Zedoary turmeric extract / cystose and preparation method thereof

Abstract

The application discloses a kind of Zaijieliu extract / chitosan composite nanometer film and preparation method thereof.The method uses traditional Chinese medicine Zaijieliu extract as raw material, carboxymethyl chitosan as film forming agent, and aldehyde sodium alginate as microcoagulant, to prepare Zaijieliu extract / chitosan composite nanometer film by combining film casting technology and solvent evaporation method.The preparation method is simple, low in cost, avoids the use of toxic chemical crosslinking agent, and the prepared Zaijieliu extract / chitosan composite nanometer film material has good mechanical properties, hydrophilicity, stability and drug release performance, and has wide application prospect in wound repair, drug controlled release and tissue engineering.

Description

Technical Field

[0001] This invention belongs to the field of biomedical materials and relates to a fluid extract / chitosan composite nanofilm and its preparation method. Background Technology

[0002] For tuberculous sinus tracts, traditional Chinese medicine currently believes that treatment should combine internal and external therapies, with external treatment as the main approach. This involves infusing the sinus tract with Chinese herbal medicine to promote the shedding of necrotic tissue or to transform thin, watery pus into thick pus, thus facilitating wound healing. Zeji fluid extract is a compound fluid extract made from the Chinese herbs Euphorbia helioscopia, Bletilla striata, and Uncaria rhynchophylla. It has the effects of promoting pus drainage, removing necrotic tissue, detoxifying, promoting blood circulation, and promoting tissue regeneration and astringency. Clinically, it is mainly used to treat chronic, difficult-to-heal wounds caused by lymph node tuberculosis, skin tuberculosis, breast tuberculosis, bone and joint tuberculosis, strong reactions to BCG vaccination leading to suppuration and rupture, or other causes (Niu Xiaohong et al., "Experimental Study on the Safety of External Application of Zeji Fluid Extract," Jiangsu Journal of Traditional Chinese Medicine 48.11(2016):4). Although Zeji fluid extract easily penetrates the sinus tract and can directly reach the affected area, it cannot achieve a stable release over a long period, resulting in a short duration of action and limited efficacy.

[0003] Hydrophilic polymer membranes, possessing high hydrophilicity, breathability, and a certain degree of mechanical strength, are ideal covering materials for treating chronic, refractory wounds. Chitosan, with its high molecular weight, exhibits excellent film-forming properties due to its intramolecular and intermolecular hydrogen bonding. Chitosan film dressings are soft, comfortable, and adhere well to the wound surface, offering both breathability and water absorption. They not only have antibacterial and anti-inflammatory effects but also provide analgesia, hemostasis, and wound healing promotion. Furthermore, they inhibit fibroblast growth, preventing secondary damage caused by adhesions during conventional dressing removal, thus reducing patient suffering and being particularly effective in resolving postoperative wound adhesions. Despite chitosan's numerous advantages in promoting wound healing, chitosan membranes suffer from low strength and poor toughness. Furthermore, their high amino and hydroxyl content leads to poor water resistance; the membrane's integrity is easily compromised after absorbing excessive wound secretions, limiting its further application. Currently, chitosan is primarily combined with other materials to improve film performance and adapt it to the needs of different environments.

[0004] Reference 1 (Journal of Biotechnology, 2020, 310: 103-113) prepared a novel polyelectrolyte membrane for wound dressings based on chitosan and hyaluronic acid biopolymers. The incorporation of glutathione into the membrane matrix significantly affected the membrane properties, improving its hydrophilicity and porosity. Reference 2 (Materials Science & Engineering C, 2019, 104: 109944) proposed an ultrasound-controlled implantable release system for doxorubicin, using an aqueous polyurethane and chitosan composite membrane as a carrier, exhibiting a high degree of flexibility in loading the membrane. Utilizing the hydrophilicity of the aqueous polyurethane and the bioactivity of the amino groups on the chitosan side chains, the prepared composite membrane exhibited good biodegradability and good cell compatibility. In vitro release studies showed that the drug-loaded membrane had a good sustained-release effect, and doxorubicin could be released under ultrasound control. Reference 3 (Carbohydrate Polymers, 2018, 197: 47-56) prepared a chitosan membrane containing excess pectin, where pectin and chitosan molecules cross-linked through physical interactions. Adjusting the pectin / chitosan ratio enhanced mechanical strength and modified hydrophilicity and cell compatibility, making it suitable for wound healing and tissue engineering. Reference 4 (Carbohydrate Polymers, 2019, 206: 428-434) used a spray-assisted layering technique with an automated device to prepare multilayer membranes of alginate and chitosan for the local and controlled release of antitumor drugs. Results showed that the antitumor drug tamoxifen was continuously released over time, and the release rate could be adjusted by the number of deposited bilayer membranes. These composite materials, by combining chitosan with other macromolecules, altered the original structure and exhibited better performance and unique functions compared to single materials. However, these materials also suffer from limitations such as limited performance improvement, complex processing procedures, and the inability to simultaneously achieve all desired properties. Summary of the Invention

[0005] The purpose of this invention is to provide a Zeji fluid extract / chitosan composite nanofilm and its preparation method. This method further prepares the Zeji fluid extract into a film dressing, giving it better mechanical properties. Combined with drug delivery, it achieves sustained drug release, allowing direct sustained drug release onto the wound surface, prolonging the drug's action time and achieving better therapeutic effects. It is suitable for repairing tuberculous wounds and other refractory wounds.

[0006] The technical solution to achieve the purpose of this invention is as follows:

[0007] The preparation method of Zeji fluid extract / chitosan composite nanofilm utilizes casting and solvent evaporation technology to promote the combination of traditional Chinese medicine and natural polysaccharide molecules, achieving membrane formation. Simultaneously, sodium alginate with aldehyde group is added during casting to improve the mechanical properties and stability of the membrane, achieving the purpose of sustained release of traditional Chinese medicine and wound treatment. The specific steps are as follows:

[0008] Step 1: According to the weight proportions, mix 800-1200 parts of Euphorbia helioscopia, 96-144 parts of Bletilla striata, 160-240 parts of Uncaria rhynchophylla, 16-240 parts of Forsythia suspensa, and 96-240 parts of Commiphora myrrha. Add 15 times the amount of water and decoct three times, 1.5 hours each time. Combine the decoctions, filter, and concentrate the filtrate to a relative density of 1.24-1.28 at 60℃ to obtain Euphorbia helioscopia fluid extract. Alternatively, add 10 times the amount of water and decoct twice, 1 hour each time. Combine the decoctions, filter, and concentrate the filtrate to a relative density of 1.24-1.28 at 60℃ to obtain Euphorbia helioscopia fluid extract.

[0009] Step 2: Dissolve carboxymethyl chitosan in the extract to obtain a traditional Chinese medicine compound solution with a carboxymethyl chitosan concentration of 0.1-0.5 wt%.

[0010] Step 3: According to the volume ratio of the extract to sodium aldehyde alkaloid solution of 5 to 20:1, add sodium aldehyde alkaloid solution of 5.0 to 10.0 wt% dropwise to the traditional Chinese medicine compound solution, stir and mix to obtain traditional Chinese medicine precipitate;

[0011] Step 4: Dry the cast film of the Chinese herbal medicine precipitate to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0012] Preferably, in step 1, the medicinal composition of the extract includes, by weight, 1000 parts of Euphorbia helioscopia, 120 parts of Bletilla striata, 200 parts of Uncaria rhynchophylla, 200 parts of Forsythia suspensa, and 120 parts of Commiphora myrrha.

[0013] Preferably, in step 2, the concentration of carboxymethyl chitosan in the traditional Chinese medicine solution is 0.2–0.3 wt%.

[0014] Preferably, in step 3, the concentration of the sodium aldehyde alginate solution is 5.0–7.0 wt%.

[0015] Preferably, in step 3, the volume ratio of the extract to the sodium alginate solution is 5-10:1.

[0016] Preferably, in step 4, the casting temperature is 40°C and the time is 24 hours.

[0017] Compared with the prior art, the significant advantages of this invention are:

[0018] (1) Toxic chemical crosslinking agents were avoided during the preparation process to ensure the biocompatibility of the material;

[0019] (2) The addition of carboxymethyl chitosan improves the extensibility of traditional Chinese medicine, enabling it to be cast into a film;

[0020] (3) Molecular precipitation significantly improves the mechanical properties and stability of the zeji fluid extract / chitosan composite nanofilm, meeting the application requirements;

[0021] (4) The Zeji fluid extract / chitosan composite nanofilm prepared by the present invention is biodegradable, which can meet the needs of drug treatment and can also be used as a tissue engineering scaffold.

[0022] (5) The process steps of the present invention are simple, safe and easy to operate, require few equipment, have mild preparation conditions and short production cycle, which meets the requirements of commercial mass production. Attached Figure Description

[0023] Figure 1 A schematic diagram of the preparation process of the Zeji fluid extract / chitosan composite nanofilm.

[0024] Figure 2 The images show the physical specimen and scanning electron microscope (SEM) image of the Zeji fluid extract / chitosan composite nanofilm prepared in Example 1.

[0025] Figure 3 The mechanical tensile properties of the zeolite / chitosan composite nanofilm prepared in Example 1 are shown.

[0026] Figure 4 The swelling performance curve of the zeolite / chitosan composite nanofilm prepared in Example 1 is shown.

[0027] Figure 5 The in vitro degradation performance curves of the Zeji fluid extract / chitosan composite nanofilm prepared in Example 1 are shown.

[0028] Figure 6 The cumulative drug release curve of the Zeji fluid extract / chitosan composite nanofilm prepared in Example 1 is shown.

[0029] Figure 7 This is a comparison diagram of the mechanical tensile strength of the composite films prepared in Example 1 and Comparative Example 1.

[0030] Figure 8 This is a comparison diagram of the mechanical tensile strength of the composite films prepared in Example 1 and Comparative Example 2. Detailed Implementation

[0031] The present invention will now be described in further detail with reference to specific embodiments and accompanying drawings.

[0032] Unless otherwise specified, all reagents used in the following examples are commercially available.

[0033] In the following examples, the Chinese herbal extract of Zeqi was prepared by Nanjing Integrated Traditional and Western Medicine Hospital. The specific preparation method is as follows: 1000 parts of Zeqi, 120 parts of Bletilla striata, 200 parts of Uncaria rhynchophylla, 200 parts of Forsythia suspensa, and 120 parts of Commiphora myrrha were mixed by weight and decocted three times with 15 times the amount of water, each time for 1.5 hours. The decoctions were combined, filtered, and the filtrate was concentrated to a relative density of 1.24-1.28 at 60°C to obtain Zeqi extract.

[0034] The preparation method of phosphate buffer (PBS) is as follows: dissolve 8g of sodium chloride, 0.2g of potassium chloride, 2.9g of disodium hydrogen phosphate, and 0.2g of potassium dihydrogen phosphate in 1000mL of distilled water and adjust the pH to 7.4.

[0035] The testing methods for thin film properties are as follows:

[0036] Microstructure observation of the thin film: After gold sputtering, the microstructure of the sample was observed using a scanning electron microscope (Quanta 250FEG, FEI).

[0037] Tensile mechanical properties of the film: The sample was cut into 50mm×10mm pieces with smooth edges, and its tensile strength and elongation were tested at a rate of 5mm / min using a universal testing machine to obtain the tensile curve.

[0038] Degradation performance of the membrane: The membrane was cut into 20mm × 20mm pieces, weighed, placed in centrifuge tubes, and 25ml of 1% PBS buffer solution with lysozyme was added. After sealing, the tubes were placed in a shaker at 37℃. The membranes were removed at fixed times, lyophilized, weighed, and the residual mass was calculated.

[0039] Drug release performance of the membrane: The drug-loaded membrane was cut into 20mm × 20mm pieces, weighed, and the total drug loading was calculated based on the mass ratio of the drug loading to the entire membrane. The samples were placed in PBS solution and incubated in a shaker at 37°C. After a certain period of time, the samples were removed, the supernatant was collected, and the absorbance was measured using a UV spectrophotometer. After measurement, the solution was poured back into the original solution. The cumulative drug release rate was calculated based on the absorbance.

[0040] Example 1

[0041] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.3 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.3 wt%;

[0042] (2) Dissolve 0.7g of sodium aldehyde alginate in 10mL of water to obtain a sodium aldehyde alginate solution with a concentration of 7.0wt%.

[0043] (3) Add 10 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate.

[0044] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a 40℃ constant temperature drying oven for 24 hours to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0045] The Zeji fluid extract / chitosan composite nanofilm prepared in this embodiment is relatively smooth and dense. The microstructure of the film was observed using scanning electron microscopy, revealing micropores distributed on the film surface (see...). Figure 2 The thickness of the thin film is 100–500 nm.

[0046] The mechanical properties of the Zeji fluid extract / chitosan composite nanofilm prepared in this embodiment are as follows: Figure 3 As shown, its maximum tensile strength reached 15.6 MPa, which is significantly better than the control blank film (nanofilm without zeolite and fluid extract, 11.2 MPa).

[0047] The Zeji extract / chitosan composite nanofilm prepared in this embodiment exhibits good stability; its swelling ratio increases slightly within 14 days, but the change is not significant (see...). Figure 4 It is stable in vitro, and the mass retention rate can reach 90.7% after 7 days (see...). Figure 5 ).

[0048] The cumulative drug release rate curve of the Zeji fluid extract / chitosan composite nanofilm prepared in this embodiment is shown in the figure below. Figure 6 As shown, the cumulative release rate after 24 hours was 24.5%, indicating a sustained-release effect.

[0049] Example 2

[0050] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.1 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.1 wt%.

[0051] (2) Dissolve 1.0g of sodium aldehyde alginate in 20mL of water to obtain a sodium aldehyde alginate solution with a concentration of 5.0wt%.

[0052] (3) Add 20 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate;

[0053] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a 50°C constant temperature drying oven for 12 hours to obtain Zeji fluid extract / chitosan composite nanofilm.

[0054] Example 3

[0055] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.5 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.5 wt%;

[0056] (2) Dissolve 0.5g of sodium aldehyde alginate in 5mL of water to obtain a sodium aldehyde alginate solution with a concentration of 10.0wt%.

[0057] (3) Add 5 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate;

[0058] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a 30°C constant temperature drying oven for 18 hours to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0059] Example 4

[0060] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.2 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.2 wt%;

[0061] (2) Dissolve 1.2g of sodium aldehyde alginate in 15mL of water to obtain a sodium aldehyde alginate solution with a concentration of 8.0wt%.

[0062] (3) Add 15 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate.

[0063] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a constant temperature drying oven at 37°C for 16 hours to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0064] Example 5

[0065] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.4 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.4 wt%.

[0066] (2) Dissolve 0.3g of sodium aldehyde alginate in 5mL of water to obtain a sodium aldehyde alginate solution with a concentration of 6.0wt%.

[0067] (3) Add 5 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate;

[0068] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a 30°C constant temperature drying oven for 20 hours to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0069] Example 6

[0070] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.25 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.25 wt%.

[0071] (2) Dissolve 0.75g of sodium aldehyde alginate in 10mL of water to obtain a sodium aldehyde alginate solution with a concentration of 7.5wt%.

[0072] (3) Add 10 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate.

[0073] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a constant temperature drying oven at 45℃ for 18 hours to obtain the Zeji fluid extract / chitosan composite nanofilm.

[0074] The properties of the Zeji fluid extract / chitosan composite nanofilms prepared in Examples 2-6 are similar to those in Example 1, all exhibiting excellent mechanical properties, stability, and drug sustained-release performance.

[0075] Comparative Example 1

[0076] (1) Prepare 10 mL of Zehe fluid extract and dissolve 0.7 g of sodium alginate in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of sodium alginate is 7.0 wt%.

[0077] (2) Dissolve 0.03g of chitosan in 10mL of lactic acid solution to obtain a chitosan solution with a concentration of 0.3wt%;

[0078] (3) Mix 10 mL of traditional Chinese medicine compound solution with 10 mL of chitosan solution and stir until viscous to obtain traditional Chinese medicine compound gel.

[0079] (4) Pour the Chinese medicine gel into a mold and dry it in a 40°C constant temperature drying oven for 24 hours to obtain a Chinese medicine film.

[0080] This comparative example uses a gelation method to prepare a traditional Chinese medicine film as a control group. Compared with the Zeji fluid extract / chitosan composite nanofilm prepared in the examples, the traditional Chinese medicine film prepared in Comparative Example 1 has lower tensile strength (see...). Figure 7 It is easily broken and degrades quickly, resulting in excessively rapid drug release that fails to meet application requirements.

[0081] Comparative Example 2

[0082] (1) Prepare 100 mL of Zehe fluid extract and dissolve 0.05 g of carboxymethyl chitosan in it to obtain a traditional Chinese medicine compound solution, wherein the concentration of carboxymethyl chitosan is 0.05 wt%.

[0083] (2) Dissolve 0.1g of sodium aldehyde alginate in 5mL of water to obtain a sodium aldehyde alginate solution with a concentration of 2.0wt%.

[0084] (3) Add 4 mL of sodium aldehyde alginate solution to 100 mL of traditional Chinese medicine compound solution and stir until all the precipitate is precipitated to obtain traditional Chinese medicine compound precipitate;

[0085] (4) Mix the Chinese medicine precipitate on a mixer, pour it into a mold, and dry it in a 40°C constant temperature drying oven for 24 hours to obtain a Chinese medicine film.

[0086] Due to the excessively low concentration of raw materials, the film forming rate in this comparative example was low, resulting in high brittleness, rapid decomposition, and significantly lower tensile strength compared to the other examples (see [examples]). Figure 8 ).

[0087] Example 7

[0088] Clinical trial of Zeji fluid extract / chitosan composite nanofilm for the treatment of tuberculous wounds

[0089] 1. Clinical Data

[0090] 1.1 General Information

[0091] A total of 276 patients with tuberculous lesions hospitalized in the Scrofula Department of Nanjing Integrated Traditional Chinese and Western Medicine Hospital, affiliated with Nanjing University of Chinese Medicine, from August 1, 2020 to February 1, 2023, were selected and randomly divided into four groups using a block randomization method: experimental group (Zeji fluid extract / chitosan composite nanofilm prepared in Example 1), control group 1 (Zeji fluid extract group), control group 2 (Zeji fluid extract + sodium zirconium phosphate alginate dressing group), and control group 3 (medical petrolatum gauze), with 69 patients in each group. This study has been reviewed and approved by the Ethics Committee of Nanjing Integrated Traditional Chinese and Western Medicine Hospital, approval number 202008020. There were no statistically significant differences in clinical data among the four groups (P>0.05), indicating comparability. See Table 1.

[0092] Table 1 Comparison of baseline data of the four groups of patients

[0093]

[0094]

[0095] Note: For comparisons of four groups of count indicators, the chi-square test is used. For comparisons of four groups of measurement indicators, analysis of variance is used, with the F-statistic, or the Kruskal-Wallis H rank-sum test is used, with the χ²-statistic. 2 In the table, M represents the median, and Q represents the interquartile range.

[0096] 1.2 Diagnostic criteria

[0097] (1) Traditional Chinese medicine diagnostic criteria and syndrome differentiation criteria: Referring to the "Standards for Diagnosis and Efficacy of Diseases and Syndromes in Traditional Chinese Medicine of the People's Republic of China", it meets the syndrome of Yin deficiency and fire excess in scrofula.

[0098] (2) Western medicine diagnostic criteria: Refer to the Clinical Practice Guidelines: Surgery Section.

[0099] ① The patient has a large abscess on the body surface, which is soft and fluctuant. After rupture, it drains pus and / or contains necrotic material; in the later stage, it liquefies and ruptures to form a wound or chronic sinus tract.

[0100] ② Acid-fast bacilli were found in the pus from aspiration of a cold abscess.

[0101] ③ Aspiration or biopsy of a cold abscess reveals typical tuberculous lesions.

[0102] ④ The pus sample tested positive for the Mycobacterium tuberculosis rpoB gene using real-time fluorescent PCR.

[0103] The main basis for this is the local symptoms and the consistency of etiology or pathological examination with tuberculosis.

[0104] 1.3 Inclusion Criteria

[0105] It meets the diagnostic and syndrome differentiation standards of traditional Chinese medicine.

[0106] Main symptoms: ruptured abscess, undermining cavity, tortuous and complex sinus tract, discharge of thin and clear pus, mixed with rotten or bean curd-like material; loose, pale or dark red granulation tissue, dark purple or dark red skin around the sore, tenderness around the sore, or pseudo-healing, repeated rupture, and slow healing.

[0107] Secondary symptoms: hot flashes and night sweats, fatigue and weakness, emaciation, red tongue with little coating, and thready and rapid pulse.

[0108] Primary symptoms are mandatory, secondary symptoms are optional. Patients aged 16 to 74 years, regardless of gender, and who meet the above-mentioned Western medical diagnostic criteria can be included in the trial cases after signing informed consent.

[0109] 1.4 Exclusion Criteria

[0110] Patients with wounds that communicate with deep organs; those allergic to this drug; those with serious primary diseases of other functional systems, creatinine (CRE) > 1 × N (N is the upper limit of normal), alanine aminotransferase (ALT) > 2 × N (N is the upper limit of normal); patients with mental illness and pregnant women are excluded.

[0111] 2. Methods

[0112] 2.1 Treatment Plan

[0113] The experimental group (local treatment with Zeji fluid extract / chitosan composite nanofilm + basic treatment) consisted of 69 patients;

[0114] Control group 1 (local treatment with Zeji fluid extract + basic treatment) 69 cases;

[0115] Two control groups (local treatment with Zeji fluid extract and sodium zirconium phosphate alginate dressing + basic treatment) included 69 patients.

[0116] The control group consisted of 69 patients (medical petroleum jelly gauze for local treatment + basic treatment).

[0117] 2.1.1 Treatment methods

[0118] 2.1.1.1 Local treatment

[0119] Experimental group: Zeji fluid extract / chitosan composite nanofilm (Example 1), specification: 10cm×10cm (piece), each time cut slightly smaller than the wound size, packed into the wound cavity, and changed every 2 days;

[0120] Control Group 1: Zeji fluid extract (provided by the preparation room of Nanjing Integrated Traditional Chinese and Western Medicine Hospital) Specification: 50mL / bottle, batch number: 20200702. Each time, 5mL of Zeji fluid extract was drawn with a syringe and slowly injected into the sinus tract. The remaining Zeji fluid extract was used to soak medical gauze to pack the sinus tract for drainage. The gauze was changed every 2 days.

[0121] Control Group 2: Zeji fluid extract (provided by the Pharmaceutical Preparation Room of Nanjing Integrated Traditional Chinese and Western Medicine Hospital) Specification: 50mL / bottle, batch number: 20200702. 5mL of Zeji fluid extract was drawn with a syringe and slowly injected into the sinus tract. Then, sodium zirconium phosphate alginate dressing (Comopec (China) Medical Supplies Co., Ltd.) Specification: 10cm×10cm (piece), batch number: W00058780 was applied. Each time, the dressing was cut slightly smaller than the size of the wound and packed into the wound cavity. It was changed every 2 days.

[0122] Control group 3: Medical Vaseline gauze (Xinxiang Huaxi Medical Supplies Co., Ltd.), batch number: 2008012461, size: 6cm×30cm, was used. Each time, it was cut slightly smaller than the size of the wound and inserted into the wound cavity for drainage. It was changed every 2 days.

[0123] 2.1.1.2 Basic Treatment

[0124] Isoniazid, rifampin, ethambutol (triple anti-tuberculosis), and bicyclol (liver protection) were all taken orally.

[0125] Isoniazid (Hangzhou Minsheng Pharmaceutical Co., Ltd.), batch number: T20J069, specification: 0.1g×100 tablets, 0.3g each time, orally, once a day.

[0126] Rifampicin (Shenyang Hongqi Pharmaceutical Co., Ltd.), batch number: G2520111, specification: 0.15g×100 tablets, 0.45g each time, taken on an empty stomach in the morning, once a day.

[0127] Ethambutol hydrochloride (Shenyang Hongqi Pharmaceutical Co., Ltd.), batch number: G0620081, specification: 0.25g×50 tablets, 0.75g each time, orally, once a day.

[0128] Bicyclol (Beijing Union Medical College Pharmaceutical Factory), batch number: 200706. Specification: 50mg x 18 tablets. Dosage: 50mg orally, twice daily.

[0129] 2.1.2 Treatment course: 28 days for all four groups.

[0130] 2.2 Observation Methods

[0131] Primary efficacy indicators: Observe wound healing, changes in systemic symptoms, and examination indicators; calculate the overall effective rate. Secondary efficacy indicators: TCM syndrome scores before and after treatment; classify major symptoms such as sinus tract length, number, amount of necrotic tissue, nature and amount of pus, hot flashes, and night sweats into five levels: none (0 points), mild (1 point), moderate (2 points), relatively severe (3 points), and severe (4 points), with weights of 2 and 1. Safety indicators: Observe changes in major indicators such as white blood cell (WBC), platelet (PLT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transferase (GGT), creatinine (CRE), and blood urea nitrogen (BUN) before and after treatment.

[0132] 2.3 Criteria for Determining Efficacy

[0133] Efficacy criteria were established based on the "Guiding Principles for Clinical Research of New Traditional Chinese Medicines".

[0134] (1) Criteria for judging the efficacy of treatment for diseases:

[0135] ① Clinical cure: Systemic symptoms disappear, wounds heal, and laboratory indicators return to normal; no recurrence was observed during a 3-month follow-up period;

[0136] ②Significant effect: The wound area is reduced by more than 1 / 2, or the number is reduced by more than 1 / 2, and the test indicators are normal;

[0137] ③ Effective: The wound area shrinks or the number of wounds decreases by less than 1 / 2, and laboratory indicators improve;

[0138] ④ Ineffective: The size and number of wounds did not improve after treatment.

[0139] Effectiveness rate = (cured + significantly effective + effective) / n × 100%.

[0140] (2) Criteria for judging syndrome efficacy: Quantitative grading standard for TCM symptoms

[0141] ① Clinical cure: TCM clinical symptoms and signs disappear or basically disappear, and the syndrome score decreases by ≥95%;

[0142] ②Significant effect: Clinical symptoms and signs in traditional Chinese medicine are significantly improved, and the syndrome score is reduced by ≥70%;

[0143] ③ Effective: Clinical symptoms and signs in TCM have improved, and the syndrome score has decreased by ≥30%;

[0144] ④ Ineffective: The clinical symptoms and signs in traditional Chinese medicine have not improved significantly, or have even worsened, and the syndrome score has decreased by less than 30%.

[0145] The efficacy index (n) = (pre-treatment score - post-treatment score) / pre-treatment score × 100%.

[0146] 2.4 Statistical Methods

[0147] SPSS 22 software was used to calculate the experimental data. Analysis of variance was used for comparisons between groups, and the chi-square test was used to compare differences in efficacy between groups. 2 The test was performed, with P < 0.05 considered statistically significant.

[0148] 3. Results

[0149] 3.1 Overall effective rate of disease treatment

[0150] The overall effective rate of the experimental group was higher than that of control group 1, control group 2, and control group 3. See Table 2.

[0151] Table 2 Comparison of total effective rates between the experimental group and the control group.

[0152]

[0153]

[0154] Note: Comparison of overall effective rates among the four groups (χ²) 2 =88.676, p=0.000), the difference was statistically significant. *When comparing the experimental group with each control group, p<0.01, the difference was statistically significant.

[0155] 3.2 Disease efficacy and cure rate

[0156] The cure rate in the experimental group was higher than that in control group 1 and control group 2. See Table 3.

[0157] Table 3 Comparison of cure rates between the experimental group and control groups 1 and 2

[0158]

[0159] Note: *Comparison between the experimental group and control group 1 and control group 2, respectively, p<0.01, the difference is statistically significant.

[0160] 3.3 Comparison of changes in syndrome efficacy scores before and after treatment

[0161] Before treatment, there was no statistically significant difference in the scores among the groups (P>0.05), indicating that the groups were balanced and comparable, and the severity of the disease was similar across groups. After treatment, the scores of each group decreased, and the differences were statistically significant. The differences in the changes in scores among the groups were statistically significant (p<0.01). Pairwise comparisons using the SNK method revealed statistically significant differences between the experimental group and control groups 1, 2, and 3 (p<0.01), statistically significant differences between control groups 1 and 2 and control groups 3 (p<0.01), and no statistically significant difference between control groups 1 and 2 (P>0.05). See Table 4.

[0162] Table 4 Comparison of syndrome scores between the experimental group and the control group

[0163]

[0164]

[0165] Note: *The variance analysis of the differences between groups showed that p<0.01, indicating that the differences between the groups were statistically significant. Pairwise comparisons using the SNK method revealed statistically significant differences between the experimental group and control group 1, control group 2, and control group 3. Statistically significant differences were also found between control group 1, control group 2, and control group 3. No statistically significant difference was found between control group 1 and control group 2.

[0166] 3.4 Safety Indicators

[0167] Safety indicators for patients in the experimental group and control groups 1, 2, and 3 were mainly determined by white blood cell count (WBC), platelet count (PLT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transferase (GGT), creatinine (CRE), and blood urea nitrogen (BUN). All indicators were within the normal range before and after treatment in all four groups. The differences between the groups were not statistically significant (P>0.05) according to ANOVA. No local skin erythema or edema, pain, bleeding, or other irritating or sensitizing reactions occurred in any of the four groups.

[0168] 3.5 Follow-up status

[0169] Three months of follow-up revealed that the wounds in the experimental group, control group 1, and control group 2 healed well, with no systemic discomfort, normal test results, and no recurrence. Two cases in control group 3 relapsed.

[0170] 4. Conclusion

[0171] The Zeji fluid extract / chitosan composite nanofilm allows the efficacy of traditional Chinese medicine to be fully realized through the sustained release and degradation effects of nanomaterials, significantly promoting the healing of tuberculous wounds. The overall effective rate and TCM syndrome scores before and after treatment were significantly better than those of Zeji fluid extract, Zeji fluid extract combined with sodium zirconium phosphate alginate dressing, and medical petrolatum gauze. The cure rate was significantly better than that of Zeji fluid extract and Zeji fluid extract combined with sodium zirconium phosphate alginate dressing. The experimental group, control group 1, and control group 2 were all better than the medical petrolatum gauze group (control group 3). There was no significant difference between Zeji fluid extract and Zeji fluid extract combined with sodium zirconium phosphate alginate dressing.

Claims

1. A method for preparing a fluid extract / chitosan composite nanofilm, characterized in that, The specific steps are as follows: Step 1: According to the weight proportions, mix 800-1200 parts of Euphorbia helioscopia, 96-144 parts of Bletilla striata, 160-240 parts of Uncaria rhynchophylla, 16-240 parts of Forsythia suspensa, and 96-240 parts of Commiphora myrrha. Add 15 times the amount of water and decoct three times, 1.5 hours each time. Combine the decoctions, filter, and concentrate the filtrate to a relative density of 1.24-1.28 at 60℃ to obtain Euphorbia helioscopia fluid extract. Alternatively, add 10 times the amount of water and decoct twice, 1 hour each time. Combine the decoctions, filter, and concentrate the filtrate to a relative density of 1.24-1.28 at 60℃ to obtain Euphorbia helioscopia fluid extract. Step 2: Dissolve carboxymethyl chitosan in the extract to obtain a traditional Chinese medicine compound solution with a carboxymethyl chitosan concentration of 0.1~0.5 wt%. Step 3: According to the volume ratio of the extract to sodium aldehyde ester solution of 5~20:1, add sodium aldehyde ester solution of 5.0~10.0 wt% dropwise to the traditional Chinese medicine compound solution, stir and mix to obtain traditional Chinese medicine precipitate; Step 4: Dry the cast film of the Chinese herbal medicine precipitate to obtain the Zeji fluid extract / chitosan composite nanofilm.

2. The preparation method according to claim 1, characterized in that, In step 1, the medicinal composition of the Zeji fluid extract, by weight, includes 1000 parts of Zeji, 120 parts of Bletilla striata, 200 parts of Uncaria rhynchophylla, 200 parts of Forsythia suspensa, and 120 parts of Myrrh.

3. The preparation method according to claim 1, characterized in that, In step 2, the concentration of carboxymethyl chitosan in the traditional Chinese medicine solution is 0.2~0.3 wt%.

4. The preparation method according to claim 1, characterized in that, In step 3, the concentration of the sodium aldehyde alginate solution is 5.0~7.0 wt%.

5. The preparation method according to claim 1, characterized in that, In step 3, the volume ratio of the extract to the sodium alginate solution is 5-10:

1.

6. The preparation method according to claim 1, characterized in that, In step 4, the casting temperature is 40℃ and the time is 24 hours.

7. The zeolite / chitosan composite nanofilm prepared by the preparation method according to any one of claims 1 to 6.

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