CN114848582A

CN114848582A - Slow-release antibacterial gel for treating female leukoplakia vulvae and preparation method thereof

Info

Publication number: CN114848582A
Application number: CN202210483994.1A
Authority: CN
Inventors: 卞忠华
Original assignee: Jiangsu Chunshentang Biotechnology Co ltd
Current assignee: Jiangsu Chunshentang Pharmaceutical Co ltd
Priority date: 2022-05-05
Filing date: 2022-05-05
Publication date: 2022-08-05
Anticipated expiration: 2042-05-05
Also published as: CN114848582B

Abstract

The invention provides a sustained-release antibacterial gel for treating female leukoplakia vulvae and a preparation method thereof, and relates to the technical field of medical antibacterial gel processing. The slow-release antibacterial gel comprises a hydrogel material and drug-loaded microspheres, wherein the hydrogel material is obtained by mixing oxidized modified hydroxypropyl carboxymethyl cellulose and modified chitosan, and the drug-loaded microspheres are prepared by loading effective drug ingredients into carboxymethyl chitosan. The invention overcomes the defects of the prior art, adopts the gel beads to mix the effective medicinal components and then uses the hydrogel to wrap the effective medicinal components to prepare the medicament release system, effectively achieves the aim of medicament slow release, ensures the long-term curative effect of the medicament, reduces toxic and side reactions, protects the physiological environment of the affected part and improves the use comfort.

Description

Slow-release antibacterial gel for treating female leukoplakia vulvae and preparation method thereof

Technical Field

The invention relates to the technical field of medical antibacterial gel processing, in particular to a sustained-release antibacterial gel for treating female leukoplakia vulvae and a preparation method thereof.

Background

The vulvar dystrophy, also known as white lesions of vulva, non-neoplastic lesions in the vulva epithelium, leukoplakia vulvae, etc., is a chronic and common disease of female vulvar skin and mucosa tissue degeneration and pigment change, which has long course of disease, is difficult to cure and easy to recur, and the pathogenesis of vulvar dystrophy is unknown so far.

At present, the clinical treatment methods for the leukoplakia vulvae comprise drug therapy, ultrasonic therapy, microwave therapy, hyperbaric oxygen therapy, laser therapy, warm acupuncture therapy, surgical treatment and the like. However, most patients can select drugs for treatment in consideration of the comfort, convenience and safety of treatment, and the drugs for treating leukoplakia vulvae generally comprise glucocorticoid, immunomodulator, antitumor drug, sex hormone, traditional Chinese medicine and the like, and generally the drugs can treat the affected part in the form of ointment or lotion.

Most of lotion contacts the affected part with the medicine when being cleaned, the medicine has short retention time, the aim of administration is usually achieved directly by adopting high-concentration medicine, the aim of sterilization treatment is achieved, the generally used mode is repeated for a plurality of times a day, the treatment process is more complicated, the ointment direct-coating treatment is relatively convenient, but the general ointment can damage the genital tract environment and the mucous membrane secretion function after being coated for a long time due to the characteristics of the ointment and the physiological structure of a patient, so that the local immunity of the genital tract is reduced, the body is wounded, the cervical secretion function is damaged, the resistance of the vagina is reduced, and the sexual life and the reproductive health are seriously influenced.

At present, materials such as hydrogel are mostly adopted to be matched with effective medicinal ingredients to treat the affected part, wherein the hydrogel has good biocompatibility, biodegradability and low toxicity, but when the materials are used, a release system of the medicine directly and quickly releases the effective ingredients to treat the affected part, however, a lot of medicines can generate large side effects after being used for a period of time, and can cause the patients to generate drug resistance, so that the treatment effect of the medicine is reduced, and certain troubles are brought to actual clinical treatment.

Disclosure of Invention

Aiming at the defects of the prior art, the invention provides the sustained-release antibacterial gel for treating female leukoplakia vulvae and the preparation method thereof, the gel beads are mixed with effective medicinal ingredients and then the effective medicinal ingredients are wrapped by hydrogel to prepare a medicament release system, so that the purpose of medicament sustained release is effectively achieved, the long-term curative effect of the medicament is ensured, the toxic and side reactions are reduced, the physiological environment of an affected part is protected, and the use comfort is improved.

In order to achieve the above purpose, the technical scheme of the invention is realized by the following technical scheme:

the slow-release antibacterial gel for treating female leukoplakia vulvae comprises a hydrogel material and drug-loaded microspheres, wherein the hydrogel material is obtained by mixing oxidized and modified hydroxypropyl carboxymethyl cellulose and modified chitosan, and the drug-loaded microspheres are prepared by loading effective drug ingredients into carboxymethyl chitosan.

Preferably, the drug-loaded microspheres account for 20% -40% of the hydrogel material content.

Preferably, the effective pharmaceutical ingredient comprises any one or more of glucocorticoid, immunomodulator, sex hormone and the like which can be applied to the treatment of leukoplakia vulvae.

The preparation method of the slow-release antibacterial gel comprises the following steps:

(1) preparation of oxidized hydroxypropyl carboxymethyl cellulose: carrying out oxidation modification on hydroxypropyl carboxymethyl cellulose by using sodium periodate solution to obtain oxidized hydroxypropyl carboxymethyl cellulose for later use;

(2) preparing modified chitosan: mixing chitosan with sulfobetaine for modification to obtain modified chitosan for later use;

(3) preparing hydrogel: mixing the oxidized hydroxypropyl carboxymethyl cellulose and the modified chitosan, adding deionized water, stirring at normal temperature to dissolve, and obtaining hydrogel for later use;

(4) treatment of effective drugs: adding the effective medicinal components for treating leukoplakia vulvae into deionized water and sodium tripolyphosphate, and performing ultrasonic treatment to obtain mixed emulsion for later use;

(5) preparing microspheres: mixing carboxymethyl chitosan with polyvinyl alcohol, then mixing with the mixed emulsion to obtain a mixed solution, adding a silver ion solution and nano zinc oxide into the mixed solution, continuously stirring for precipitation, separating, and freeze-drying the precipitate to obtain microspheres for later use;

(6) preparing an antibacterial gel: and (4) placing the microspheres into the hydrogel obtained in the step (3), and uniformly mixing and stirring to obtain the antibacterial gel.

Preferably, the specific manner of the oxidation modification in the step (1) is as follows: preparing 10% hydroxypropyl carboxymethyl cellulose solution, mixing and stirring uniformly in a dark place according to the volume ratio of the hydroxypropyl carboxymethyl cellulose solution, the sodium periodate solution and the ethylene glycol of 5: 1: 6, dialyzing for 3d by using distilled water, and freeze-drying to obtain the oxidized hydroxypropyl carboxymethyl cellulose.

Preferably, the specific modification mode in the step (2) is as follows: mixing chitosan with 2% acetic acid water solution to prepare 0.1mol/L chitosan solution, adding equimolar sulfobetaine, reacting at 90 deg.C for 24h, precipitating, washing, and lyophilizing to obtain modified chitosan.

Preferably, the content of the mixture of the oxidized hydroxypropyl carboxymethyl cellulose and the modified chitosan in the hydrogel prepared in the step (3) is 2% and 3%.

Preferably, the content of the effective pharmaceutical ingredient in the mixed emulsion obtained in the step (4) is 1%.

Preferably, the volume ratio of the mixture of the carboxymethyl chitosan, the polyvinyl alcohol and the mixed emulsion in the preparation process of the microspheres in the step (5) is 2: 1, and the concentration of the polyvinyl alcohol is 3%.

Preferably, the mass ratio of the mixed solution, the silver ion solution and the nano zinc oxide in the step (5) is 10: 1: 0.3, and the concentration of the silver ions in the silver ion solution is 0.1 mol/L.

The invention provides a sustained-release antibacterial gel for treating female leukoplakia vulvae and a preparation method thereof, and compared with the prior art, the sustained-release antibacterial gel has the advantages that:

(1) the hydrogel microspheres are compounded with effective medicinal ingredients to form a slow-release drug delivery system, so that the sustained-release drug delivery system can be used for continuously and slowly delivering and treating the affected part after being mixed with the hydrogel system, the affected part is prevented from being stimulated by overhigh concentration of the drug, meanwhile, the toxic and side effects of the drug are effectively prevented, and the treatment effect is improved;

(2) according to the invention, the gel microspheres are prepared from carboxymethyl chitosan by adopting an emulsification crosslinking mode, the carboxymethyl chitosan is chelated with metal ions through coordination bonds to form hydrogel in the preparation process, effective medicine components can be effectively combined, the hydrogel microspheres formed in the later stage have a good coating and combining effect on medicines, the hydrogel microspheres mixed with other components after drying have good biocompatibility with the affected part when being coated on the affected part, the medicines are effectively and slowly released after being coated, the uniform and slow-release administration process is achieved, the concentration of the medicines at the affected part is ensured, the problem of uneven administration caused by overhigh or overlow medicine concentration during administration is prevented, the toxic and side effects of the medicines are effectively reduced, the resistance of a human body to the medicines is prevented, and the action effect and the action time of the medicines are improved.

(3) The preparation method adopts the oxidation-modified hydroxypropyl carboxymethyl cellulose and the sulfobetaine-modified chitosan as the preparation materials of the hydrogel, so that the biocompatibility and the swelling property of the hydrogel are effectively improved, the use comfort level is improved, a good drug slow-release system is established in cooperation with hydrogel microbeads, and the hydrogel sustained-release system has a good synergistic sterilization effect, so that a good sterilization and itching-relieving effect is achieved, the physiological balance of affected parts is ensured, and the treatment effect is improved.

Description of the drawings:

FIG. 1 is a schematic diagram of the gelling speed of the antibacterial hydrogel with different microsphere concentrations at different temperatures;

FIG. 2 is a schematic diagram illustrating the sterilization effect of the antimicrobial hydrogel according to the present invention on Staphylococcus aureus with different contact times;

FIG. 3 is a schematic diagram showing the sterilization effect of the antibacterial hydrogel of the present invention on Escherichia coli with different contact times.

Detailed Description

In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention are clearly and completely described below in conjunction with the embodiments of the present invention, and it is obvious that the described embodiments are some embodiments of the present invention, but not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

Example 1:

preparing a hydrogel raw material:

(1) mixing a proper amount of hydroxypropyl carboxymethyl cellulose with distilled water, and uniformly stirring to obtain a hydroxypropyl carboxymethyl cellulose solution with the mass concentration of 10%;

(2) uniformly mixing the hydroxypropyl carboxymethyl cellulose solution and the sodium periodate solution according to the volume ratio of 5: 1 to obtain a mixed solution;

(3) adding equal volume of ethylene glycol into the mixed solution in a dark environment, mixing and stirring uniformly, dialyzing with distilled water for 3 days, and freeze-drying at the low temperature of minus 50 ℃ to obtain the oxidized hydroxypropyl carboxymethyl fiber;

(4) stirring and dissolving a proper amount of chitosan in 2% acetic acid water solution to prepare 0.1mol/L chitosan solution;

(5) adding the chitosan solution into sulfonic acid betaine with equal mole, heating to 90 ℃ in a water bath, mixing and reacting for 24 hours, evaporating to remove part of solvent, mixing with acetone solution, performing flocculation precipitation, washing the precipitate, and freeze-drying at the low temperature of 50 ℃ below zero to obtain modified chitosan;

(6) and (3) mixing and stirring the modified chitosan, the oxidized hydroxypropyl carboxymethyl cellulose and the deionized water at normal temperature according to the mass ratio of 3: 2: 95 to obtain the hydrogel.

Example 2:

(in the actual preparation process, the selection of effective drugs is determined according to actual clinical requirements, and hydrocortisone is used as an experimental detection drug)

Preparation of loaded hydrogel microspheres:

(1) treatment of active ingredients: hydrocortisone butyrate is used as an effective medicinal component, and then deionized water and sodium tripolyphosphate are added for ultrasonic mixing to prepare a mixed emulsion with the concentration of the effective medicinal component of 1% and the concentration of sodium tripolyphosphate of 5%;

(2) uniformly mixing carboxymethyl chitosan, the mixed emulsion and 3% polyvinyl alcohol according to the volume ratio of 2: 1 to obtain a mixed solution;

(3) adding nano zinc oxide into the mixed solution, stirring while adding a silver ion solution with the silver ion concentration of 0.1mol/L, wherein the mass ratio of the nano zinc oxide to the mixed solution to the silver ion solution is 0.3: 1: 10, continuously stirring, separating and precipitating, washing, and freeze-drying the precipitate at the low temperature of minus 50 ℃ to obtain the drug-loaded hydrogel microsphere.

Example 3:

preparation of antibacterial gel:

the drug-loaded hydrogel microspheres in example 2 were added to the hydrogel in example 1 in the proportions of 20%, 30% and 40%, and the mixture was thoroughly mixed and stirred to obtain the antibacterial gel.

And (3) detection:

1. the gelling time of the antibacterial gel in example 3 above at different temperatures was examined:

and (3) gelling time: 20ml of the antibacterial hydrogel added with hydrogel microspheres with different components in the example 3 is sucked into an open glass bottle, the glass bottle is placed at different temperatures, and when the glass bottle shakes, the fluidity of the solution in the bottle is visually observed, so that gel is formed, and the specific results are shown in the following table 1 and the figure 1:

TABLE 1

The above table shows that the antibacterial hydrogel can rapidly form gel texture when the temperature of the antibacterial hydrogel is close to the temperature of a human body, so that the antibacterial hydrogel is convenient to smear and seal when the antibacterial hydrogel is actually used, and the excessive infection of bacteria is prevented.

2. Removing the drug components according to the procedure of the above example 2, preparing hydrogel microspheres containing no effective drug components, mixing the hydrogel microspheres containing no effective drug components with the hydrogel of the example 1 according to the addition amounts of (20%, 30%, 40%) to prepare the drug-free antibacterial gel, and testing the antibacterial performance of the drug-free antibacterial gel:

selecting Staphylococcus aureus (ATCC 6538, gram positive bacteria) and Escherichia coli (ATCC 43895, gram negative bacteria) as test strains, coating the non-drug antibacterial gel on 2.54cm × 2.54cm flat plate, gelatinizing, drying to form membrane, sterilizing, and dripping 25 μ L of pre-cultured bacteria suspension (the concentration of Staphylococcus aureus is 2.33 × 10) into the center of the membrane ⁶ CFU/ml, E.coli concentration 1.47X 10 ⁶ CFU/mL), then covering another film on the film to ensure that the two layers of films are fully contacted with bacteria, taking down the film and transferring the film to a sterile centrifuge tube filled with 5mL of PBS solution after contacting for 10, 20, 30, 40 and 60min respectively, vortexing for 2min, taking 0.1mL of the vortexed solution to perform gradient dilution by using the PBS solution, dropwise adding the diluted bacterial solution into LB culture medium for culture, and culturing for 24h in a 37 ℃ constant temperature incubator. The antibacterial effect was evaluated by counting colonies after completion of the culture, and the specific results are shown in table 2 below and fig. 2 and 3:

TABLE 2

From the above table, the hydrogel and the microspheres in the present invention have good sterilization effects, and the higher the content of the microspheres is, the better the sterilization effect is (the microspheres contain chitosan component and silver ions, and have good sterilization effects).

3. The antibacterial hydrogel with 10%, 20%, 30%, 40%, 50% hydrogel microspheres content prepared according to the above examples 1-3 was used as an experimental group, and the hydrocortisone butyrate cream on the market was used as a control group (hydrocortisone butyrate cream, wherein the hydrocortisone acid content is 0.1%) and physiological saline was used as a blank control group for skin irritation test:

the skin irritation of multiple administrations of the antibacterial hydrogel with different hydrogel microsphere addition amounts is observed by adopting an experimental adult white rabbit:

removing rabbit hair of 5cm × 5cm on both sides of back of adult rabbit, marking 3cm × 3cm range of central mark of hair-removed part as administration region, breeding in suitable environment for 48 hr, and selecting rabbit without erythema edema at hair-shaving position for detection;

selecting seven groups of shaved rabbits, wherein each group comprises three rabbits, smearing 1g of the product of the experimental group and the comparative group in the dosage area on one side of each group of rabbits every 8 hours, smearing the product on the other side of each group of rabbits by using normal saline, cleaning the dosage area by using warm water before each dosage, continuously administering for 72 hours, and observing the dosage position of each group of rabbits, wherein the specific conditions are shown in the following table 3:

TABLE 3

As shown in the table, the antibacterial hydrogel prepared by the invention basically has no irritation to skin, can relieve the irritation of a medicinal component (hydrocortisone butyrate) to skin, and achieves a good relieving effect.

4. Clinical detection shows that the antibacterial hydrogel with 30% of hydrogel microsphere content in the invention and hydrocortisone butyrate cream on the market are used as a comparison group (hydrocortisone butyrate cream is selected, wherein the hydrocortisone acid content is 0.1%) to compare the treatment slow-cross effect on the leukoplakia vulvae:

172 patients with vulvar dystrophy diagnosed by pathological examination in 2019, 4 months to 2019, 11 months were selected, wherein the age range was 36-67 years, the average age was 52 years, 37 patients with squamous epithelial hyperplasia type, 12 patients with lichen sclerosis type and 123 patients with mixed type, all of the patients excluded other lesions, and the 172 patients were randomly divided into a control group (eugler hydrocortisone butyrate cream treatment) and a treatment group (antibacterial hydrogel treatment), wherein the grouping results are as shown in the following table 4:

TABLE 4

The treatment method adopts each group of medicines, the medicines are respectively cleaned in the morning and at night and then coated on the affected part, 3 weeks are a treatment course, the follow-up visit and the re-diagnosis are carried out on the patient after four treatment courses, and the itching symptom, the change of skin physical signs and the structural change condition of the vulva skin under a skin histoscope are used as the evaluation index of the curative effect when the patient is re-diagnosed after four treatment courses.

(healing: the itching disappears completely, the mucous membrane of the skin of the affected part shows normal color, the mucous membrane becomes soft and the skin recovers elasticity; relieving: the itching symptom is controlled, the color and the elasticity of the skin of the affected part are closer to normal compared with those before treatment; ineffective: the itching symptom still exists and the affected part is not changed basically)

The treatment status of the control and treatment groups after four treatment sessions is shown in table 5 below:

TABLE 5

Group of	Cure of disease	Mitigation	Invalidation
				Treatment group	45	39	2
Control group	42	39	5

From the above table, it can be seen that the treatment effect of the treatment group is better than that of the control group, and in the follow-up process, three patients in the control group are inefficiently transferred to other medicines after one week of treatment, one patient in the treatment group is inefficiently transferred to other medicines after two weeks of treatment, and 12 patients in the control group show that the treatment effect of the third and fourth treatment courses is not good as the treatment effect of the first and second treatment courses, but the treatment group does not have such feedback.

In conclusion, the antibacterial gel prepared by the invention can be effectively combined with corresponding medicines to treat leukoplakia vulvae, and has better treatment effect compared with the same type of medicine components.

It is noted that, herein, relational terms such as first and second, and the like may be used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Also, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising an … …" does not exclude the presence of other identical elements in a process, method, article, or apparatus that comprises the element.

The above examples are only intended to illustrate the technical solution of the present invention, but not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions of the embodiments of the present invention.

Claims

1. The slow-release antibacterial gel for treating female leukoplakia vulvae is characterized by comprising a composite composition of a hydrogel material and drug-loaded microspheres, wherein the hydrogel material is obtained by mixing oxidized modified hydroxypropyl carboxymethyl cellulose and modified chitosan, and the drug-loaded microspheres are prepared by loading effective drug ingredients into carboxymethyl chitosan.

2. The sustained-release antibacterial gel for treating female leukoplakia vulvae according to claim 1, which is characterized in that: the drug-loaded microspheres account for 20% -40% of the hydrogel material.

3. The sustained-release antibacterial gel for treating female leukoplakia vulvae according to claim 1, which is characterized in that: the effective medicine component comprises any one or more of glucocorticoid, immunomodulator and sex hormone for treating leukoplakia vulvae.

4. A preparation method of a sustained-release antibacterial gel for treating female leukoplakia vulvae is characterized in that the preparation method of the sustained-release antibacterial gel comprises the following steps:

5. The method for preparing the sustained-release antibacterial gel for treating female leukoplakia vulvae according to claim 4, wherein the oxidation modification in the step (1) is specifically as follows: preparing 10% hydroxypropyl carboxymethyl cellulose solution, mixing and stirring uniformly in a dark place according to the volume ratio of the hydroxypropyl carboxymethyl cellulose solution to the sodium periodate solution to the ethylene glycol of 5: 1: 6, dialyzing for 3d by using distilled water, and freeze-drying to obtain the oxidized hydroxypropyl carboxymethyl cellulose.

6. The method for preparing the slow-release antibacterial gel for treating female leukoplakia vulvae according to claim 4, wherein the modification in the step (2) is as follows: mixing chitosan with 2% acetic acid water solution to prepare 0.1mol/L chitosan solution, adding equimolar sulfobetaine, reacting at 90 deg.C for 24h, precipitating, washing, and lyophilizing to obtain modified chitosan.

7. The method for preparing a sustained-release antibacterial gel for treating female leukoplakia vulvae according to claim 4, wherein the content of the mixture of the oxidized hydroxypropyl carboxymethyl cellulose and the modified chitosan in the hydrogel prepared in the step (3) is 2% and 3%.

8. The method for preparing a sustained-release antibacterial gel for treating leukoplakia vulvae of women as claimed in claim 4, wherein the content of effective pharmaceutical ingredient in the mixed emulsion obtained in the step (4) is 1%.

9. The method for preparing a sustained-release antibacterial gel for treating leukoplakia vulvae of women as claimed in claim 4, wherein the volume ratio of the mixture of the carboxymethyl chitosan, the polyvinyl alcohol and the mixed emulsion in the preparation process of the microspheres in step (5) is 2: 1, and the concentration of the selected polyvinyl alcohol is 3%.

10. The method for preparing a sustained-release antibacterial gel for treating female leukoplakia vulvae according to claim 4, wherein the mass ratio of the mixed solution, the silver ion solution and the nano zinc oxide in the step (5) is 10: 1: 0.3, and the concentration of silver ions in the silver ion solution is 0.1 mol/L.