Chitosan genital tract biological antibacterial composition and preparation method thereof

Abstract

The invention provides a chitosan genital tract biological antibacterial composition and a preparation method thereof, and belongs to the technical field of biological antibacterial products, the chitosan genital tract biological antibacterial composition comprises the following components by mass: 7-10% of modified chitosan, 3-6% of a plant extract, 3-6% of glycerin, 1-3% of a surfactant, and the balance of deionized water. According to the invention, the plant extract prepared by extracting a plurality of Chinese herbal medicines and the modified chitosan are compounded as the antibacterial component, so that the antibacterial property is improved, and the recurrence rate of the antibacterial composition on the cured mycotic vaginitis is reduced. The chitosan genital tract biological antibacterial composition disclosed by the invention has the advantages of single and definite treatment purpose, pertinence in treatment of colpitis mycotica, lasting antibacterial property, good treatment effect, low possibility of relapse, high product stability and convenience in storage.

Description

Technical Field

[0001] The invention belongs to the technical field of biological antibacterial products, and specifically relates to a chitosan reproductive tract biological antibacterial composition and a preparation method thereof. Background Art

[0002] There is a series of normal microbial communities in the female reproductive tract, which work together to maintain the stability of the vaginal environment and prevent the excessive reproduction of pathogenic microorganisms and cause infection. Fungal vaginitis, also known as vulvovaginal candidiasis, is a common female vaginal inflammation caused by the excessive proliferation of Candida (especially Candida albicans). The main symptoms include vulvar itching, abnormal leucorrhea, vaginal discomfort, edema, and vaginitis changes. In terms of treatment, antifungal drugs are usually used, such as topically applied vaginal suppositories or creams such as clotrimazole and miconazole. Severe or recurrent cases may require oral antifungal drugs. However, long-term use of antifungal drugs may cause Candida to develop resistance to antifungal drugs, making the treatment effect weakened or even ineffective, and prone to recurrence. Improper use may lead to the destruction of the pH value of the private part, acid-base imbalance, and lower immunity in the private part, thus causing various gynecological problems.

[0003] Chitosan's antibacterial effect belongs to physical antibacterial, and it does not produce drug resistance. At the same time, chitosan has the effect of promoting the repair of damaged tissues and the proliferation of lactobacilli. In recent years, this special selective antibacterial effect of chitosan has attracted much attention in the medical community. Most of the gynecological chitosan gel dressings on the market are for the treatment of gynecological diseases, but the treatment purpose is relatively broad, not specifically for candidal vaginitis, and the effect of use is not ideal, and it is easy to relapse. In addition, chitosan, as a polysaccharide compound, is insoluble in water. If it is not handled properly, it may cause uneven dispersion of chitosan in the antibacterial solution or form precipitation, and it cannot fully contact with the lesion, affecting the effect of use. At the same time, the stability of chitosan in the solution is affected by fluctuations in external environmental conditions, which affects the clarity of the solution, and the antibacterial effect weakens over time.

[0004] Therefore, there is an urgent need for an antibacterial product containing chitosan that has good therapeutic effect, long-lasting antibacterial effect, and high storage stability for candidal vaginitis. Summary of the invention

[0005] The purpose of the present invention is to provide a chitosan reproductive tract biological antibacterial composition and a preparation method thereof. The chitosan reproductive tract biological antibacterial composition has a single and clear therapeutic purpose, specifically treats candidal vaginitis, has long-lasting antibacterial properties, good therapeutic effects, is not prone to recurrence, has high product stability, and is easy to store.

[0006] In order to achieve the above object, the present invention provides the following technical solutions: A chitosan reproductive tract biological antibacterial composition comprises the following components in percentage by mass: 7-10% of modified chitosan, 3-6% of plant extract, 3-6% of glycerol and 1-3% of surfactant, and the balance is deionized water.

[0007] Furthermore, the preparation method of the modified chitosan is: (1) Mix chitosan and glacial acetic acid aqueous solution in a mass ratio of 0.1-0.3:100, stir evenly, and obtain a chitosan solution; (2) adding polyhexamethylene guanidine to the chitosan solution, wherein the mass ratio of polyhexamethylene guanidine to chitosan is 0.2-0.6:1; reacting at 25-30° C. for 80-100 min to obtain a chitosan solution modified with polyhexamethylene guanidine; (3) Mix N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, cysteine ​​and phosphate buffer solution in a mass ratio of 6-8:4-7:1:10-12, stir at 30-40°C for 2-4 hours to obtain a mixed solution, add the mixed solution to a polyhexamethyleneguanidine-modified chitosan solution, wherein the mass ratio of cysteine ​​to chitosan is 0.4-0.8:1, and react at 30-35°C for 40-45 hours to obtain a cysteine-modified chitosan solution; (4) The cysteine-modified chitosan solution is placed in a dialysis bag and dialyzed in deionized water. The molecular weight cutoff of the dialysis bag is 7000Da. The water is changed every 6 hours. After changing the water 7 times, the product in the dialysis bag is frozen and dried to obtain modified chitosan.

[0008] Chitosan, deacetylation degree ≥90%. Zhejiang Jinke Pharmaceutical Co., Ltd., chitosan for medical use.

[0009] Polyhexamethyleneguanidine, Tianjin Liangneng Biotechnology Co., Ltd., PHMG powder.

[0010] N-Hydroxysuccinimide, CAS: 6066-82-6.

[0011] 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, CAS: 25952-53-8.

[0012] Phosphate buffer solution, pH 6.6. Merck, Cat. No. P8165.

[0013] Furthermore, the mass fraction of glacial acetic acid in the glacial acetic acid aqueous solution is 1-3%.

[0014] In order to solve the problem that chitosan is insoluble in water, the present invention modifies chitosan with polyhexamethylene guanidine, introduces amino groups on chitosan, improves the water solubility of chitosan, and at the same time, the amino groups can effectively interfere with the synthesis and metabolic pathways of the bacterial cell wall of Candida albicans, inhibit bacterial growth, and enhance the antibacterial properties of chitosan. The inventor further modified chitosan with cysteine ​​to improve the antibacterial stability of chitosan. Analysis shows that the sulfhydryl group on cysteine ​​participates in maintaining the three-dimensional conformation of chitosan and forms a stable structure with other molecules in the system, thereby enhancing the antibacterial stability.

[0015] Furthermore, the preparation method of the plant extract is: S1: Weigh 10-15 parts by weight of Smilax glabra, 3-8 parts by weight of Cimicifuga heracleifolia, 14-19 parts by weight of Sijiqing, 5-10 parts by weight of Herba Lycopodii and 9-14 parts by weight of Lophatherum gracile, mix and grind, and pass through a 60-mesh sieve to obtain a mixed powder; S2: Mix the mixed powder and deionized water in a mass ratio of 1:8-10, sterilize, and obtain a mixture; mix Lactobacillus brevis, Bifidobacterium longum, and Lactobacillus bulgaricus in a mass ratio of 1:3.2-3.5:0.4-0.7 as a mixed bacterial agent; add the mixed bacterial agent and the mixture in a mass ratio of 0.1-0.3:10 into a fermentation tank, adjust the pH to 6.1-6.3, perform anaerobically fermentation at 36-40°C for 3-5 days, sterilize, and obtain a fermentation system; S3: The fermentation system is subjected to ultrasound, and then filtered through a 0.22 μm filter membrane, and then subjected to ultrafiltration treatment. The molecular weight cutoff of the ultrafiltration membrane is 20,000 Da. The permeate is concentrated under reduced pressure at 50-60° C. and dried to obtain a plant extract.

[0016] Furthermore, the ultrasonic condition is: the ultrasonic device is set in an alternating magnetic field, and the strength of the alternating magnetic field is 5000-5500 GS.

[0017] Furthermore, the ultrasound time is 80-100 min, and the frequency is 180-200 Hz.

[0018] Lactobacillus brevis, with an activity of 10 billion CFU / g. Collection number: SHBCC D14346. Purchased from Shanghai Microbiological Collection Center.

[0019] Bifidobacterium longum, bacterial activity 10 billion CFU / g. Collection number: CCTCC HB 20082718. Purchased from China Center for Type Culture Collection.

[0020] Lactobacillus bulgaricus, bacterial activity 20 billion CFU / g. Collection number: CCTCC CB 20082295; purchased from China Center for Type Culture Collection.

[0021] The inventors found that the antibacterial composition using only modified chitosan as an antibacterial agent is prone to recurrence of cured fungal vaginitis. The inventors compounded plant extracts prepared by extracting a variety of Chinese herbal medicines with modified chitosan as antibacterial components, reducing the recurrence rate of cured fungal vaginitis with the antibacterial composition. The analysis shows that the active components in the plant extracts have high biocompatibility with the modified chitosan, play a synergistic role, and can improve the drug resistance of Candida albicans during the medication process. The combination of plant extracts and modified chitosan can improve the treatment effect and reduce the risk of recurrence.

[0022] Furthermore, the surfactant is selected from one or more of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil, and sodium myristyl alcohol polyether sulfate.

[0023] Furthermore, the surfactant is a compound of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil and sodium myristyl alcohol polyether sulfate in a mass ratio of 1:1.4-1.6:0.3-0.6.

[0024] Sodium cocoamphoacetate, CAS: 68334-21-4.

[0025] Polyoxyethylene ether 40 hydrogenated castor oil, CAS: 61788-85-0.

[0026] Sodium myristeth sulfate, CAS: 25446-80-4.

[0027] After the modified chitosan and the complex active ingredients of the plant extract are mixed in the present invention, the storage stability of the antibacterial composition obtained is not ideal. The present invention improves the storage stability of the antibacterial composition by adding a specific surfactant. The analysis shows that the surfactant of the present invention can adjust the interfacial tension between the components of the system, improve the solubility of the components, form a microscopically stable interface layer, and improve the physical stability of the system. At the same time, the surfactant can form a more stable complex with the components in the system, protect them from environmental factors, and improve the stability during storage.

[0028] The present invention also provides a method for preparing a chitosan reproductive tract biological antibacterial composition, which comprises mixing modified chitosan, plant extracts, glycerol, a surfactant and deionized water, stirring evenly, filtering and sterilizing to obtain the chitosan reproductive tract biological antibacterial composition.

[0029] Compared with the prior art, the advantages and beneficial effects of the present invention are: 1. The present invention modifies chitosan with polyhexamethylene guanidine, introduces amino groups into chitosan, improves the water solubility of chitosan, and further modifies chitosan with cysteine, thereby improving the antibacterial stability of chitosan.

[0030] 2. The present invention combines plant extracts prepared by extracting a variety of Chinese herbal medicines with modified chitosan as antibacterial components, thereby reducing the recurrence rate of candidal vaginitis after the antibacterial composition is cured.

[0031] 3. The present invention improves the storage stability of the antibacterial composition by adding a specific surfactant.

[0032] In summary, the chitosan reproductive tract biological antibacterial composition of the present invention has a single and clear therapeutic purpose, specifically treats candidal vaginitis, has long-lasting antibacterial properties, good therapeutic effects, is not prone to recurrence, has high product stability, and is easy to store. DETAILED DESCRIPTION

[0033] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0034] Example 1: This example provides a chitosan reproductive tract biological antibacterial composition, comprising the following components in percentage by mass: 9% modified chitosan, 5% plant extract, 4% glycerol and 2% surfactant, with the remainder being deionized water.

[0035] The preparation method of the modified chitosan is: (1) mixing chitosan and glacial acetic acid aqueous solution in a mass ratio of 0.2:100, stirring evenly, to obtain a chitosan solution; (2) adding polyhexamethylene guanidine to the chitosan solution, wherein the mass ratio of polyhexamethylene guanidine to chitosan is 0.4:1; reacting at 27° C. for 90 min to obtain a chitosan solution modified with polyhexamethylene guanidine; (3) N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, cysteine ​​and phosphate buffer solution in a mass ratio of 7:5:1:11 were mixed, stirred at 35°C for 3 h to obtain a mixed solution, and the mixed solution was added to a polyhexamethyleneguanidine-modified chitosan solution in a mass ratio of cysteine ​​to chitosan of 0.6:1. The mixture was reacted at 32°C for 43 h to obtain a cysteine-modified chitosan solution; (4) The cysteine-modified chitosan solution is placed in a dialysis bag and dialyzed in deionized water. The molecular weight cutoff of the dialysis bag is 7000Da. The water is changed every 6 hours. After changing the water 7 times, the product in the dialysis bag is frozen and dried to obtain modified chitosan.

[0036] Chitosan, deacetylation degree ≥90%. Zhejiang Jinke Pharmaceutical Co., Ltd., chitosan for medical use.

[0037] Polyhexamethyleneguanidine, Tianjin Liangneng Biotechnology Co., Ltd., PHMG powder.

[0038] N-Hydroxysuccinimide, CAS: 6066-82-6.

[0039] 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, CAS: 25952-53-8.

[0040] Phosphate buffer solution, pH 6.6. Merck, Cat. No. P8165.

[0041] The mass fraction of glacial acetic acid in the glacial acetic acid aqueous solution is 2%.

[0042] The preparation method of the plant extract is: S1: Weigh 12 parts by weight of Smilax glabra, 6 parts by weight of Cimicifuga heracleifolia, 17 parts by weight of Sijiqing, 8 parts by weight of Herba Lysimachiae and 10 parts by weight of Folium Bambusae, mix and grind, and pass through a 60-mesh sieve to obtain a mixed powder; S2: Mix the mixed powder and deionized water in a mass ratio of 1:9, sterilize, and obtain a mixture; mix Lactobacillus brevis, Bifidobacterium longum, and Lactobacillus bulgaricus in a mass ratio of 1:3.4:0.5 to serve as a mixed bacterial agent; add the mixed bacterial agent and the mixture in a mass ratio of 0.2:10 to a fermentation tank, adjust the pH to 6.2, perform anaerobically fermentation at 38°C for 4 days, and sterilize to obtain a fermentation system; S3: The fermentation system is ultrasonicated, and then filtered through a 0.22 μm filter membrane, and then ultrafiltration is performed. The molecular weight cutoff of the ultrafiltration membrane is 20,000 Da. The permeate is concentrated under reduced pressure at 55° C. and dried to obtain a plant extract.

[0043] The ultrasonic conditions are as follows: the ultrasonic device is set in an alternating magnetic field with an intensity of 5200 GS, an ultrasonic time of 90 min, and a frequency of 190 Hz.

[0044] Lactobacillus brevis, with an activity of 10 billion CFU / g. Collection number: SHBCC D14346. Purchased from Shanghai Microbiological Collection Center.

[0045] Bifidobacterium longum, bacterial activity 10 billion CFU / g. Collection number: CCTCC HB 20082718. Purchased from China Center for Type Culture Collection.

[0046] Lactobacillus bulgaricus, bacterial activity 20 billion CFU / g. Collection number: CCTCC CB 20082295; purchased from China Center for Type Culture Collection.

[0047] The surfactant is a compound of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil and sodium myristyl alcohol polyether sulfate in a mass ratio of 1:1.5:0.4.

[0048] Sodium cocoamphoacetate, CAS: 68334-21-4.

[0049] Polyoxyethylene ether 40 hydrogenated castor oil, CAS: 61788-85-0.

[0050] Sodium myristeth sulfate, CAS: 25446-80-4.

[0051] The preparation method of the chitosan reproductive tract biological antibacterial composition comprises the following steps: mixing modified chitosan, plant extract, glycerol, surfactant and deionized water, stirring evenly, filtering and sterilizing to obtain the chitosan reproductive tract biological antibacterial composition.

[0052] Example 2: This example provides a chitosan reproductive tract biological antibacterial composition, comprising the following components in percentage by mass: 7% modified chitosan, 6% plant extract, 3% glycerol and 3% surfactant, with the remainder being deionized water.

[0053] The preparation method of the modified chitosan is: (1) mixing chitosan and glacial acetic acid aqueous solution in a mass ratio of 0.1:100, stirring evenly to obtain a chitosan solution; (2) adding polyhexamethylene guanidine to the chitosan solution, wherein the mass ratio of polyhexamethylene guanidine to chitosan is 0.6:1; reacting at 25° C. for 100 min to obtain a chitosan solution modified with polyhexamethylene guanidine; (3) N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, cysteine ​​and phosphate buffer solution in a mass ratio of 6:7:1:12 were mixed, stirred at 30°C for 4 h to obtain a mixed solution, and the mixed solution was added to a polyhexamethyleneguanidine-modified chitosan solution in a mass ratio of cysteine ​​to chitosan of 0.4:1, and reacted at 35°C for 45 h to obtain a cysteine-modified chitosan solution; (4) The cysteine-modified chitosan solution is placed in a dialysis bag and dialyzed in deionized water. The molecular weight cutoff of the dialysis bag is 7000Da. The water is changed every 6 hours. After changing the water 7 times, the product in the dialysis bag is frozen and dried to obtain modified chitosan.

[0054] Chitosan, deacetylation degree ≥90%. Zhejiang Jinke Pharmaceutical Co., Ltd., chitosan for medical use.

[0055] Polyhexamethyleneguanidine, Tianjin Liangneng Biotechnology Co., Ltd., PHMG powder.

[0056] N-Hydroxysuccinimide, CAS: 6066-82-6.

[0057] 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, CAS: 25952-53-8.

[0058] Phosphate buffer solution, pH 6.6. Merck, Cat. No. P8165.

[0059] The mass fraction of glacial acetic acid in the glacial acetic acid aqueous solution is 1-3%.

[0060] The preparation method of the plant extract is: S1: Weigh 15 parts by weight of Smilax glabra, 3 parts by weight of Cimicifuga heracleifolia, 19 parts by weight of Sijiqing, 5 parts by weight of Herba Lysimachiae and 14 parts by weight of Folium Bambusae, mix and grind, and pass through a 60-mesh sieve to obtain a mixed powder; S2: Mix the mixed powder and deionized water in a mass ratio of 1:10, sterilize, and obtain a mixture; mix Lactobacillus brevis, Bifidobacterium longum, and Lactobacillus bulgaricus in a mass ratio of 1:3.5:0.7 as a mixed bacterial agent; add the mixed bacterial agent and the mixture in a mass ratio of 0.3:10 into a fermentation tank, adjust the pH to 6.1, perform anaerobically fermentation at 40°C for 3 days, sterilize, and obtain a fermentation system; S3: The fermentation system is ultrasonicated, and then filtered through a 0.22 μm filter membrane, and then ultrafiltration is performed. The molecular weight cutoff of the ultrafiltration membrane is 20,000 Da. The permeate is concentrated under reduced pressure at 50° C. and dried to obtain a plant extract.

[0061] The ultrasonic conditions are as follows: the ultrasonic device is set in an alternating magnetic field with an intensity of 5500 GS, an ultrasonic time of 80 min, and a frequency of 200 Hz.

[0062] Lactobacillus brevis, with an activity of 10 billion CFU / g. Collection number: SHBCC D14346. Purchased from Shanghai Microbiological Collection Center.

[0063] Bifidobacterium longum, bacterial activity 10 billion CFU / g. Collection number: CCTCC HB 20082718. Purchased from China Center for Type Culture Collection.

[0064] Lactobacillus bulgaricus, bacterial activity 20 billion CFU / g. Collection number: CCTCC CB 20082295; purchased from China Center for Type Culture Collection.

[0065] The surfactant is a compound of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil and sodium myristyl alcohol polyether sulfate in a mass ratio of 1:1.4:0.6.

[0066] Sodium cocoamphoacetate, CAS: 68334-21-4.

[0067] Polyoxyethylene ether 40 hydrogenated castor oil, CAS: 61788-85-0.

[0068] Sodium myristeth sulfate, CAS: 25446-80-4.

[0069] The preparation method of the chitosan reproductive tract biological antibacterial composition comprises the following steps: mixing modified chitosan, plant extract, glycerol, surfactant and deionized water, stirring evenly, filtering and sterilizing to obtain the chitosan reproductive tract biological antibacterial composition.

[0070] Comparative Example 1 The difference between this comparative example and Example 1 is: a chitosan reproductive tract biological antibacterial composition, comprising the following components in percentage by mass: 14% modified chitosan, 4% glycerol and 2% surfactant, and the balance is deionized water.

[0071] Comparative Example 2 The difference between this comparative example and Example 1 is that the modified chitosan is replaced by carboxymethyl chitosan with a carboxylation degree of 80%. Xi'an Prius Bioengineering Co., Ltd.

[0072] Comparative Example 3 The difference between this comparative example and Example 1 is that the raw material ratios of the Chinese medicine are different.

[0073] Specifically, 6 parts by weight of Smilax glabra, 16 parts by weight of Cimicifuga heracleifolia, 8 parts by weight of Sijiqing, 17 parts by weight of Jixiangcao and 6 parts by weight of Leptochloa chinensis were weighed, mixed, crushed, and passed through a 60-mesh sieve to obtain a mixed powder. Comparative Example 4 The difference between this comparative example and Example 1 is that the types of Chinese medicinal raw materials are different.

[0074] Specifically, 12 parts by mass of polygonatum, 6 parts by mass of sophora flavescens, 17 parts by mass of cnidium monnieri, 8 parts by mass of dandelion and 10 parts by mass of panax notoginseng were weighed, mixed, crushed, and passed through a 60-mesh sieve to obtain a mixed powder.

[0075] Comparative Example 5 The difference between this comparative example and Example 1 is that the ratio of the mixed bacterial agent is different.

[0076] Lactobacillus brevis, Bifidobacterium longum and Lactobacillus bulgaricus were mixed in a mass ratio of 1:1:1 to prepare a mixed bacterial agent.

[0077] Comparative Example 6 The difference between this comparative example and Example 1 is that the types of mixed bacterial agents are different.

[0078] Lactobacillus plantarum, Lactobacillus acidophilus and Lactobacillus casei were mixed in a mass ratio of 1:3.4:0.5 to form a mixed bacterial agent.

[0079] Lactobacillus plantarum, with an activity of 10 billion CFU / g. Collection number: CCTCC AB 2010210. Purchased from China Center for Type Culture Collection.

[0080] Lactobacillus acidophilus, bacterial activity 20 billion CFU / g. Collection number: CCTCC AB 2010208. Purchased from China Center for Type Culture Collection.

[0081] Lactobacillus casei, with an activity of 10 billion CFU / g. Accession number: SHBCC D24737. Purchased from Shanghai Microbiological Collection Center.

[0082] Comparative Example 7 The difference between this comparative example and Example 1 is that the surfactant is a compound of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil and sodium myristyl alcohol polyether sulfate in a mass ratio of 1:1:1.

[0083] Comparative Example 8 The difference between this comparative example and Example 1 is that the surfactant is a compound of Tween 60, Tween 80 and Span 60 in a mass ratio of 1:1.5:0.4.

[0084] Comparative Example 9 The difference between this comparative example and Example 1 is that the preparation method of the modified chitosan is as follows: adding carbodiimide to a citric acid aqueous solution with a pH of 6 for activation for 1 h, adding a chitosan solution, reacting at 4°C for 25 h, precipitating with anhydrous ethanol, washing, and filtering to obtain the modified chitosan; the chitosan solution is a solution formed by dissolving chitosan in 2.5% by mass acetic acid; and the mass ratio of chitosan to citric acid is 1:1.

[0085] Performance Testing The chitosan reproductive tract biological antibacterial compositions prepared in Examples 1-2 and Comparative Examples 1-9 were subjected to performance tests.

[0086] 1. Antibacterial effect According to GB / T15979-2002, the bactericidal rates of Escherichia coli 8099, Staphylococcus aureus ATCC 6538 and Candida albicans ATCC 10231 were tested, and the results are shown in Table 1.

[0087] Table 1 Sterilization rate (%)

[0088] The results show that the chitosan reproductive tract biological antibacterial composition of Examples 1 to 2 has a high antibacterial rate, especially the antibacterial rate against Candida albicans is as high as 99.99%. Comparative Examples 1 to 9 change the preparation method of modified chitosan and plant extracts, which has a greater impact on the antibacterial rate and reduces it to varying degrees.

[0089] 2. Antibacterial stability According to GB / T15979-2002, the bactericidal stability against Escherichia coli 8099, Staphylococcus aureus ATCC 6538 and Candida albicans ATCC 10231 was tested.

[0090] (1) Natural sampling: Place the sealed packaged samples of the chitosan reproductive tract biological antibacterial composition at 25°C for 1 year and measure the sterilization rate.

[0091] (2) Accelerated test: The sealed packaged samples of the chitosan reproductive tract biological antibacterial composition were placed in a constant temperature box at 55°C for 14 days, with a relative humidity of 80%, and the sterilization rate was measured.

[0092] The results are shown in Table 2.

[0093] Table 2 Sterilization rate (%)

[0094] The results show that the chitosan reproductive tract biological antibacterial composition of Examples 1-2 has good antibacterial stability, while Comparative Example 1 without adding plant extracts has poor antibacterial stability, and Comparative Example 2 using carboxymethyl chitosan also has significantly reduced antibacterial stability. Comparative Examples 3-6 and 9 change the plant extracts and modified chitosan, which also reduces the antibacterial stability to varying degrees. Comparative Examples 7-8 show that surfactants can also affect antibacterial stability.

[0095] 3. Clinical trials A total of 440 patients diagnosed with candidal vaginitis were collected and randomly divided into 10 groups, including women aged 22 to 56 years who were sexually active, with an average age of 30.5 years. Excluded were patients with severe liver, kidney, heart, hematopoietic system and nervous system diseases, patients with other infections, patients with a history of drug allergy, patients who used similar drugs for candidal vaginitis within one month before the visit, and pregnant and lactating women.

[0096] Gynecological examination and laboratory secretion microscopy revealed that Candida albicans was positive at the same time. Before going to bed every night, 15 mL of the chitosan reproductive tract biological antibacterial composition of Examples 1-2 and Comparative Examples 1-9 was pushed deep into the vagina for 10 consecutive days as a course of treatment, for a total of 3 courses of treatment. The number of people with three types of efficacy after 3 courses of treatment was counted.

[0097] Efficacy evaluation criteria Cure: The above symptoms completely disappear, the body returns to normal, and there is no recurrence within two years.

[0098] Improvement: The above symptoms disappear, but may recur.

[0099] Uncured: The above symptoms have not changed.

[0100] Adverse reactions: The above symptoms worsen, and the vulva becomes red and swollen.

[0101] Table 3 Clinical results (number of people)

[0102] The results show that the chitosan reproductive tract biological antibacterial composition of Examples 1-2 has a high cure rate and no recurrence. Comparative Example 1 uses only modified chitosan, and has a high recurrence rate. Comparative Example 8 has a high recurrence rate. The analysis shows that only the surfactants of Examples 1-2 of the present invention can improve the wettability of the antibacterial composition to the vaginal mucosal tissue, making it easier for the antibacterial composition to penetrate and distribute to every corner of the vaginal wall, including wrinkles and places where secretions are difficult to remove, so as to more effectively contact pathogenic microorganisms.

[0103] 4. Storage stability test (1) Natural sampling: Place the chitosan reproductive tract biological antibacterial composition transparent sealed packaged sample at 25°C for 1 year and observe whether it is stratified.

[0104] (2) Accelerated test: Place the transparent sealed packaged sample of the chitosan reproductive tract biological antibacterial composition in a constant temperature box at 55°C for 14 days, maintaining a relative humidity of 80%, and observe whether stratification occurs.

[0105] Table 4 Stability test

[0106] The results show that the chitosan reproductive tract biological antibacterial composition of Examples 1 to 2 has high stability. Comparative Example 2 uses carboxymethyl chitosan and Comparative Example 4 uses different Chinese medicine raw materials, and the stability is poor under natural sample retention conditions. Due to the different bacterial agents, the compatibility of the active product and the modified chitosan in Comparative Example 6 is low, and stratification also occurs. Comparative Examples 7 and 8 change the surfactant, and the storage stability decreases overall. Comparative Example 9 uses different modified chitosans, and stratification occurs after storage. Analysis shows that it is due to unsatisfactory compatibility with the system.

[0107] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A chitosan reproductive tract biological antibacterial composition, characterized in that: The invention comprises the following components in percentage by mass: 7-10% of modified chitosan, 3-6% of plant extract, 3-6% of glycerol and 1-3% of surfactant, and the balance is deionized water; The preparation method of the modified chitosan is: (1) Mix chitosan and glacial acetic acid aqueous solution in a mass ratio of 0.1-0.3:100, stir evenly, and obtain a chitosan solution; (2) adding polyhexamethylene guanidine to the chitosan solution, wherein the mass ratio of polyhexamethylene guanidine to chitosan is 0.2-0.6:1; reacting at 25-30° C. for 80-100 min to obtain a chitosan solution modified with polyhexamethylene guanidine; (3) Mix N-hydroxysuccinimide, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, cysteine ​​and phosphate buffer solution in a mass ratio of 6-8:4-7:1:10-12, stir at 30-40°C for 2-4 hours to obtain a mixed solution, add the mixed solution to a polyhexamethyleneguanidine-modified chitosan solution, wherein the mass ratio of cysteine ​​to chitosan is 0.4-0.8:1, and react at 30-35°C for 40-45 hours to obtain a cysteine-modified chitosan solution; (4) The cysteine-modified chitosan solution is placed in a dialysis bag and dialyzed in deionized water. The molecular weight cutoff of the dialysis bag is 7000Da. The water is changed every 6 hours. After changing the water 7 times, the product in the dialysis bag is frozen and dried to obtain modified chitosan.

2. The chitosan reproductive tract biological antibacterial composition according to claim 1, characterized in that: The deacetylation degree of the chitosan is ≥90%.

3. The chitosan reproductive tract biological antibacterial composition according to claim 1, characterized in that: The mass fraction of glacial acetic acid in the glacial acetic acid aqueous solution is 1-3%.

4. The chitosan reproductive tract biological antibacterial composition according to claim 1, characterized in that: The preparation method of the plant extract is: S1: Weigh 10-15 parts by weight of Smilax glabra, 3-8 parts by weight of Cimicifuga heracleifolia, 14-19 parts by weight of Sijiqing, 5-10 parts by weight of Herba Lycopodii and 9-14 parts by weight of Lophatherum gracile, mix and grind, and pass through a 60-mesh sieve to obtain a mixed powder; S2: Mix the mixed powder and deionized water in a mass ratio of 1:8-10, sterilize, and obtain a mixture; mix Lactobacillus brevis, Bifidobacterium longum, and Lactobacillus bulgaricus in a mass ratio of 1:3.2-3.5:0.4-0.7 as a mixed bacterial agent; add the mixed bacterial agent and the mixture in a mass ratio of 0.1-0.3:10 into a fermentation tank, adjust the pH to 6.1-6.3, perform anaerobically fermentation at 36-40°C for 3-5 days, sterilize, and obtain a fermentation system; S3: The fermentation system is subjected to ultrasound, and then filtered through a 0.22 μm filter membrane, and then subjected to ultrafiltration treatment. The molecular weight cutoff of the ultrafiltration membrane is 20,000 Da. The permeate is concentrated under reduced pressure at 50-60° C. and dried to obtain a plant extract.

5. The chitosan reproductive tract biological antibacterial composition according to claim 4, characterized in that: The ultrasonic conditions are as follows: the ultrasonic device is set in an alternating magnetic field, and the strength of the alternating magnetic field is 5000-5500 GS.

6. The chitosan reproductive tract biological antibacterial composition according to claim 4, characterized in that: The ultrasonic time is 80-100min and the frequency is 180-200Hz.

7. The chitosan reproductive tract biological antibacterial composition according to claim 4, characterized in that: Lactobacillus brevis, with an active bacterial count of 10 billion CFU / g; Bifidobacterium longum, with an active bacterial count of 10 billion CFU / g; Lactobacillus bulgaricus, with an active bacterial count of 20 billion CFU / g.

8. The chitosan reproductive tract biological antibacterial composition according to claim 1, characterized in that: The surfactant is selected from one or more of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil, and sodium myristyl alcohol polyether sulfate.

9. The chitosan reproductive tract biological antibacterial composition according to claim 8, characterized in that: The surfactant is a compound of sodium cocoamphoacetate, polyoxyethylene ether 40 hydrogenated castor oil and sodium myristyl alcohol polyether sulfate in a mass ratio of 1:1.4-1.6:0.3-0.

6.

10. A method for preparing the chitosan reproductive tract biological antibacterial composition according to any one of claims 1 to 9, characterized in that: The preparation method comprises the following steps: mixing modified chitosan, plant extract, glycerol, surfactant and deionized water, stirring evenly, filtering and sterilizing to obtain the chitosan reproductive tract biological antibacterial composition.

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