buergerianum* and *Sapindus mukorossi* seeds are mixed and pulverized, then water is added in a volume of 8 times the total weight of *Syzygium buergerianum* and *Sapindus mukorossi* seeds for vacuum reflux extraction. The vacuum reflux extraction temperature is 65℃, the vacuum reflux extraction pressure is 20 kPa, the vacuum reflux extraction time is 1.5 hours, and the vacuum reflux extraction is performed twice. The extract is filtered, the resulting water extracts are combined, concentrated under reduced pressure, and dried to obtain the natural extract.

[0069] The moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of 5:2.

[0070] The preservative is sodium benzoate, and the flavoring is blueberry flavoring.

[0071] The preparation method of the above-mentioned exosome oral spray includes the following steps:

[0072] Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture;

[0073] Add preservatives and flavorings to the first mixture and stir to obtain the second mixture;

[0074] The second mixture is homogenized to obtain the exosome oral spray;

[0075] The working conditions for ultrasonic stirring and mixing include: ultrasonic power of 300W, stirring speed of 160rpm, and time of 10 minutes; the working conditions for homogenization include: homogenization pressure of 18MPa and time of 7 minutes.

[0076] Example 2

[0077] This example provides an exosome oral spray, which, by weight, comprises the following components:

[0078] 0.2 parts human umbilical cord mesenchymal stem cell exosomes, 1 part natural extract, 8 parts moisturizing solubilizer, 0.2 parts preservative, 0.1 parts fragrance and 78 parts deionized water;

[0079] The human umbilical cord mesenchymal stem cell exosomes were selected from Shanghai Fuyu Biotechnology Co., Ltd., with the product number WM-23GK0032;

[0080] The natural extract is obtained by water extraction of *Alligator haematomarginatus* and *Sapindus mukorossi* seeds in a weight ratio of 20:11. The preparation method includes the following steps: mixing and pulverizing *Alligator haematomarginatus* and *Sapindus mukorossi* seeds, then adding water equal to 8 times the total weight of the two seeds for vacuum reflux extraction. The vacuum reflux extraction temperature is 60℃, the vacuum reflux extraction pressure is 16 kPa, the vacuum reflux extraction time is 1.5 hours, and the vacuum reflux extraction is performed twice. The extract is then filtered, and the resulting water extracts are combined, concentrated under reduced pressure, and dried to obtain the natural extract.

[0081] The moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of 7:3.

[0082] The preservative is sodium benzoate, and the flavoring is blueberry flavoring.

[0083] The preparation method of the above-mentioned exosome oral spray includes the following steps:

[0084] Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture;

[0085] Add preservatives and flavorings to the first mixture and stir to obtain the second mixture;

[0086] The second mixture is homogenized to obtain the exosome oral spray;

[0087] The working conditions for ultrasonic stirring and mixing include: ultrasonic power of 400W, stirring speed of 200rpm, and time of 5 minutes; the working conditions for homogenization include: homogenization pressure of 15MPa and time of 10 minutes.

[0088] Example 3

[0089] This example provides an exosome oral spray, which, by weight, comprises the following components:

[0090] 0.7 parts human umbilical cord mesenchymal stem cell exosomes, 5 parts natural extracts, 15 parts moisturizing solubilizers, 0.5 parts preservatives, 0.5 parts fragrances, and 90 parts deionized water;

[0091] The human umbilical cord mesenchymal stem cell exosomes were selected from Shanghai Fuyu Biotechnology Co., Ltd., with the product number WM-23GK0032;

[0092] The natural extract is obtained by water extraction of *Syzygium buergerianum* and *Sapindus mukorossi* seeds in a weight ratio of 14:5. The preparation method includes the following steps: *Syzygium buergerianum* and *Sapindus mukorossi* seeds are mixed and pulverized, and then water is added in a volume of 8 times the total weight of *Syzygium buergerianum* and *Sapindus mukorossi* seeds for vacuum reflux extraction. The vacuum reflux extraction temperature is 68℃, the vacuum reflux extraction pressure is 23 kPa, the vacuum reflux extraction time is 1.5 hours, and the vacuum reflux extraction is performed twice. The extract is filtered, the resulting water extracts are combined, concentrated under reduced pressure, and dried to obtain the natural extract.

[0093] The moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of 4:1.

[0094] The preservative is sodium benzoate, and the flavoring is blueberry flavoring.

[0095] The preparation method of the above-mentioned exosome oral spray includes the following steps:

[0096] Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture;

[0097] Add preservatives and flavorings to the first mixture and stir to obtain the second mixture;

[0098] The second mixture is homogenized to obtain the exosome oral spray;

[0099] The working conditions for ultrasonic stirring and mixing include: ultrasonic power of 200W, stirring speed of 100rpm, and time of 15 minutes; the working conditions for homogenization include: homogenization pressure of 20MPa and time of 5 minutes.

[0100] Example 4

[0101] This example provides an exosome oral spray and its preparation method, which differs from Example 1 only in that the temperature of the vacuum reflux extraction in the preparation method of the natural extract is adjusted to 86°C and the pressure of the vacuum reflux extraction is adjusted to 55 kPa; the remaining steps and parameters are the same.

[0102] Comparative Example 1

[0103] This example provides an exosome oral spray and its preparation method, which differs from Example 1 only in that the amount of natural extract added is 0 parts (i.e., no natural extract is added and the corresponding preparation process of adding natural extract is omitted); the other steps and parameters are the same.

[0104] Comparative Example 2

[0105] This example provides an exosome oral spray and its preparation method, which differs from Example 1 only in that the natural extract is obtained by water extraction of *Xanthoceras sorbifolium* (i.e., without the addition of *Xanthoceras sorbifolium* seeds); the remaining steps and parameters are the same.

[0106] Comparative Example 3

[0107] This example provides an exosome oral spray and its preparation method, which differs from Example 1 only in that the natural extract is obtained by water extraction using the kernel of *Sapindus mukorossi* (i.e., without the addition of *Gynostemma pentaphyllum*); the remaining steps and parameters are the same.

[0108] Comparative Example 4

[0109] This example provides an exosome oral spray and its preparation method, which differs from Example 1 only in that the natural extract is obtained by water extraction using alligator crocodile flower and Sapindus mukorossi seed kernels in a weight ratio of 9:25; the remaining steps and parameters are the same.

[0110] Test case

[0111] In this example, the exosome oral sprays obtained from Examples 1-4 and Comparative Examples 1-4 were tested for their skin repair effects.

[0112] The testing method is as follows:

[0113] 1. Experimental animals: Healthy male BALB / c mice aged 6-8 weeks were selected as experimental subjects and subjected to an acclimatization period of 2 weeks to ensure that the mice's physiological state was stable.

[0114] 2. Grouping and Sample Size: Mice were randomly divided into 9 groups of 5 mice each, for a total of 45 mice. One group served as the blank control group (treated only with saline, labeled as group C), and the other 8 groups served as experimental groups, which were treated with the exosome oral sprays obtained in Examples 1 to 4 (corresponding to groups E1-E4) and Comparative Examples 1 to 4 (corresponding to groups P1-P4).

[0115] 3. Wound model establishment: On day 0 of the experiment, mice were anesthetized with 5% chloral hydrate. Then, two circular full-thickness skin wounds with a diameter of about 10 mm were created on both sides of the back of the mice using a skin puncher to ensure that the wound conditions of each group of mice were consistent.

[0116] 4. Sample application: Starting from the day the wound is created, apply the corresponding test sample to the mouse wound once a day, 100 μL per wound each time, ensuring that the sample evenly covers the entire wound area. This process is repeated for 3 consecutive days to observe the initial treatment effect.

[0117] 5. Data Collection and Evaluation:

[0118] Wound photography record: On the 4th day after sample application, high-resolution photographs were taken of the wounds on the backs of mice to record changes in wound morphology. Wound healing rate calculation: The wound healing rate was calculated using the following formula: Wound healing rate (%) = (S1-S2) / S1×100%, where S1 is the initial wound area and S2 is the wound area after the experimental test.

[0119] The test results are shown in Table 1.

[0120] Table 1

[0121]

[0122] As shown in Table 1, compared with the control group (Comparative Example 1, P1), Comparative Example 2, and Comparative Example 3 (P3), the exosome oral sprays developed in the four embodiments of the present invention—Example 1 (E1), Example 2 (E2), Example 3 (E3), and Example 4 (E4)—exhibited higher therapeutic efficiency and better therapeutic effects in promoting short-term wound healing of damaged skin. This indicates that the present invention, by using human umbilical cord mesenchymal stem cell exosomes and a specific composition of natural extracts as active ingredients, achieves synergy between the two, thereby significantly enhancing the therapeutic and repair capabilities of the oral spray for skin damage. It is applicable to the treatment of common oral diseases such as oral ulcers and oral mucositis, providing patients with a new and more effective treatment option. Furthermore, by comparing Comparative Example 4 (P4) with Example 1 (E1), the skin repair effect of Example 1 (E1) of the present invention is more significant, indicating that the dosage of the natural extract *Cornus officinalis* seed kernel should not be excessive.

[0123] In addition, this example refers to the YY / T0127.13-2009 pharmaceutical industry standard to conduct an oral mucosal irritation test on the composition provided in the embodiment of the present invention. The test results show that the composition provided in the embodiment of the present invention has a reaction grade of 0 on oral mucosal tissue, that is, no tissue irritation reaction, which can meet the use requirements of oral care products.

[0124] In summary, the embodiments of the present invention provide an exosome oral spray and its preparation method. By using human umbilical cord mesenchymal stem cell exosomes and a specific composition of natural extracts as active ingredients, the two work synergistically to significantly improve the repair effect of the resulting exosome oral spray on skin damage, which is beneficial for the treatment of common oral diseases such as oral ulcers and oral mucositis, and has a wide range of applications.

[0125] Various embodiments of the present invention may exist in the form of a range; it should be understood that the description in the form of a range is merely for convenience and brevity and should not be construed as a hard limitation on the scope of the invention; therefore, it should be considered that the range description has specifically disclosed all possible subranges and single numerical values ​​within that range. For example, it should be considered that the range description from 1 to 6 has specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and single numbers within the range, such as 1, 2, 3, 4, 5, and 6, regardless of the range. Furthermore, whenever a numerical range is referred to herein, it means including any referenced number (fraction or integer) within the range referred to.

[0126] The above description is merely a specific embodiment of the present invention, enabling those skilled in the art to understand or implement the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the present invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features claimed herein.

Claims

1. The application of an exosome spray in the preparation of a drug to promote the healing of damaged skin wounds, characterized in that, The exosome spray comprises the following components in parts by weight: 0.4 parts human umbilical cord mesenchymal stem cell exosomes, 3.5 parts natural extracts, 12 parts moisturizing solubilizers, 0.2 parts preservatives, 0.4 parts fragrances, and 83.5 parts deionized water; The human umbilical cord mesenchymal stem cell exosomes were selected from Shanghai Fuyu Biotechnology Co., Ltd., with the product number WM-23GK0032; The natural extract is obtained by water extraction using alligator crocodile flower and *Sapindus mukorossi* seed kernels in a weight ratio of 16:

9. Its preparation method is as follows. The process includes the following steps: Alligator's mouth flower and *Sapindus mukorossi* seed kernels are mixed and pulverized, then water is added in an amount equal to 8 times the total weight of the alligator's mouth flower and *Sapindus mukorossi* seed kernels for vacuum reflux extraction. The vacuum reflux extraction temperature is 65℃, the vacuum reflux extraction pressure is 20 kPa, the vacuum reflux extraction time is 1.5 hours, and the vacuum reflux extraction is performed twice. The mixture is filtered, the resulting aqueous extracts are combined, concentrated under reduced pressure, and dried to obtain the natural extract. The moisturizing solubilizer is composed of betaine and 1,2-propanediol in a weight ratio of 5:

2. The preservative is sodium benzoate, and the flavoring is blueberry flavoring; The preparation method of the exosome spray includes the following steps: Human umbilical cord mesenchymal stem cell exosomes, natural extracts, moisturizing solubilizers, and water were ultrasonically stirred and mixed to obtain the first mixture; Add preservatives and flavorings to the first mixture and stir to obtain the second mixture; The second mixture is homogenized to obtain the exosome spray; The working conditions for ultrasonic stirring and mixing include: ultrasonic power of 300W, stirring speed of 160rpm, and time of 10 minutes; the working conditions for homogenization include: homogenization pressure of 18MPa and time of 7 minutes.

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