Antibacterial emulsion for skin and preparation method thereof

Abstract

The invention discloses an antibacterial emulsion for skin and a preparation method thereof, and belongs to the technical field of emulsion medicines. The antibacterial emulsion is prepared from the following raw materials in parts by weight: 3 to 5 parts by weight of modified chitosan, 10 to 20 parts by weight of cross-linked oxidized cyclodextrin inclusion compound, 0.1 to 0.15 part by weight of genipin, 60 to 70 parts by weight of water phase matrix and 10 to 15 parts by weight of oil phase matrix. The antibacterial emulsion prepared by the preparation method disclosed by the invention has a good antibacterial effect and a transdermal effect.

Description

Technical Field

[0001] This invention belongs to the field of emulsion pharmaceutical technology, specifically relating to an antibacterial emulsion for skin and its preparation method. Background Technology

[0002] As the largest organ in the human body, the skin not only has important functions such as protecting internal organs, regulating body temperature, and sensing external stimuli, but also plays a key role in resisting the invasion of pathogenic microorganisms. However, when the skin is damaged or infected, it is often necessary to use antibacterial drugs to assist in treatment. Although antibiotics are widely used in the treatment of skin infections, their abuse and improper use have led to the emergence of drug-resistant bacteria. Drug-resistant bacteria have become a global public health crisis, causing serious consequences such as treatment failure, prolonged course of disease, and increased medical costs. In recent years, with the widespread use of antibiotics, the problem of drug resistance of antibacterial drugs has become increasingly apparent, prompting people to explore new antibacterial treatment methods. Traditional Chinese medicine has unique advantages in antibacterial effects, mainly reflected in the following aspects: (1) Multi-target effect: Traditional Chinese medicine is usually composed of multiple components, which can act on multiple biological pathways at the same time, reducing the risk of bacterial drug resistance; (2) Natural source: Traditional Chinese medicine extracts are derived from natural plants, and their components are complex and diverse, with good biocompatibility and relatively few side effects.

[0003] For skin treatment, traditional Chinese medicine often involves grinding medicinal herbs into powder, mixing them with other base materials to prepare patches, and then applying them to the skin. While this method is simple and easy to implement, it has some drawbacks. For example, manual grinding may result in the loss of some volatile or photosensitive active ingredients in the herbs, affecting the consistency and stability of the efficacy and reducing the therapeutic effect. Patches are adhered to the skin for extended periods, which can lead to local itching due to lack of breathability, affecting the treatment effect. Furthermore, the inconvenience of wearing the patches may impact patient compliance. Additionally, the release of active ingredients from the herbal powder is very slow, resulting in prolonged treatment time and unstable efficacy.

[0004] Traditional Chinese medicine (TCM) extracts are active pharmaceutical ingredients prepared by extracting TCM herbs through methods such as water extraction, alcohol extraction, or ultrasonic extraction to release their active components. This method can improve the stability of the active ingredients and reduce the dosage. However, TCM extracts are often prepared into decoctions, pills, tablets, or capsules for oral treatment, with fewer preparations for skin ointments. This is mainly because the antibacterial active ingredients in TCM extracts are often prematurely oxidized upon contact with air on the skin surface, thus reducing their therapeutic effect. Furthermore, the skin barrier makes drug absorption difficult. Although penetration enhancers such as azone, menthol, and propylene glycol have been developed, the different types and structures of these drugs lead to varying effects on drug penetration, limiting the development of skin emulsions.

[0005] Honeysuckle is a traditional Chinese medicine that is both food and medicine. It is rich in antibacterial and anti-inflammatory active ingredients such as organic acids, flavonoids, and volatile oils. Among them, chlorogenic acid is the most important and abundant antibacterial active ingredient. Currently, honeysuckle is mainly used by decocting it into a soup for oral administration. It is rarely used externally on the skin. This is mainly because the efficacy of topical powder is relatively poor. The active ingredients extracted from honeysuckle contain antibacterial active ingredients with phenolic hydroxyl groups. The antioxidant activity of phenolic hydroxyl groups can consume intracellular ROS to affect the signaling pathways of cell metabolism and induce bacterial apoptosis. However, when extracted and applied to the skin, the phenolic hydroxyl groups are easily oxidized by contact with air, losing their antioxidant activity and thus reducing the antibacterial activity and failing to achieve a long-term therapeutic effect.

[0006] Therefore, it is of great significance to find a way to utilize honeysuckle extract with antibacterial activity to prepare antibacterial emulsions for skin. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention provides an antibacterial emulsion prepared by mixing and emulsifying modified chitosan, cross-linked oxidized cyclodextrin inclusion complex, genipin, an aqueous matrix, and an oil matrix. This emulsion exhibits excellent transdermal and antibacterial effects, thus solving the technical problems mentioned in the background art. Specifically, the technical solution of this invention includes the following: An antibacterial emulsion for skin use, the antibacterial emulsion comprising the following raw materials in parts by weight: 3-5 parts by weight of modified chitosan, 10-20 parts by weight of cross-linked oxidized cyclodextrin inclusion complex, 0.1-0.15 parts by weight of genipin, 60-70 parts by weight of aqueous matrix and 10-15 parts by weight of oil matrix.

[0008] Furthermore, the preparation method of the modified chitosan includes the following steps: Chitosan, aqueous acetic acid solution, and azone were mixed and reacted at 25°C for 4-5 hours to obtain a reaction solution; After the reaction solution and reducing agent are mixed and reacted at 25°C for 2 hours, modified chitosan is obtained by precipitation with anhydrous ethanol and drying. The acidic environment provided by acetic acid is conducive to enhancing the positive charge of the carbonyl carbon in the azone, promoting the affinity attack of the amino group in the chitosan on the carbonyl carbon, and thus generating an imine structure.

[0009] Furthermore, the pH of the acetic acid aqueous solution is 4-5.

[0010] Furthermore, the reducing agent includes sodium borohydride.

[0011] Furthermore, the weight ratio of chitosan: aqueous acetic acid: azone: reducing agent is 1:25~30:0.5~0.6:0.6~0.7.

[0012] Furthermore, the preparation method of the cross-linked oxidized cyclodextrin inclusion complex includes the following steps: Cyclodextrin, deionized water, and sodium periodate were mixed in a weight ratio of 1:15~20:2~3 and reacted at 30℃ for 2~3 hours to obtain oxidized cyclodextrin. Oxidized cyclodextrin, anhydrous ethanol, and ethylenediamine were mixed and dispersed in a weight ratio of 1:10:0.5~0.6 and reacted at 30℃ for 12 h to obtain cross-linked oxidized cyclodextrin. Cross-linked oxidized cyclodextrin, honeysuckle extract and deionized water were mixed in a weight ratio of 5~7:1:40~50 and treated at 30℃~40℃ for 6h~8h, followed by freeze drying to obtain the cross-linked oxidized cyclodextrin inclusion complex.

[0013] Furthermore, the cyclodextrin includes hydroxypropyl-β-cyclodextrin.

[0014] Furthermore, the preparation method of the honeysuckle extract includes the following steps: Honeysuckle powder and deionized water were mixed at a weight ratio of 1:30 and then ultrasonically extracted at 600W~700W ultrasonic power and 50℃ for 30min~40min to obtain the first filtrate and filter residue. The filter residue and deionized water were mixed at a weight ratio of 1:20 and then heated to 80°C for 1 hour to obtain the second filtrate. After the first and second filtrates were combined, they were purified by precipitation with anhydrous ethanol and then centrifuged to obtain a centrifuged liquid. The centrifuged liquid was then evaporated under reduced pressure to obtain the honeysuckle extract.

[0015] Furthermore, the centrifugal separation conditions include a centrifugal speed of 7000 r / min and a centrifugation time of 10 min.

[0016] Furthermore, the conditions for the reduced pressure evaporation include a vacuum of 5 kPa and a temperature of 50°C.

[0017] Furthermore, the aqueous matrix is ​​composed of water and glycerol in a weight ratio of 15:1.

[0018] Furthermore, the oil phase matrix is ​​composed of PEG-8 caprylic / capric triglyceride, ethylhexyl palmitate, and cetearyl alcohol polyether-13 in a weight ratio of 1:1:0.4.

[0019] A method for preparing an antibacterial emulsion for skin use, the method comprising the following steps: After mixing 60-70 parts by weight of aqueous matrix and 10-15 parts by weight of oil matrix, the mixture is heated to 70℃-80℃ and stirred for 3-5 minutes, then cooled to 30℃. Then, 3-5 parts by weight of modified chitosan, 10-20 parts by weight of cross-linked oxidized cyclodextrin inclusion complex and 0.1-0.15 parts by weight of genipin are added, and the mixture is stirred at 500 r / min for 15-20 minutes to obtain the antibacterial emulsion.

[0020] Compared with the prior art, the beneficial effects of the present invention are as follows: This invention first involves multiple solution extractions of honeysuckle to maximize the release of active ingredients into the extraction solution, resulting in an extract. After purification to remove proteins and polysaccharides, the extract is freeze-dried to obtain honeysuckle extract powder. Subsequently, the honeysuckle extract is encapsulated and protected with cyclodextrin, which has a hydrophobic cavity structure, and then prepared as an antibacterial ointment. However, this ointment showed poor transdermal sustained-release and antibacterial effects. Next, the cyclodextrin is oxidized to generate aldehyde groups. Ethylenediamine is then cross-linked with the oxidized cyclodextrin to form imine bonds with pH-responsive hydrolysis cleavage, yielding cross-linked oxidized cyclodextrin. This cross-linked oxidized cyclodextrin inclusion complex is then encapsulated with the honeysuckle extract, allowing for gradual hydrolysis and release of the honeysuckle extract in the weakly acidic environment of the skin. The cross-linked oxidized cyclodextrin inclusion complex exhibits poor antibacterial effect and transdermal penetration due to the low content of active ingredients in the extracted honeysuckle extract and the lack of additional antibacterial substances. Chitosan was then cross-linked with the cross-linked oxidized cyclodextrin inclusion complex using the natural cross-linking agent genipin. Chitosan provides antibacterial activity, synergistically enhancing the antibacterial effect with the honeysuckle extract. However, it was found that the cross-linked mixture tended towards a gel state with poor flowability, failing to form a good emulsion, which was not only unfavorable for subsequent emulsion preparation but also resulted in poor transdermal penetration. Therefore, by condensing the ketone group on the azone structure with the amino group on the chitosan structure to form an imine bond, and then using a reducing agent to reduce the imine bond to a stable secondary amine structure, azone is introduced into chitosan to obtain modified chitosan. The modified chitosan, through the hydrophobic long carbon chain on the azone structure, weakens the crosslinking density during chitosan crosslinking, preventing excessive crosslinking that leads to gelation and is detrimental to emulsion preparation. Furthermore, the introduced azone can synergistically increase the transdermal effect of honeysuckle extract with chitosan. The reduced imine bond stabilizes the binding stability of azone and chitosan, preventing the hydrolysis of the imine bond from increasing the crosslinking density of chitosan with genipin and crosslinked oxidized cyclodextrin inclusion complexes. Modified chitosan, crosslinked oxidized cyclodextrin inclusion complexes, genipin, an aqueous matrix, and an oil matrix are mixed, homogenized, emulsified, and dispersed to obtain an antibacterial emulsion with good transdermal and antibacterial effects. Detailed Implementation

[0021] The technical solution of the present invention will be clearly and completely described below through embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. 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.

[0022] Unless otherwise stated, all raw materials and reagents used in this invention are commercially available or can be prepared by known methods.

[0023] Preparation Example 1 The preparation method of modified chitosan is as follows: Ten parts by weight of chitosan were weighed and dispersed in 250 parts by weight of an acetic acid aqueous solution with a pH of 4. The mixture was stirred until the chitosan was dissolved and evenly dispersed. Then, five parts by weight of azone were added to the dispersed chitosan solution and the mixture was placed in a 25°C environment with stirring at 200 rpm for 4–5 hours. After the reaction, the pH of the resulting reaction solution was adjusted to neutral. Six parts by weight of sodium borohydride were then added, and the mixture was stirred at 200 rpm for another 2 hours at 25°C. After the reaction, anhydrous ethanol was added and stirred to induce precipitation. The precipitate was then centrifuged at 1000 rpm for 10 minutes to obtain the precipitate. The precipitate was repeatedly washed with deionized water until the pH of the washing wastewater reached neutral. Finally, the washed precipitate was dried in a vacuum drying oven at 55°C for 12 hours to obtain modified chitosan.

[0024] Preparation Example 2 The preparation method of modified chitosan is as follows: Weigh 10 parts by weight of chitosan and disperse it in 270 parts by weight of an acetic acid aqueous solution with a pH of 4, stirring until the chitosan is dissolved and evenly dispersed. Then, add 5.5 parts by weight of azone to the dispersed chitosan solution and place it in a 25°C environment, stirring at 200 rpm for 4-5 hours. After the reaction, adjust the pH of the resulting reaction solution to neutral, then add 6.5 parts by weight of sodium borohydride and continue stirring at 200 rpm for 2 hours at 25°C. After the reaction, pour in anhydrous ethanol and stir to form a precipitate. Centrifuge at 1000 rpm for 10 minutes to obtain the precipitate. Wash the precipitate repeatedly with deionized water until the pH of the washing wastewater reaches neutral. Finally, place the washed precipitate in a vacuum drying oven at 55°C and dry for 12 hours to obtain modified chitosan.

[0025] Preparation Example 3 The preparation method of modified chitosan is as follows: Ten parts by weight of chitosan were weighed and dispersed in 300 parts by weight of an acetic acid aqueous solution with a pH of 5. The mixture was stirred until the chitosan was dissolved and evenly dispersed. Then, 6 parts by weight of azone were added to the dispersed chitosan solution and the mixture was placed in a 25°C environment with stirring at 200 rpm for 4–5 hours. After the reaction, the pH of the resulting reaction solution was adjusted to neutral. Then, 7 parts by weight of sodium borohydride were added, and the mixture was stirred at 200 rpm for another 2 hours at 25°C. After the reaction, anhydrous ethanol was added and stirred to induce precipitation. The precipitate was then centrifuged at 1000 rpm for 10 minutes. The precipitate was repeatedly washed with deionized water until the pH of the rinsing wastewater reached neutral. Finally, the washed precipitate was dried in a vacuum drying oven at 55°C for 12 hours to obtain modified chitosan.

[0026] Preparation Example 4 The preparation method of modified chitosan is as follows: Ten parts by weight of chitosan were weighed and dispersed in 300 parts by weight of an aqueous acetic acid solution with a pH of 5. The mixture was stirred until the chitosan was dissolved and evenly dispersed. Then, 6 parts by weight of azone were added to the dispersed chitosan solution and placed in a 25°C environment. The mixture was stirred at 200 rpm for 4–5 hours. After the reaction was complete, anhydrous ethanol was added and stirred to induce precipitation. The precipitate was then centrifuged at 1000 rpm for 10 minutes to obtain the precipitate. The precipitate was repeatedly washed with deionized water until the pH of the washing wastewater reached neutral. Finally, the washed precipitate was dried in a vacuum drying oven at 55°C for 12 hours to obtain modified chitosan.

[0027] Preparation Example 5 The preparation method of modified chitosan is as follows: Ten parts by weight of chitosan were added to 300 parts by weight of deionized water and dispersed evenly using ultrasonic power of 300W. Then, six parts by weight of azone were added to the dispersed chitosan dispersion and placed in a 25°C environment with stirring at 200 rpm for 4-5 hours. After the reaction, the pH of the resulting reaction solution was adjusted to neutral. Then, seven parts by weight of sodium borohydride were added, and the reaction was continued at 25°C with stirring at 200 rpm for 2 hours. After the reaction, anhydrous ethanol was added and stirred to induce precipitation. The precipitate was then centrifuged at 1000 rpm for 10 minutes to obtain the precipitate. The precipitate was repeatedly washed with deionized water until the pH of the washing wastewater reached neutral. Finally, the washed precipitate was dried in a vacuum drying oven at 55°C for 12 hours to obtain modified chitosan.

[0028] Preparation Example 6 The preparation method of honeysuckle extract is as follows: After cleaning the honeysuckle, it was dried in an oven at 50℃ for 24 hours. After cooling to room temperature, it was pulverized and passed through a 50-mesh sieve to obtain honeysuckle powder. 10 parts by weight of honeysuckle powder and 300 parts by weight of deionized water were weighed and mixed thoroughly. The mixture was then placed in a 600W ultrasonic disperser at 50℃ for 30 minutes of ultrasonic extraction. After extraction, the mixture was removed and filtered to obtain the first filtrate and filter residue. 20 times the weight of the filter residue was mixed with deionized water and stirred thoroughly. The mixture was then heated to 80℃ and boiled at this temperature for 1 hour. After this process, the second filtrate and filter residue were obtained by filtration. The first and second filtrates were combined, and 2 times the weight of the mixed filtrate was added to anhydrous ethanol. The mixture was stirred at 100 rpm for 30 minutes and then allowed to stand for 1 hour. After settling, the mixture was centrifuged at 7000 r / min for 10 min to obtain a centrifuged liquid. The centrifuged liquid was then placed in a rotary evaporator flask with a vacuum of 5 kPa and the temperature was controlled at 50℃ for vacuum evaporation and drying until the weight remained constant, thus obtaining honeysuckle extract. The chlorogenic acid content in the honeysuckle extract was determined to be 19.04 mg / g by high performance liquid chromatography (column temperature 25℃, mobile phase composed of acetonitrile, water and glacial acetic acid in a volume ratio of 15:85:0.2, flow rate 0.8 mL / min, detection wavelength 327 nm).

[0029] Preparation Example 7 The preparation method of honeysuckle extract is as follows: After cleaning the honeysuckle, it was dried in an oven at 50℃ for 24 hours. After cooling to room temperature, it was pulverized using a pulverizer. The pulverized particles were passed through a 50-mesh sieve to obtain honeysuckle powder. 10 parts by weight of honeysuckle powder and 300 parts by weight of deionized water were weighed and mixed thoroughly. The mixture was then placed in a 600W ultrasonic disperser at 50℃ for a random ultrasonic extraction process of up to 40 minutes. After extraction, the mixture was removed and filtered to obtain the first filtrate and filter residue. 20 times the weight of the filter residue was added to the deionized water and mixed thoroughly. The mixture was then heated to 80℃ and boiled at this temperature for 1 hour. After this process, the second filtrate and filter residue were obtained by filtration. The first and second filtrates were combined, and 2 times the weight of the mixed filtrate was added to anhydrous ethanol. The mixture was stirred at 100 rpm for 30 minutes and then allowed to stand for 1 hour. After settling, the mixture was centrifuged at 7000 r / min for 10 min to obtain a centrifuged liquid. The centrifuged liquid was then placed in a rotary evaporator flask with a vacuum of 5 kPa and the temperature was controlled at 50℃ for vacuum evaporation and drying until the weight remained constant, thus obtaining honeysuckle extract. The chlorogenic acid content in the honeysuckle extract was determined to be 20.11 mg / g by high performance liquid chromatography (column temperature 25℃, mobile phase composed of acetonitrile, water and glacial acetic acid in a volume ratio of 15:85:0.2, flow rate 0.8 mL / min, detection wavelength 327 nm).

[0030] Preparation Example 8 The preparation method of honeysuckle extract is as follows: After cleaning the honeysuckle, it was dried in an oven at 50℃ for 24 hours. After cooling to room temperature, it was pulverized using a pulverizer. The pulverized particles were passed through a 50-mesh sieve to obtain honeysuckle powder. 10 parts by weight of honeysuckle powder and 300 parts by weight of deionized water were weighed and mixed thoroughly. The mixture was then placed in a 700W ultrasonic disperser at 50℃ for a random ultrasonic extraction process of up to 40 minutes. After extraction, the mixture was removed and filtered to obtain the first filtrate and filter residue. 20 times the weight of the filter residue was added to the deionized water and mixed thoroughly. The mixture was then heated to 80℃ and boiled at this temperature for 1 hour. After this process, the second filtrate and filter residue were obtained by filtration. The first and second filtrates were combined, and 2 times the weight of the mixed filtrate was added to anhydrous ethanol. The mixture was stirred at 100 rpm for 30 minutes and then allowed to stand for 1 hour. After settling, the mixture was centrifuged at 7000 r / min for 10 min to obtain a centrifuged liquid. The centrifuged liquid was then placed in a rotary evaporator flask with a vacuum of 5 kPa and the temperature was controlled at 50℃ for vacuum evaporation and drying until the weight remained constant, thus obtaining honeysuckle extract. The chlorogenic acid content in the honeysuckle extract was determined to be 20.45 mg / g by high performance liquid chromatography (column temperature 25℃, mobile phase composed of acetonitrile, water and glacial acetic acid in a volume ratio of 15:85:0.2, flow rate 0.8 mL / min, detection wavelength 327 nm).

[0031] Preparation Example 9 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: 10 parts by weight of hydroxypropyl-β-cyclodextrin and 150 parts by weight of deionized water were weighed and added to a flask, mixed and stirred until completely dissolved and evenly dispersed. Then, 20 parts by weight of sodium periodate were added, mixed and stirred evenly, and the mixture was protected from light. The temperature was then controlled at 30°C, and the oxidation reaction was timed at this temperature for 2 hours. After the reaction was completed, 40 parts by weight of ethylene glycol were added to quench the oxidative catalysis of sodium periodate. The resulting reaction mixture was poured into a dialysis bag with a molecular weight cutoff of 1 kDa, and dialyzed with distilled water for 2 days. The solution in the dialysis bag was then placed in a rotary evaporator flask with a vacuum degree of 5 kPa, and the temperature was controlled at 50°C for vacuum evaporation and drying until the weight remained constant, yielding oxidized cyclodextrin. 1 part by weight of oxidized cyclodextrin, 10 parts by weight of anhydrous ethanol, and 0.5 parts by weight of ethylenediamine were weighed and mixed together, and dispersed using 200 W of ultrasonic power for 20 minutes to ensure even dispersion. The mixture was then reacted in a water bath at 30°C for 12 hours. After the reaction was completed, the mixture was cooled to 5°C and allowed to stand until precipitation occurred. The solid was then collected by filtration, washed with anhydrous ethanol, and finally dried in a vacuum drying oven at 55°C for 12 hours to obtain cross-linked oxidized cyclodextrin. Five parts by weight of cross-linked oxidized cyclodextrin, one part by weight of honeysuckle extract obtained in Preparation Example 6, and 40 parts by weight of deionized water were weighed and mixed. The mixture was then placed in a water bath at 30°C and stirred at 200 r / min for 6 hours to obtain an aqueous dispersion of cyclodextrin inclusion complex. This dispersion was then freeze-dried at -50°C to obtain the cross-linked oxidized cyclodextrin inclusion complex.

[0032] Preparation Example 10 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: 10 parts by weight of hydroxypropyl-β-cyclodextrin and 180 parts by weight of deionized water were weighed and added to a flask, mixed and stirred until completely dissolved and evenly dispersed. Then, 25 parts by weight of sodium periodate were added, mixed and stirred evenly, and the mixture was protected from light. The temperature was then controlled at 30°C, and the oxidation reaction was timed at this temperature for 2.5 hours. After the reaction was completed, 50 parts by weight of ethylene glycol were added to quench the oxidative catalytic effect of sodium periodate. The resulting reaction mixture was poured into a dialysis bag with a molecular weight cutoff of 1 kDa, and dialyzed with distilled water for 2 days. The solution in the dialysis bag was then placed in a rotary evaporator flask with a vacuum degree of 5 kPa, and the temperature was controlled at 50°C for vacuum evaporation and drying until the weight remained constant, yielding oxidized cyclodextrin. 1 part by weight of oxidized cyclodextrin, 10 parts by weight of anhydrous ethanol, and 0.55 parts by weight of ethylenediamine were weighed and mixed together, and dispersed using an ultrasonic power of 200 W for 20 minutes to ensure even dispersion. The mixture was then reacted in a water bath at 30°C for 12 hours. After the reaction was completed, the mixture was cooled to 5°C and allowed to stand until precipitation occurred. The solid was then collected by filtration, washed with anhydrous ethanol, and finally dried in a vacuum drying oven at 55°C for 12 hours to obtain cross-linked oxidized cyclodextrin. Six parts by weight of cross-linked oxidized cyclodextrin, one part by weight of honeysuckle extract obtained in Preparation Example 7, and 45 parts by weight of deionized water were weighed and mixed. The mixture was then placed in a water bath at 35°C and stirred at 200 r / min for 7 hours to obtain an aqueous dispersion of cyclodextrin inclusion complex. This dispersion was then freeze-dried at -50°C to obtain the cross-linked oxidized cyclodextrin inclusion complex.

[0033] Preparation Example 11 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: 10 parts by weight of hydroxypropyl-β-cyclodextrin and 200 parts by weight of deionized water were weighed and added to a flask, mixed and stirred until completely dissolved and evenly dispersed. Then, 30 parts by weight of sodium periodate were added, mixed and stirred evenly, and the mixture was protected from light. The temperature was then controlled at 30°C, and the oxidation reaction was timed at this temperature for 3 hours. After the reaction was completed, 60 parts by weight of ethylene glycol were added to quench the oxidative catalysis of sodium periodate. The resulting reaction mixture was poured into a dialysis bag with a molecular weight cutoff of 1 kDa, and dialyzed with distilled water for 2 days. The solution in the dialysis bag was then placed in a rotary evaporator flask with a vacuum degree of 5 kPa, and the temperature was controlled at 50°C for vacuum evaporation and drying until the weight remained constant, yielding oxidized cyclodextrin. 1 part by weight of oxidized cyclodextrin, 10 parts by weight of anhydrous ethanol, and 0.6 parts by weight of ethylenediamine were weighed and mixed together, and dispersed using 200 W of ultrasonic power for 20 minutes to ensure even dispersion. The mixture was then reacted in a water bath at 30°C for 12 hours. After the reaction was completed, the mixture was cooled to 5°C and allowed to stand until precipitation occurred. The solid was then collected by filtration, washed with anhydrous ethanol, and finally dried in a vacuum drying oven at 55°C for 12 hours to obtain cross-linked oxidized cyclodextrin. Seven parts by weight of cross-linked oxidized cyclodextrin, one part by weight of honeysuckle extract obtained in Preparation Example 8, and 50 parts by weight of deionized water were weighed and mixed. The mixture was then placed in a water bath at 40°C and stirred at 200 r / min for 8 hours to obtain an aqueous dispersion of cyclodextrin inclusion complex. This dispersion was then freeze-dried at -50°C to obtain the cross-linked oxidized cyclodextrin inclusion complex.

[0034] Preparation Example 12 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: Weigh 10 parts by weight of hydroxypropyl-β-cyclodextrin and 200 parts by weight of deionized water and add them to a flask. Mix and stir until completely dissolved and evenly dispersed. Then add 40 parts by weight of sodium periodate, mix and stir evenly, and then protect from light. Then control the temperature at 30°C and perform an oxidation reaction at this temperature for 4 hours. The rest of the preparation process is the same as in Preparation Example 11.

[0035] Preparation Example 13 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: In Preparation Example 11, hydroxypropyl-β-cyclodextrin was replaced with β-cyclodextrin, and the rest of the preparation process remained the same as in Preparation Example 11.

[0036] Preparation Example 14 The preparation method of cross-linked oxidized cyclodextrin inclusion complex is as follows: In Preparation Example 11, ethylenediamine was replaced with 1,6-hexanediamine, and the rest of the preparation process remained the same as in Preparation Example 11.

[0037] Preparation Example 15 The preparation method of cyclodextrin inclusion complex is as follows: Weigh 7 parts by weight of hydroxypropyl-β-cyclodextrin, 1 part by weight of honeysuckle extract obtained in Preparation Example 8, and 50 parts by weight of deionized water, mix them, and then place them in a water bath at 40°C. Stir at 200 r / min for 8 h to obtain an aqueous dispersion of cyclodextrin inclusion complex. Then, freeze dry the dispersion at -50°C to obtain the cyclodextrin inclusion complex.

[0038] Example 1 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: Weigh 60 parts by weight of the aqueous phase matrix (composed of deionized water and glycerol in a weight ratio of 15:1) and 10 parts by weight of the oil phase matrix (composed of PEG-8 caprylic / capric glyceride, ethylhexyl palmitate and cetearyl alcohol polyether-13 in a weight ratio of 1:1:0.4), mix them, and then heat to 70°C. Stir at 100 r / min for 3 min at this temperature, and then cool to 30°C. Add 3 parts by weight of the modified chitosan obtained in Preparation Example 1, 10 parts by weight of the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 9 and 0.1 parts by weight of genipin at this temperature. Then increase the speed to 500 r / min and stir for 15 min to obtain the antibacterial emulsion.

[0039] Example 2 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: Weigh 65 parts by weight of the aqueous phase matrix (composed of deionized water and glycerol in a weight ratio of 15:1) and 12 parts by weight of the oil phase matrix (composed of PEG-8 caprylic / capric glyceride, ethylhexyl palmitate and cetearyl alcohol polyether-13 in a weight ratio of 1:1:0.4), mix them, and then heat to 80°C. Stir at 100 r / min for 4 min at this temperature, and then cool to 30°C. At this temperature, add 4 parts by weight of the modified chitosan obtained in Preparation Example 2, 15 parts by weight of the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 10, and 0.13 parts by weight of genipin. Then increase the speed to 500 r / min and stir for 18 min to obtain the antibacterial emulsion.

[0040] Example 3 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: Weigh 70 parts by weight of the aqueous phase matrix (composed of deionized water and glycerol in a weight ratio of 15:1) and 15 parts by weight of the oil phase matrix (composed of PEG-8 caprylic / capric glyceride, ethylhexyl palmitate and cetearyl alcohol polyether-13 in a weight ratio of 1:1:0.4), mix them, and then heat to 80°C. Stir at 100 r / min for 5 min at this temperature, and then cool to 30°C. Add 5 parts by weight of the modified chitosan obtained in Preparation Example 3, 20 parts by weight of the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 11 and 0.15 parts by weight of genipin at this temperature, and then increase the speed to 500 r / min and stir for 20 min to obtain the antibacterial emulsion.

[0041] Comparative Example 1 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The modified chitosan in Example 3 was replaced with the modified chitosan obtained in Preparation Example 4, and the rest of the preparation process was the same as in Example 3.

[0042] Comparative Example 2 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The modified chitosan in Example 3 was replaced with the modified chitosan obtained in Preparation Example 5, and the rest of the preparation process was the same as in Example 3.

[0043] Comparative Example 3 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The cross-linked oxidized cyclodextrin inclusion complex in Example 3 was replaced with the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 12, and the rest of the preparation process was the same as in Example 3.

[0044] Comparative Example 4 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The cross-linked oxidized cyclodextrin inclusion complex in Example 3 was replaced with the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 13, and the rest of the preparation process was the same as in Example 3.

[0045] Comparative Example 5 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The cross-linked oxidized cyclodextrin inclusion complex in Example 3 was replaced with the cross-linked oxidized cyclodextrin inclusion complex obtained in Preparation Example 14, and the rest of the preparation process was the same as in Example 3.

[0046] Comparative Example 6 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: Weigh 70 parts by weight of the aqueous phase matrix (composed of deionized water and glycerol in a weight ratio of 15:1) and 15 parts by weight of the oil phase matrix (composed of PEG-8 caprylic / capric glyceride, ethylhexyl palmitate and cetearyl alcohol polyether-13 in a weight ratio of 1:1:0.4), mix them, then heat to 80°C, stir at 100 r / min for 5 min at this temperature, then cool to 30°C, add 20 parts by weight of the cyclodextrin inclusion complex obtained in Preparation Example 15 at this temperature, then increase the stirring speed to 500 r / min and stir for 20 min to obtain the antibacterial emulsion.

[0047] Comparative Example 7 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The modified chitosan and genipin in Example 3 were removed and not added, while the rest of the preparation process remained the same as in Example 3.

[0048] Comparative Example 8 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: Replacing the modified chitosan in Example 3 with chitosan and maintaining the remaining preparation process as in Example 3, it was found that the emulsion's fluidity gradually decreased during mixing and emulsification, and gel solidification occurred after a period of time, resulting in preparation failure. This may be because chitosan easily gels through cross-linking interactions such as intermolecular hydrogen bonds. Without treatment, excessive cross-linking can easily lead to gel solidification, which is detrimental to emulsion preparation.

[0049] Comparative Example 9 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: In Example 3, the modified chitosan was increased to 10 parts by weight, while the rest of the preparation process remained the same as in Example 3. It was found that during the mixing and emulsification process, the emulsion's fluidity gradually decreased, and after a period of time, gel solidification was observed, resulting in preparation failure. This may be because the steric hindrance of azone can weaken the cross-linking strength of chitosan, thereby preventing gel solidification. However, in this preparation method, if too much modified chitosan is used, it is equivalent to increasing the concentration of the substance, which increases the probability of contact between substances and may instead cause stronger cross-linking, which is detrimental to emulsion preparation.

[0050] Comparative Example 10 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The modified chitosan and genipin in Example 3 were removed and not added, and the amount of cross-linked oxidized cyclodextrin inclusion complex was increased to 30 parts by weight. The rest of the preparation process was the same as in Example 3.

[0051] Comparative Example 11 A method for preparing an antibacterial emulsion for skin use, specifically including the following steps: The modified chitosan and genipin in Example 3 were removed and not added. The amount of cross-linked oxidized cyclodextrin inclusion complex was increased to 30 parts by weight, and an additional 3 parts by weight of azone were added. The rest of the preparation process was the same as in Example 3.

[0052] (1) In vitro transdermal test: Healthy male mice were euthanized by cervical dislocation. Hair was removed, and the abdomen was inflated until taut. After peeling off the abdominal skin, subcutaneous adipose tissue and connective tissue were cleaned with cotton balls soaked in physiological saline. The mice were then repeatedly rinsed with distilled water and physiological saline, dried with non-woven fabric, and cut into 2.5cm × 2.5cm pieces. These pieces were then fixed in a transdermal diffusion apparatus with the stratum corneum facing upwards, ensuring complete contact between the dermis and the liquid in the receiving tank (7mL volume). The water bath temperature was set to 32℃, and the stirring speed was 200 rpm. Physiological saline at 32℃ was added to the receiving tank to remove air bubbles. Then, 2mL of the antibacterial emulsions obtained in Examples 1-3, Comparative Examples 1-7, and Comparative Examples 10-11 were applied to the skin, covering an area of ​​2.2cm². 2 At 2h, 4h, 6h, 12h, and 24h, 2mL of receiving liquid was drawn from the receiving cell into centrifuge tubes for high-performance liquid chromatography (HPLC) analysis, according to the cumulative transmittance P=Q. n / M calculates the cumulative transmittance of chlorogenic acid at each time point, where Q n The calculation formula is as follows: Where Cn is the chlorogenic acid content at the nth sampling, V is the volume of the receiving cell, Ci is the chlorogenic acid content at the ith sampling, Vi is the sampling volume at the ith sampling, A is the area of ​​the skin covered with the drug, and M is the chlorogenic acid content. The results are shown in Table 1 below.

[0053] Table 1. Cumulative Transmittance (%) Source of materials 2h 4h 6h 12h 24h Example 1 8.13 13.32 21.94 44.37 70.11 Example 2 7.54 12.94 21.06 42.87 69.03 Example 3 6.12 12.75 20.41 42.05 68.27 Comparative Example 1 1.84 2.76 3.72 12.71 26.59 Comparative Example 2 1.27 2.03 3.22 11.48 23.91 Comparative Example 3 6.33 13.11 21.31 42.91 69.05 Comparative Example 4 1.91 2.82 3.83 12.82 26.74 Comparative Example 5 6.24 12.97 20.81 42.46 68.82 Comparative Example 6 2.21 2.98 4.24 13.75 27.92 Comparative Example 7 1.96 2.84 3.88 12.89 26.82 Comparative Example 10 2.12 2.91 4.03 13.21 27.31 Comparative Example 11 2.27 3.03 4.75 14.01 28.32 The antibacterial emulsions obtained in Examples 1-3, Comparative Examples 1-7, and Comparative Examples 10-11 were mixed with an equal volume of an initial viable count of 1×10⁻⁶ cells. 7 ~1×10 8 After mixing the Staphylococcus aureus culture medium at CFU / mL, the mixture was placed in a constant temperature incubator at 37℃ for 24 hours. The remaining viable bacteria number was then recorded by plate counting and the sterilization rate was calculated. Sterilization rate = (initial viable bacteria number - remaining viable bacteria number) / initial viable bacteria number × 100%. The results are shown in Table 2 below.

[0054] Table 2 Antibacterial effect Source of materials Sterilization rate (%) Example 1 90.69 Example 2 93.17 Example 3 94.23 Comparative Example 1 94.08 Comparative Example 2 94.11 Comparative Example 3 46.13 Comparative Example 4 93.97 Comparative Example 5 52.66 Comparative Example 6 35.41 Comparative Example 7 31.26 Comparative Example 10 94.02 Comparative Example 11 93.91 Based on Tables 1 and 2 above, the following conclusions can be drawn: (1) As can be seen from Examples 1 to 3, the antibacterial emulsion prepared by the preparation method of the present invention has good transdermal effect and antibacterial effect.

[0055] (2) Comparative Example 1 shows that the prepared antibacterial emulsion has good antibacterial effect, but poor transdermal effect. This may be because chitosan forms an imine bond through the condensation of amino groups and carbonyl groups on azone. If the imine bond is not further reduced to a stable secondary amine structure, the imine bond will gradually break and separate due to hydrolysis in the weakly acidic environment of the skin. As time goes on, the transdermal effect becomes worse and worse, which is not conducive to the long-term treatment of the antibacterial emulsion.

[0056] (3) Comparative Example 2 shows that the transdermal effect of the prepared antibacterial emulsion is poor. This may be because although chitosan has an amino group in its structure, it can condense with the carbonyl group on azone to form an imine bond and be further reduced to a secondary amine structure by a reducing agent. However, the carbonyl group on the azone molecule is a hydrophobic long carbon chain with twelve carbon atoms, which has a strong steric hindrance effect. If the carbonyl carbon on the azone is not treated with acid to increase its positive charge, chitosan and azone may not be able to react and connect in this system. Relying solely on the penetration-promoting effect of azone and chitosan, the penetration-promoting effect of honeysuckle extract in this system is poor, which is not conducive to the long-term treatment of antibacterial emulsion.

[0057] (4) Comparative Example 3 shows that the antibacterial emulsion prepared has poor antibacterial effect. This may be because the excessive use of sodium periodate as an oxidant in this system may lead to excessive oxidation and destruction of hydroxypropyl-β-cyclodextrin, which may affect the hydrophobic cavity structure of hydroxypropyl-β-cyclodextrin, resulting in a poorer encapsulation effect on honeysuckle extract, and thus a poor antibacterial effect of the final antibacterial emulsion.

[0058] (5) Comparative Example 4 shows that the transdermal effect of the prepared antibacterial emulsion is poor. This may be because β-cyclodextrin has poor hydrophilicity compared to hydroxypropyl-β-cyclodextrin in this system, which may hinder the hydration of the skin and make it difficult for the drug to be released through the skin barrier into the body.

[0059] (6) Comparative Example 5 shows that the antibacterial emulsion prepared has poor antibacterial effect. This may be because the carbon chain of 1,6-hexanediamine is too long in this system, which affects the hydrophilic properties of the outer surface of cyclodextrin, which may competitively affect the encapsulation of honeysuckle extract by hydrophobic cavity, and the encapsulation effect may be worse, resulting in a weaker antibacterial effect.

[0060] (7) Comparative Examples 6 and 7 show that the transdermal and antibacterial effects of the prepared antibacterial emulsions are poor. This may be because, in this system, on the one hand, the content of active ingredients in honeysuckle extract is low and the loading of hydroxypropyl-β-cyclodextrin is limited. Without the introduction of other antibacterial substances, the antibacterial effect is weak. On the other hand, relying solely on the encapsulation effect of hydroxypropyl-β-cyclodextrin, although hydroxypropyl-β-cyclodextrin has a certain degree of permeation-promoting effect, its permeation-promoting effect on the drug is obviously poor under the preparation method of this system.

[0061] (8) Comparative Example 10 shows that although the antibacterial emulsion prepared has a good antibacterial effect, the transdermal effect is poor. This may be because, although hydroxypropyl-β-cyclodextrin has a certain degree of penetration-enhancing effect in this system, the preparation method of this system relies solely on hydroxypropyl-β-cyclodextrin, which is obviously not suitable for improving the penetration-enhancing effect of drugs.

[0062] (9) Comparative Example 11 shows that although the antibacterial emulsion prepared has a good antibacterial effect, the transdermal effect is poor. This may be because although azone is a commonly used penetration enhancer in this system, not all penetration enhancers can achieve a good penetration enhancement effect under the preparation method of this system. It may be necessary to adjust the penetration enhancer in order to play an auxiliary penetration enhancement role.

[0063] The embodiments described above provide a detailed explanation of the technical solutions and beneficial effects of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Various changes and modifications can be made to the present invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed.

Claims

1. An antibacterial emulsion for skin use, characterized in that, The antibacterial emulsion agent comprises the following raw materials by weight: 3-5 parts by weight of modified chitosan, 10-20 parts by weight of cross-linked oxidized cyclodextrin inclusion compound, 0.1-0.15 parts by weight of genipin, 60-70 parts by weight of aqueous phase matrix and 10-15 parts by weight of oil phase matrix.

2. The antibacterial emulsion agent for skin according to claim 1, characterized by, The preparation method of the modified chitosan comprises the following steps: Chitosan, aqueous acetic acid solution and azone are mixed and reacted at 25℃ for 4-5 hours to obtain a reaction solution; After the reaction solution and a reducing agent are mixed and reacted at 25℃ for 2 hours, the modified chitosan is obtained by precipitation and drying with anhydrous ethanol.

3. The antibacterial emulsion for skin use according to claim 2, characterized by, The reducing agent comprises sodium borohydride.

4. The antibacterial emulsion agent for skin according to claim 2, characterized by, The weight ratio of chitosan, aqueous acetic acid solution, azone and reducing agent is 1:25-30:0.5-0.6:0.6-0.

7.

5. The antibacterial emulsion agent for skin according to claim 1, wherein The preparation method of the cross-linked oxidized cyclodextrin inclusion compound comprises the following steps: Cyclodextrin, deionized water and sodium periodate are mixed at a weight ratio of 1:15-20:2-3 and reacted at 30℃ for 2-3 hours to obtain oxidized cyclodextrin; The oxidized cyclodextrin, anhydrous ethanol and ethylenediamine are mixed and dispersed at a weight ratio of 1:10:0.5-0.6, and then reacted at 30℃ for 12 hours to obtain cross-linked oxidized cyclodextrin; The cross-linked oxidized cyclodextrin, honeysuckle extract and deionized water are mixed at a weight ratio of 5-7:1:40-50, and then treated at 30-40℃ for 6-8 hours, and then freeze-dried to obtain the cross-linked oxidized cyclodextrin inclusion compound.

6. The antibacterial emulsion for skin use according to claim 5, wherein The cyclodextrin comprises hydroxypropyl-β-cyclodextrin.

7. The antibacterial emulsion agent for skin according to claim 5, wherein The preparation method of the honeysuckle extract comprises the following steps: Honeysuckle powder and deionized water are mixed at a weight ratio of 1:30, and then ultrasonic extraction is performed at 600-700W ultrasonic power and 50℃ for 30-40 minutes to obtain a first filtrate and a filter residue; The filter residue and deionized water are mixed at a weight ratio of 1:20, and then heated to 80℃ for 1 hour to obtain a second filtrate; The first filtrate and the second filtrate are combined, impurities are removed by precipitation with anhydrous ethanol, and then centrifuged to obtain a centrifugal liquid, and the centrifugal liquid is evaporated under reduced pressure to obtain the honeysuckle extract.

8. The antibacterial emulsion agent for skin according to claim 1, wherein The aqueous phase matrix is composed of water and glycerol at a weight ratio of 15:

1.

9. The antibacterial emulsion agent for skin according to claim 1, wherein The oil phase matrix is composed of PEG-8 caprylic / capric glycerides, ethylhexyl palmitate and ceteareth-13 at a weight ratio of 1:1:0.

4.

10. A process for the preparation of an antibacterial emulsion for skin use according to any one of claims 1 to 9, characterized in that The preparation method comprises the following steps: 60-70 parts by weight of aqueous phase matrix and 10-15 parts by weight of oil phase matrix are mixed, heated to 70-80℃, stirred for 3-5 minutes, cooled to 30℃, and then 3-5 parts by weight of modified chitosan, 10-20 parts by weight of cross-linked oxidized cyclodextrin inclusion compound and 0.1-0.15 parts by weight of genipin are added, and stirred at a speed of 500r / min for 15-20 minutes to obtain the antibacterial emulsion agent.

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