A traditional Chinese medicine paste for treating clostridium difficile and bacterial flora reconstruction and a preparation method thereof

Abstract

This invention discloses a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance, and its preparation method. It relates to the field of traditional Chinese medicine plaster technology, comprising a substrate layer, a plaster, and a protective layer. The plaster is made from active ingredients, a penetrant, excipients, an adsorbent, and deionized water. The active ingredients are made from Atractylodes macrocephala volatile oil, Magnolia officinalis volatile oil, Artemisia argyi volatile oil, Coptis chinensis extract, Taraxacum mongolicum extract, Paeonia lactiflora extract, and Poria cocos extract. This invention achieves precise intervention in the intestinal microenvironment through targeted drug delivery via the umbilicus, combining transdermal absorption and acupoint stimulation mechanisms. The porous sustained-release structure of the adsorbent prolongs the drug's action time, simultaneously regulating flora balance and inhibiting pathogenic bacteria proliferation. This overcomes the limitations of traditional oral or injection therapies, providing an efficient and convenient external treatment solution for drug-resistant Clostridium difficile infection.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine plaster technology, specifically to a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance, and its preparation method. Background Technology

[0002] Clostridium difficile (CDI) infection is a serious intestinal disease primarily caused by toxins produced by Clostridium difficile. It can lead to a range of clinical symptoms, from mild diarrhea to pseudomembranous colitis, toxic megacolon, and even death. In recent years, with the widespread use of antibiotics, the incidence and recurrence rate of CDI have been on the rise, making it a significant hospital-acquired infection and public health problem worldwide.

[0003] Clostridium difficile infections primarily occur in hospital settings, with the vast majority being exogenous infections; the hospital environment is the main source of Clostridium difficile. This bacterium can form spores and survive for extended periods under adverse conditions, increasing the difficulty of infection control. High-risk groups include the elderly, those with weakened immune systems, those on long-term antibiotic use, and patients with kidney failure, inflammatory bowel disease, or chronic liver disease.

[0004] Currently, the main treatments for Clostridium difficile infection include:

[0005] Antibiotic treatment: Vancomycin, metronidazole, and other antibiotics are first-line drugs for treating CDI. However, antibiotic treatment has significant limitations, including a high failure rate, a high relapse rate (approximately 20%–30% of patients will experience a relapse), and the potential to further disrupt the balance of gut microbiota.

[0006] Fecal microbiota transplantation (FMT): For patients with recurrent CDI, FMT is an effective treatment, but it has problems such as complex operation, low acceptance, and potential risks.

[0007] Probiotics: used as an adjunct therapy, but their efficacy remains controversial.

[0008] Comprehensive Western medicine treatment includes discontinuing antibiotics that may cause CDI, supplementing electrolytes, and using probiotics.

[0009] The main problems with current treatment methods are high relapse and treatment failure rates, and long-term use of antibiotics may lead to increased drug resistance, further complicating treatment. In addition, current treatments are mostly systemic, which may have unnecessary effects on tissues and organs outside the intestines, and it is difficult to maintain effective drug concentrations in the intestinal tract.

[0010] Traditional Chinese medicine has a long history and rich experience in the treatment of digestive system diseases. Traditional Chinese medicine theory believes that Clostridium difficile infection falls under the categories of "diarrhea" and "dysentery" and is related to factors such as internal damp-heat and spleen and stomach weakness.

[0011] However, currently, traditional Chinese medicine (TCM) is mainly administered orally in the treatment of Clostridium difficile infection (CDI), which suffers from low bioavailability and insufficient local drug concentration in the intestines. Furthermore, there is a lack of topical TCM preparations specifically targeting Clostridium difficile infection, particularly those that can directly act on the colon.

[0012] Traditional Chinese medicine plasters, as a traditional method of drug delivery, have advantages such as local application, sustained release, and ease of use. However, existing traditional Chinese medicine plasters are mostly used for external injuries and rheumatic pain, with limited research on plaster formulations for internal organ diseases, especially intestinal infections.

[0013] The development of a traditional Chinese medicine plaster for Clostridium difficile infection, which can deliver the drug directly to the intestinal lesion through transdermal absorption, avoids the first-pass effect of oral administration, improves the bioavailability of the drug, and reduces systemic adverse reactions, has important clinical application value. Summary of the Invention

[0014] To address the shortcomings of existing technologies, the present invention aims to provide a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance, as well as its preparation method. The present invention delivers medication through the umbilicus (Shenque acupoint), combining transdermal absorption and acupoint stimulation mechanisms to achieve precise intervention in the intestinal microenvironment. The porous sustained-release structure of the adsorbent prolongs the drug's action time, simultaneously regulating the flora balance and inhibiting the proliferation of pathogenic bacteria. This overcomes the limitations of traditional oral or injection therapies and provides an efficient and convenient external treatment solution for drug-resistant Clostridium difficile infection.

[0015] To achieve the above objectives, the present invention adopts the following technical solution:

[0016] A traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance includes a base layer, a medicated plaster, and a protective layer. The medicated plaster is made of 1-6 wt% active ingredients, 0.5-2 wt% penetrant, 0.5-2 wt% excipient, 40-52 wt% adsorbent, and 38-58 wt% deionized water. The active ingredients, by weight, are made of the following components: 4-5 parts of Atractylodes macrocephala volatile oil, 3-6 parts of Magnolia officinalis volatile oil, 1-3 parts of Artemisia argyi volatile oil, 16-24 parts of Coptis chinensis extract, 8-12 parts of Taraxacum mongolicum extract, 12-18 parts of Paeonia lactiflora extract, and 12-20 parts of Poria cocos extract.

[0017] Preferably, the preparation method of volatile oil of traditional Chinese medicine is as follows: take Atractylodes macrocephala, Magnolia officinalis, and Artemisia argyi, crush them, soak them in deionized water, reflux to extract, collect the volatile oil, and obtain the volatile oil of Atractylodes macrocephala, volatile oil of Magnolia officinalis, and volatile oil of Artemisia argyi, and obtain the volatile residue for later use.

[0018] After appropriate pulverization of medicinal materials such as Atractylodes macrocephala, the cell walls are mechanically broken, creating micro-cracks. When soaked in water, the medicinal materials absorb water and swell, further softening the cell structure. During reflux, water molecules penetrate into the cell interior and form an azeotropic system with the volatile oils. Under continuous heating, the volatile oil components evaporate along with the steam due to the decrease in vapor pressure. After condensation, the oil and water immiscibility is utilized to achieve layer separation. In this process, the dual cell wall disruption (mechanical pulverization + hydration swelling) promotes the release of volatile oils, while the physical mass transfer characteristics of steam distillation selectively extract heat-sensitive volatile components, completely preserving the chemical structure of active substances such as terpenes and phenylpropanoids.

[0019] Preferably, the Chinese herbal medicine is pulverized into 40-80 mesh, the weight of deionized water is 10-20 times that of the Chinese herbal medicine, soaked for 30-60 minutes, and the reflux extraction temperature is 100℃.

[0020] Preferably, the preparation method of the Chinese herbal extract is as follows: Coptis chinensis, dandelion, white peony root, and Poria cocos are added to ethanol, heated under reflux, the extracts are combined, filtered, the filtrate is reduced under reduced pressure to recover ethanol, and concentrated into a thick paste to obtain the Coptis chinensis extract, dandelion extract, white peony root extract, and Poria cocos extract, and the extract residue is obtained for later use.

[0021] When Coptis chinensis and other medicinal materials are extracted with ethanol solution, the moderately polar ethanol molecules selectively dissolve active ingredients such as flavonoids and alkaloids based on the principle of "like dissolves like". During the heating process, ethanol permeates the cell membrane and generates an osmotic pressure difference, which promotes the diffusion of intracellular active ingredients to the solvent phase through the concentration gradient. At the same time, thermal motion enhances the molecular diffusion rate. The segmented reflux extraction breaks the solid-liquid two-phase distribution equilibrium step by step through the principle of dynamic equilibrium, which improves the solute migration efficiency. In the vacuum concentration stage, the low boiling point of ethanol is used to preferentially vaporize and separate the components, avoiding the destruction of heat-sensitive components by high temperature. Finally, a thick paste system rich in polar to moderately polar active molecules is formed, which fully preserves antibacterial, anti-inflammatory and other functional components.

[0022] Preferably, the ethanol concentration is 60-80 wt%, and the weight of the ethanol is 8-15 times that of the traditional Chinese medicine; the extraction is carried out by heating and reflux twice, each time for 1-2 hours.

[0023] Atractylodes macrocephala: It is bitter, sweet, and warm in nature, and enters the spleen and stomach meridians. It has the effects of tonifying qi and strengthening the spleen, drying dampness and promoting diuresis, and consolidating the exterior and stopping sweating. It is mainly used to treat abdominal distension and diarrhea due to spleen deficiency and dampness. Its volatile oil contains sesquiterpenoid components such as atractylone and atractylodes lactone, which can destroy the sterol structure of Clostridium difficile cell membrane and inhibit bacterial proliferation.

[0024] Magnolia officinalis: bitter, pungent, and warm in nature, it enters the spleen, stomach, lung, and large intestine meridians. Its functions include drying dampness and resolving phlegm, lowering qi and relieving fullness, and it is effective for epigastric fullness, vomiting, and diarrhea caused by dampness obstructing the middle jiao. The volatile oil contains magnolol and other biphenyl lignans, which block toxin release by inhibiting bacterial β-glucosidase activity.

[0025] Artemisia argyi: Bitter, pungent, and warm in nature, it enters the liver, spleen, and kidney meridians. It can warm the meridians to stop bleeding, dispel cold, and relieve pain. It is mainly used to treat bleeding due to deficiency and cold, as well as skin eczema. Its volatile oil contains monoterpenoid compounds such as eucalyptol and camphor, which can interfere with bacterial biofilm formation and promote transdermal drug absorption.

[0026] Coptis chinensis: Bitter and cold in nature, it enters the heart, spleen, stomach, liver, and gallbladder meridians. It clears heat and dries dampness, drains fire and detoxifies, and is specifically used to treat damp-heat fullness and vomiting. Berberine and other isoquinoline alkaloids in its extract can inhibit bacterial nucleic acid metabolism and downregulate the expression of inflammatory factors.

[0027] Dandelion: Bitter, sweet, and cold in nature, it enters the liver and stomach meridians. It clears heat and detoxifies, reduces swelling and dissipates nodules, and is effective in treating carbuncles and boils caused by heat toxins. Extracts contain flavonoids such as luteolin, which can reduce inflammatory infiltration of the intestinal mucosa.

[0028] White peony root: bitter, sour, and slightly cold in nature; it enters the liver and spleen meridians; it nourishes blood, astringes yin, softens the liver, and relieves pain; it is used for abdominal pain due to blood deficiency. Extracts such as paeoniflorin and other monoterpenoid glycosides can regulate immune balance to repair the intestinal mucosal barrier.

[0029] Poria cocos: sweet, bland, and neutral in nature; it enters the heart, spleen, and kidney meridians; it promotes diuresis and eliminates dampness, strengthens the spleen and calms the mind; it is mainly used to treat edema and oliguria. Extracts containing pachymic acid and other triterpenoids can enhance the phagocytic function of macrophages.

[0030] Preferably, the preparation method of the adsorbent includes the following steps: taking volatile residue and extract residue, grinding them to obtain residue powder; dispersing the residue powder into a starch solution, stirring and mixing evenly to obtain a residue starch solution; adding calcium carbonate powder to the residue starch solution, stirring and mixing evenly to obtain calcium carbonate coated with residue starch, calcining, and then washing with hydrochloric acid to obtain the adsorbent;

[0031] Preferably, the calcium carbonate powder has a particle size of 0.5 to 2 mm.

[0032] First, the residue powder is mixed with a phosphoric acid-modified starch solution. The phosphoric acid promotes the cross-linking of starch molecules to form a three-dimensional network framework. Then, calcium carbonate powder is introduced as a template agent. During the stepped heating pyrolysis process, the organic matter is carbonized at low temperature to form a porous carbon matrix. At medium temperature, calcium carbonate reacts with the carbon layer to generate active calcium oxide and releases gas to create pores. At high temperature, calcium carbonate decomposes completely to produce carbon dioxide, which reacts with the carbon matrix to form a gasification pore-expanding reaction. Finally, hydrochloric acid selectively etches the calcium oxide to generate soluble salts, exposing hierarchical pores. At the same time, the oxygen-containing functional groups activated by phosphoric acid endow the surface with chemical adsorption activity, forming a composite adsorption material with both physical adsorption channels and chemical binding sites.

[0033] Preferably, the dregs are ground to 500-1000 mesh; the starch solution is obtained by dissolving starch in a 10wt% phosphoric acid aqueous solution to obtain a starch solution with a concentration of 30-40wt%; the ratio of dregs powder to starch solution is 10g:15-30mL; the ratio of calcium carbonate powder to dregs starch solution is 10g:10-15mL; the calcination conditions are pyrolysis at 400-600℃ for 30-60min, 600-800℃ for 30-60min, and 800-900℃ for 30-60min.

[0034] Preferably, the penetrant is one or more of menthol, glycerin, hydroquinone, lauryl ketone, acetone, propylene glycol, and ethanol.

[0035] Preferably, the excipient is one or more of petrolatum, dimethicone, lanolin, and beeswax.

[0036] This invention also claims a method for preparing a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance, comprising the following steps: mixing the active ingredient with an adsorbent, then dispersing it in deionized water, adding a penetrant and excipients, stirring and mixing evenly to obtain a plaster, applying the plaster to a substrate layer, and covering it with a protective layer to obtain the traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance.

[0037] This invention also claims the application of a traditional Chinese medicine plaster for the adjuvant treatment of Clostridium difficile infection and the restoration of intestinal flora balance, characterized by the following steps: cleaning the navel and surrounding skin, keeping it dry, aligning the core of the plaster with the center of the navel, applying it for 4-8 hours daily, 1-2 times daily, for 5-10 days.

[0038] Compared with the prior art, the present invention has the following beneficial effects:

[0039] 1. This invention provides a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance. It leverages the advantages of umbilical administration, where the navel (Shenque acupoint) serves as a meridian intersection point. After drug penetration through the skin, it directly stimulates acupoints to activate meridian qi, regulating gastrointestinal qi and blood circulation via the Ren meridian. Simultaneously, the thin epidermis and rich blood vessels of the navel significantly improve drug bioavailability. The adsorbent-loaded active ingredients achieve sustained release through a hollow porous structure, avoiding the rapid dissipation of volatile oils in traditional plasters. The penetrant reduces skin barrier resistance by altering the lipid arrangement of the stratum corneum, promoting the penetration of small molecule drugs. The excipients provide suitable adhesion and spreadability, ensuring close contact between the plaster and the skin and maintaining continuous drug release.

[0040] 2. This invention provides a medicated ointment in which the active ingredients include Atractylodes macrocephala volatile oil to help increase beneficial intestinal bacteria, and Poria cocos polysaccharides to enhance the protective function of the gut microbiota, jointly inhibiting the growth of harmful bacteria; the active ingredients of Magnolia officinalis and Artemisia argyi disrupt the structure of Clostridium difficile, reducing toxin production, while Coptis chinensis inhibits bacterial division and reproduction; Paeonia lactiflora and Atractylodes macrocephala extracts promote tight junctions of intestinal cells and repair the mucosal barrier; and Taraxacum mongolicum and Magnolia officinalis components reduce intestinal damage by inhibiting inflammatory response pathways. These components form a synergistic mechanism of "antibacterial-detoxification-repair-anti-inflammatory," gradually taking effect from clearing pathogens to restoring intestinal health.

[0041] 3. This invention provides an adsorbent where the microporous-mesoporous system formed by the CO2 produced by the pyrolysis of calcium carbonate significantly increases drug loading, simultaneously encapsulating volatile oils and extracts. The active sites remaining in the medicinal residue bind drug components through non-covalent bonds, and the pore confinement effect slows the volatile oil escape rate, ensuring stable release of the active ingredients throughout the entire usage cycle. Furthermore, the resource utilization of the medicinal residue combines environmental friendliness with optimized drug loading performance. It solves the problem of rapid volatile oil evaporation in traditional plasters and prolongs drug retention time through the natural closed environment of the umbilical depression, enhancing the synergistic effect of intestinal mucosal immune repair and flora regulation, thus improving the therapeutic effect of Clostridium difficile infection. Attached Figure Description

[0042] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly described below. It should be understood that the following drawings only show some schematic diagrams of certain embodiments of the present invention, and therefore should not be regarded as a limitation of the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.

[0043] Figure 1 The survival rate of mice treated with the herbal plaster of this invention for Clostridium difficile infection and for restoring intestinal flora balance was measured. Detailed Implementation

[0044] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. Of course, the specific embodiments described herein are only for explaining the invention and are not intended to limit the invention.

[0045] Unless otherwise specified, all chemical reagents and materials in this invention are purchased from the market or synthesized from raw materials purchased from the market.

[0046] A method for preparing a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance includes the following steps:

[0047] (1) Take Atractylodes macrocephala, Magnolia officinalis and Artemisia argyi, crush them into 40-80 mesh, add them to 10-20 times the weight of the Chinese medicine in deionized water and soak for 30-60 minutes, reflux at 100℃ to extract, collect the volatile oil, and obtain the volatile oil of Atractylodes macrocephala, Magnolia officinalis and Artemisia argyi respectively, and obtain the volatile residue for later use.

[0048] (2) Take Coptis chinensis, dandelion, white peony root, and Poria cocos, add them to 60-80 wt% ethanol at 8-15 times their weight, heat and reflux to extract twice, each time for 1-2 hours, combine the extracts, filter, recover the ethanol from the filtrate under reduced pressure, concentrate into a thick paste, and obtain the Coptis chinensis extract, dandelion extract, white peony root extract, and Poria cocos extract, and obtain the extract residue for later use;

[0049] (3) Take the volatile residue and extract residue, grind them to 500-1000 mesh to obtain residue powder; disperse 10g of residue powder into 15-30mL of 30-40wt% starch solution (obtained by dissolving starch in 10wt% phosphoric acid aqueous solution), stir and mix evenly to obtain residue starch solution; add 10g of calcium carbonate powder to 10-15mL of residue starch solution, stir and mix evenly to obtain calcium carbonate coated with residue starch, pyrolyze at 400-600℃ for 30-60min, 600-800℃ for 30-60min, and 800-900℃ for 30-60min, and then wash with hydrochloric acid to obtain the adsorbent;

[0050] (4) Take 4-5 parts of Atractylodes macrocephala volatile oil, 3-6 parts of Magnolia officinalis volatile oil, 1-3 parts of Artemisia argyi volatile oil, 16-24 parts of Coptis chinensis extract, 8-12 parts of Taraxacum mongolicum extract, 12-18 parts of Paeonia lactiflora extract, and 12-20 parts of Poria cocos extract to obtain the effective components; mix 1-6 wt% of the effective components with 40-52 wt% of adsorbent, then disperse them in 38-58 wt% deionized water, add 0.5-2 wt% of penetrant and 0.5-2 wt% of excipient, stir and mix evenly to obtain the ointment, apply the ointment to the substrate layer, cover with a protective layer, and obtain the Chinese herbal plaster for adjuvant treatment of Clostridium difficile infection and reconstruction of intestinal flora balance.

[0051] The penetrant is one or more of menthol, glycerin, hydroquinone, lauryl ketone, acetone, propylene glycol, and ethanol; the excipient is one or more of petrolatum, dimethicone, lanolin, and beeswax.

[0052] The present invention will be further described below through specific embodiments.

[0053] Example 1

[0054] A method for preparing a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance includes the following steps:

[0055] (1) Take Atractylodes macrocephala, Magnolia officinalis and Artemisia argyi, grind them into 50 mesh, add them to 15 times the weight of the Chinese medicine in deionized water and soak for 45 minutes, reflux at 100℃ to extract, collect the volatile oil, and obtain the volatile oil of Atractylodes macrocephala, volatile oil of Magnolia officinalis and volatile oil of Artemisia argyi, and obtain the volatile residue for later use.

[0056] (2) Take Coptis chinensis, dandelion, white peony root and Poria cocos, add them to 70wt% ethanol at 12 times their weight, heat and reflux to extract twice, 1.5h each time, combine the extracts, filter, recover the ethanol from the filtrate under reduced pressure, concentrate into a thick paste, and obtain the Coptis chinensis extract, dandelion extract, white peony root extract and Poria cocos extract respectively, and obtain the extract residue for later use.

[0057] (3) Take the volatile residue and extract residue, grind them to 800 mesh to obtain residue powder; disperse 10g of residue powder into 25mL of 35wt% starch solution (obtained by dissolving starch in 10wt% phosphoric acid aqueous solution), stir and mix evenly to obtain residue starch solution; add 10g of calcium carbonate powder to 12mL of residue starch solution, stir and mix evenly to obtain calcium carbonate coated with residue starch, pyrolyze at 500℃ for 45min, 700℃ for 45min, and 850℃ for 45min, and then wash with hydrochloric acid to obtain the adsorbent;

[0058] (4) Take 4.5g of Atractylodes macrocephala volatile oil, 4.5g of Magnolia officinalis volatile oil, 2g of Artemisia argyi volatile oil, 20g of Coptis chinensis extract, 10g of Taraxacum mongolicum extract, 15g of Paeonia lactiflora extract, and 16g of Poria cocos extract to obtain the effective components; mix 3.5g of the effective components with 46g of adsorbent, then disperse them in 48g of deionized water, add 1g of penetrant laurocapram and 1.5g of excipient beeswax, stir and mix evenly to obtain the ointment, apply the ointment to the non-woven fabric substrate layer with a diameter of 2-5cm, cover with a non-woven fabric protective layer, and obtain the Chinese herbal plaster for adjuvant treatment of Clostridium difficile infection and reconstruction of intestinal flora balance.

[0059] Comparative Example 1

[0060] A method for preparing a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection and restoration of intestinal flora balance includes the following steps:

[0061] (1) Take Atractylodes macrocephala, Magnolia officinalis and Artemisia argyi, grind them into 50 mesh, add them to 15 times the weight of the Chinese medicine in deionized water and soak for 45 minutes, reflux at 100℃ to extract, collect the volatile oil, and obtain the volatile oil of Atractylodes macrocephala, volatile oil of Magnolia officinalis and volatile oil of Artemisia argyi, and obtain the volatile residue for later use.

[0062] (2) Take Coptis chinensis, dandelion, white peony root and Poria cocos, add them to 70wt% ethanol at 12 times their weight, heat and reflux to extract twice, 1.5h each time, combine the extracts, filter, recover the ethanol from the filtrate under reduced pressure, concentrate into a thick paste, and obtain the Coptis chinensis extract, dandelion extract, white peony root extract and Poria cocos extract respectively, and obtain the extract residue for later use.

[0063] (3) Take the volatile residue and extract residue, grind them to 800 mesh to obtain residue powder; 10g of residue powder is pyrolyzed at 500℃ for 45min, 700℃ for 45min, and 850℃ for 45min, and then washed with hydrochloric acid to obtain the adsorbent.

[0064] (4) Take 4.5g of Atractylodes macrocephala volatile oil, 4.5g of Magnolia officinalis volatile oil, 2g of Artemisia argyi volatile oil, 20g of Coptis chinensis extract, 10g of Taraxacum mongolicum extract, 15g of Paeonia lactiflora extract, and 16g of Poria cocos extract to obtain the effective components; mix 3.5g of the effective components with 46g of adsorbent, then disperse them in 48g of deionized water, add 1g of penetrant laurocapram and 1.5g of excipient beeswax, stir and mix evenly to obtain the ointment, apply the ointment to the non-woven fabric substrate layer with a diameter of 2-5cm, cover with a non-woven fabric protective layer, and obtain the Chinese herbal plaster for adjuvant treatment of Clostridium difficile infection and reconstruction of intestinal flora balance.

[0065] A therapeutic experiment on mice with diarrhea using traditional Chinese medicine plasters.

[0066] I. Experimental Materials

[0067] 1. Laboratory animals

[0068] SPF-grade KM mice (6 weeks old, weighing 20±2g) were divided into 4 groups of 10 each, half male and half female (normal group, infection group, Example 1 group, and Comparative Example 1 group).

[0069] 2. Strains and reagents

[0070] Escherichia coli ATCC25922 (bacterial concentration 1×10⁹ CFU / mL);

[0071] Antibiotic mixture (gentamicin sulfate 65g / L + cephalexin 65g / L);

[0072] Example 1: Traditional Chinese Medicine Plaster;

[0073] Comparative Example 1: Traditional Chinese Medicine Plaster.

[0074] II. Experimental Methods

[0075] 1. Establishment of animal models

[0076] Antibiotic pretreatment: Mice in the infection group, Example 1 group, and Comparative Example 1 group were administered 25 mL / (kg·d) of antibiotic mixture by gavage daily for 5 consecutive days to disrupt the intestinal flora.

[0077] Pathogen infection: After pretreatment, 0.2 mL of Escherichia coli ATCC25922 bacterial suspension (1×10⁹ CFU / mL) was administered by gavage, and the patient was observed for 24 hours until symptoms of drowsiness, loose stools, and perianal contamination appeared.

[0078] 2. Grouping and Dosing

[0079] Normal group: No infection, no intervention.

[0080] Infected group: No treatment after infection.

[0081] Example 1 group: 24 hours after infection, the patch of Example 1 was applied to the navel of the mice (changed once a day for 7 days).

[0082] Comparative Example 1: Same as Example 1, but with Comparative Example 1 plaster applied.

[0083] 3. Observation indicators

[0084] Fecal score: The score was determined by the size of the stain formed on the filter paper due to fecal contamination, and was divided into four levels: Level 1 was a stain diameter < 1.0 cm, indicated by "+"; Level 2 was a stain diameter 1.0–1.9 cm, indicated by "++"; Level 3 was a stain diameter 2.0–3.0 cm, indicated by "+++"; and Level 4 was a stain diameter > 3.0 cm, indicated by "++++". Specific results are shown in Table 1.

[0085] Table 1. Fecal Score Results

[0086] Post-modeling fecal scoring + ++++ ++++ ++++ Post-treatment stool score + ++++ + ++

[0087] Diarrhea rate: In accordance with "Experimental Methodology in Pharmacology" (4th edition), the diarrhea rate was measured on days 0, 3, and 7 of the experiment. Diarrhea rate = number of animals with diarrhea / total number in the group × 100%. The specific results are shown in Table 2.

[0088] Table 2. Results of diarrhea rate detection

[0089]

[0090] Inflammatory factors: 24 hours after the last administration, serum and small intestinal tissue were collected from mice in each group, and the levels of TNF-α and IL-6 were detected by ELISA. Specific data are shown in Table 3.

[0091] Table 3 Results of inflammatory factor detection

[0092]

[0093] Note: * indicates p<0.05 compared to the normal group; # indicates p<0.05 compared to the infected group.

[0094] Experimental study on the therapeutic effect of traditional Chinese medicine plaster on mice infected with Clostridium difficile.

[0095] I. Experimental Materials

[0096] 1. Laboratory animals

[0097] SPF-grade BALB / c mice (6 weeks old, male, weighing 20±2g), 10 mice per group, for a total of 5 groups (normal group, infection group, Example 1 group, Comparative Example 1 group, vancomycin group).

[0098] 2. Strains and reagents

[0099] Clostridium difficile ATCC43255 (anaerobic culture, bacterial concentration 1×10⁹ CFU / mL);

[0100] Antibiotic mixture (kanamycin 0.4 mg / mL, gentamicin 0.035 mg / mL, colistin 850 U / mL, metronidazole 0.215 mg / mL, vancomycin 0.045 mg / mL);

[0101] Clindamycin injection (10 mg / kg);

[0102] Vancomycin suspension (50 mg / kg, orally);

[0103] Example 1: Traditional Chinese Medicine Plaster;

[0104] Comparative Example 1: Traditional Chinese Medicine Plaster.

[0105] II. Experimental Methods

[0106] 1. Establishment of animal models

[0107] Antibiotic pretreatment: Mice were allowed free access to water containing an antibiotic mixture for 5 days, followed by 3 days of sterilized water.

[0108] Infection induction: Clindamycin (10 mg / kg) was injected intraperitoneally, followed by gavage administration of 0.2 mL of Clostridium difficile ATCC43255 bacterial suspension 24 hours later.

[0109] 2. Grouping and Dosing

[0110] Normal group: No infection, no intervention.

[0111] Infected group: No treatment after infection.

[0112] Example 1 group: 16 hours after infection, apply the Example 1 patch to the navel (change once a day for 5 days).

[0113] Comparative Example 1: Same as Example 1, but with Comparative Example 1 plaster applied.

[0114] Vancomycin group: Vancomycin suspension (50 mg / kg, once daily for 5 consecutive days) was administered by gavage 16 hours after infection.

[0115] 3. Observation indicators

[0116] Survival rate: Records deaths within 7 days. See details below. Figure 1 .

[0117] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection, comprising a substrate layer, an ointment, and a protective layer, characterized in that, The ointment is made from 1-6 wt% active ingredients, 0.5-2 wt% penetrant, 0.5-2 wt% excipient, 40-52 wt% adsorbent, and 38-58 wt% deionized water; the active ingredients, by weight, are made from the following components: 4-5 parts of Atractylodes macrocephala volatile oil, 3-6 parts of Magnolia officinalis volatile oil, 1-3 parts of Artemisia argyi volatile oil, 16-24 parts of Coptis chinensis extract, 8-12 parts of Taraxacum mongolicum extract, 12-18 parts of Paeonia lactiflora extract, and 12-20 parts of Poria cocos extract; The preparation method of volatile oil of Chinese medicine is as follows: take Atractylodes macrocephala, Magnolia officinalis and Artemisia argyi, crush them, soak them in deionized water, reflux to extract, collect the volatile oil, and obtain the volatile oil of Atractylodes macrocephala, volatile oil of Magnolia officinalis and volatile oil of Artemisia argyi, and obtain the volatile residue for later use. The preparation method of the Chinese herbal extract is as follows: take Coptis chinensis, dandelion, white peony root, and Poria cocos, add them to ethanol, heat and reflux, combine the extracts, filter, recover the ethanol from the filtrate under reduced pressure, concentrate into a thick paste, and obtain the Coptis chinensis extract, dandelion extract, white peony root extract, and Poria cocos extract, and obtain the extract residue for later use. The preparation method of the adsorbent includes the following steps: taking volatile residue and extract residue, grinding them to obtain residue powder; dispersing the residue powder into a starch solution, stirring and mixing evenly to obtain residue starch solution; adding calcium carbonate powder into the residue starch solution, stirring and mixing evenly to obtain calcium carbonate coated with residue starch, calcining, and then washing with hydrochloric acid to obtain the adsorbent.

2. The traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection according to claim 1, characterized in that, The Chinese herbal medicine is pulverized into 40-80 mesh, and the weight of deionized water is 10-20 times that of the Chinese herbal medicine. The herbs are soaked for 30-60 minutes, and the reflux extraction temperature is 100℃.

3. The traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection according to claim 1, characterized in that, The ethanol concentration is 60-80 wt%, and the weight of ethanol is 8-15 times that of the Chinese medicine; the extraction is carried out by heating and reflux twice, each time for 1-2 hours.

4. The traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection according to claim 1, characterized in that, The dregs are ground to 500-1000 mesh; the starch solution is obtained by dissolving starch in a 10wt% phosphoric acid aqueous solution to obtain a starch solution with a concentration of 30-40wt%; the ratio of dregs powder to starch solution is 10g:15-30mL; the ratio of calcium carbonate powder to dregs starch solution is 10g:10-15mL; the calcination conditions are pyrolysis at 400-600℃ for 30-60min, 600-800℃ for 30-60min, and 800-900℃ for 30-60min.

5. The traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection according to claim 1, characterized in that, The penetrant is one or more of menthol, glycerin, hydroquinone, azone, propylene glycol, and ethanol.

6. The traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection according to claim 1, characterized in that, The excipient is one or more of petrolatum, dimethicone, lanolin, and beeswax.

7. A method for preparing a traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection as described in any one of claims 1 to 6, characterized in that, The process includes the following steps: mixing the active ingredient with the adsorbent, then dispersing it in deionized water, adding the penetrant and excipients, stirring and mixing evenly to obtain an ointment, applying the ointment to the substrate layer, and covering it with a protective layer to obtain the traditional Chinese medicine plaster for adjuvant treatment of Clostridium difficile infection.

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