Polyethylene glycol-polycaprolactone carried nano-micelle type chemical fiber aerosol inhalant as well as preparation method and application of polyethylene glycol-polycaprolactone carried nano-micelle type chemical fiber aerosol inhalant

Abstract

The invention discloses a polyethylene glycol-polycaprolactone carried nano-micelle type chemical fiber aerosol inhalant as well as a preparation method and application thereof, and belongs to the technical field of biological medicines. The chemical fiber formula comprises the following traditional Chinese medicine raw materials by weight: 15g of honeysuckle, 10g of scutellaria baicalensis, 15g of caulis bambusae in taeniam, 15g of tussilago farfara, 20g of astragalus membranaceus, 20g of salvia miltiorrhiza, 5g of safflower carthamus, 5g of pseudo-ginseng powder and 9g of liquorice. The self-assembly characteristic of high-molecular-weight PEG5000-PCL10000 is utilized to provide a large enough hydrophobic inner core and a stable hydrophilic shell, the nano-micelle type chemical fiber square atomizing agent is prepared, various components in a chemical fiber square can be carried to a large extent and stably exist in water, and the chemical fiber square atomizing agent can be used for preparing the chemical fiber square atomizing agent through modification of an atomizing inhalant dosage form. The economic and time cost of a patient can be reduced, and the comprehensive improvement efficiency of the chemical fiber formula on the radiation-induced lung injury is improved. The invention has great significance in the field of aerosol inhalation treatment of lung diseases.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of biological medicine, and particularly relates to a polyethylene glycol-poly-caprolactone-carrying nanomicellar type chemical fiber aerosol inhalation agent and a preparation method and application thereof. BACKGROUND

[0002] Radiation-induced lung injury (RILI) is one of the serious complications caused by chest radiotherapy. When the lung tissue receives a radiation dose exceeding its biological effect threshold, it may trigger this disease. RILI can be divided into early radiation pneumonia (RP) and late radiation-induced pulmonary fibrosis (RIPF). RP usually occurs 4-12 weeks after the start of radiotherapy, with an average incidence of about 35%. Its clinical manifestations mainly include symptoms such as fever, cough, chest pain and dyspnea. The current clinical treatment for RP mainly includes the use of antibiotics, anti-inflammatory agents, bronchodilators and glucocorticoids. These measures can alleviate symptoms to a certain extent, but the overall efficacy is limited, and long-term use may cause serious side effects. The development of RILI to the later stage will cause irreversible fibrosis and scarring damage to the lung tissue, and the pathogenesis of RILI is not fully understood, and there is currently a lack of effective intervention measures that can be used to treat RIPF. Therefore, the key to the management strategy of RILI is to develop safer and more effective prevention and treatment measures, strictly control the probability and severity of RP, and avoid its transformation into RIPF in the later stage and fall into a difficult situation.

[0003] A large number of basic and clinical studies show that some Chinese patent medicines and decoctions have significant effects on the prevention and treatment of radiation-induced lung injury under the guidance of traditional Chinese medicine theory, and have high safety and broad clinical application prospects. The basic prescription of Hua Xian Fang is established by Professor Lin Bing, a famous TCM doctor in Sichuan Province, based on years of clinical experience. It has been used in clinical practice for more than ten years and has great potential for the early prevention and treatment of RIPF. A small sample clinical trial of 34 patients showed that Hua Xian Fang can significantly reduce the incidence of RILI in patients receiving chest radiotherapy at 6 months and 1 year, and relieve symptoms such as cough, sputum, and dyspnea, with total effective rates of 85.3% and 94.1%, respectively. Further, the team of the present application proved the mechanism of action of Hua Xian Fang by inhibiting the pro-fibrotic function of macrophages, thereby inhibiting the overactivation of fibroblasts and the accumulation of extracellular matrix, based on transcriptomics and in vitro and in vivo experiments. In addition, the main active ingredients of Hua Xian Fang, such as 5,7,8-trimethoxyflavone, hyperoside, helicin, and isoflavonoid, were screened by serum drug chemistry detection and network pharmacology analysis.

[0004] However, many effective components of Hua Xian Fang, including the above-mentioned components, such as flavonoids and saponins, have the problem of poor water solubility and low absorption efficiency in the human body, which limits the efficacy of traditional dosage forms such as decoctions. In recent years, the widespread application of nanotechnology in the field of drug delivery has provided new strategies for the development of poorly soluble active ingredients of traditional Chinese medicine. The application of nanomodification technology can significantly improve the water solubility of poorly soluble active substances, enhance their stability, and prolong their retention time in the body, thereby improving the bioavailability and efficacy of drugs. Among them, nanomicelles, as an advanced nanodelivery system, have unique advantages in the delivery of active ingredients of traditional Chinese medicine due to their simple preparation process, excellent stability, and low toxicity. This new drug delivery system provides a new research idea and technical support for the efficient use of poorly soluble components of traditional Chinese medicine.

[0005] PEG-PCL (polyethylene glycol-poly caprolactone) nanomicelles, as a new type of nanocarrier system, exhibit many excellent properties. Its high biocompatibility and biodegradability make it have good safety, and the PEG part prolongs the blood circulation time and reduces the risk of immune system clearance, while the PCL part can be degraded by in vivo hydrolysis to avoid toxicity accumulation and ensure the safety of long-term use. PEG-PCL nanomicelles also have excellent water solubility and stability, can efficiently load hydrophobic drugs, improve their bioavailability, and achieve sustained release of drugs through controlled release mechanism, thereby significantly enhancing the therapeutic effect. In addition, the hydrophilic shell of PEG endows it with the ability to achieve passive targeted delivery through the enhanced permeability and retention effect (EPR effect), and combined with specific active targeting strategies can further optimize the targeting performance, increase the drug concentration at the lesion site, and reduce the toxic side effects on normal tissues.

[0006] However, at present, there is no reported technology that can be applied to improve the physicochemical properties of Huaxianfang, thereby improving its water solubility and bioavailability to improve its clinical use value. Therefore, the development of a self-assembled Huaxianfang nanomicelle (NHXF) based on PEG-PCL can solve the problem of poor water solubility and low bioavailability of multiple effective components of Huaxianfang, and further improve the efficacy of Huaxianfang, which has important theoretical significance and practical application value.

[0007] The inventors' previous publications have fully demonstrated the preventive and therapeutic effects of Huaxian formula on RILI (Lin Bing, Zhang Peng, Lang Jinyi. Clinical observation on the efficacy of Huaxian decoction in preventing and treating 34 cases of radiation-induced lung injury. Sichuan Journal of Traditional Chinese Medicine. 2012;30(07):76-8., Chen Junyang, Zou Pingjin, Fang Zengyi, et al. Huaxian formula alleviates radiation-induced pulmonary fibrosis by upregulating peripheral and tissue IFN-γ levels [J]. Chinese Journal of Radiation Oncology, 2024,33(6):554-561, Chen J, Zou P, Quan L, et al. Huaxian formula prevents the progression of radiation-induced pulmonary fibrosis by inhibiting the pro-fibrotic effects of macrophages. JEthnopharmacol. 2025 Feb 10;338(Pt 2):119026., Gong C, Chen J, Zou P et al. Serum Pharmacochemistry and Network Pharmacology Reveal Active Compounds and Mechanisms of the Huaxian Formula in Alleviating Radiation-Induced Pulmonary Fibrosis[J]. Drug Des Devel Ther. 2025:19:627-644. doi: 10.2147 / DDDT.S490844.), but this formula has a complex composition and high cost. The benefits obtained depend on the patient's long-term adherence to medication. However, traditional Chinese medicine decoctions have drawbacks such as a special odor, poor taste, and troublesome decoction, which may make it difficult for patients to adhere to medication, thus limiting its efficacy and widespread application. Therefore, it is necessary to improve the dosage form of the Huaxian Formula to improve its therapeutic efficiency.

[0008] For lung diseases, nebulized inhalation formulations have the advantages of small dosage, rapid effect, and good targeting. In addition, the application of traditional Chinese medicine nebulized formulations in acute and chronic lung diseases has been extensively studied in clinical practice. At present, the main preparation method of traditional Chinese medicine nebulized inhalation formulations is to directly concentrate the decoction of traditional Chinese medicine to a certain concentration and sterilize it before using it for nebulized inhalation in patients (Chen Juan. Clinical study on the treatment of acute viral pharyngitis by combining Qingre Liyan Decoction with ultrasonic nebulization of traditional Chinese medicine [J]. Journal of Integrated Traditional and Western Medicine, 2025, 17(03): 165-168., Chen Jie, Guo Yu. Clinical observation on the treatment of acute rhinosinusitis of the lung meridian wind-heat type by ultrasonic nebulization of Tongqiao Sanjie Decoction [J]. Chinese Journal of Integrated Traditional and Western Medicine Otorhinolaryngology, 2023, 31(04): 257-263.DOI:10.16542 / j.cnki.issn.1007-4856.2023.04.005.). The primary problem with this type of technology is that the main active ingredients in many traditional Chinese medicine formulas are flavonoids, saponins, and other components that are difficult to dissolve in water. This makes it difficult for the human body to absorb and utilize simple traditional Chinese medicine inhalers, thus limiting their efficacy.

[0009] Therefore, when preparing nebulized inhalers using traditional Chinese medicine decoctions, their physicochemical properties, such as water solubility and stability, should be modified and improved first to achieve the expected effects of the new dosage form. Nanomaterial modification technology is widely used in the field of drug delivery, and its application in improving the bioavailability, solubility, and stability of drugs is gradually attracting attention. However, research on this technology in the field of traditional Chinese medicine is mostly limited to the modification of single active ingredients (Wang H, Jiao Y, Ma S, et al. Nebulized inhalation of peptide-modified DNA origami to alleviate acute lung injury. Nano Lett. 2024;24(20):6102–6111.), which makes traditional Chinese medicine lose its advantage of multi-component efficacy, thus neglecting the essential and pursuing the trivial. Therefore, a technology is needed to improve the water solubility and bioavailability of the overall components of traditional Chinese medicine compound extracts in order to fully exert their therapeutic effects. Summary of the Invention

[0010] To address the problems existing in the prior art, this invention utilizes the self-assembly properties of high molecular weight PEG5000-PCL10000 to provide a sufficiently large hydrophobic core and a stable hydrophilic shell, thus preparing a nanomicelle-type synthetic fiber nebulizer. This nebulizer can carry a large number of components from the synthetic fiber and remain stable in water. By modifying the nebulizer formulation, the economic and time costs for patients can be reduced, and the overall effectiveness of the synthetic fiber in improving radiation-induced lung injury can be improved.

[0011] To achieve the above-mentioned technical objectives, the present invention adopts the following technical solution: One objective of this invention is to provide a polyethylene glycol-polycaprolactone-loaded nanomicelle-type chemical fiber nebulizer, comprising nanomicelles formed by nanomicelle assembly technology of chemical fiber extract and polyethylene glycol-polycaprolactone copolymer. The chemical fiber extract is prepared from the following traditional Chinese medicine raw materials in the following weight ratios: honeysuckle 14-16g, scutellaria baicalensis 8-12g, bamboo shavings 14-16g, coltsfoot flower 14-16g, astragalus membranaceus 18-22g, salvia miltiorrhiza 18-22g, safflower 4-6g, notoginseng powder 4-6g, and licorice 8-10g.

[0012] Furthermore, the molecular weight of the polyethylene glycol-polycaprolactone copolymer is PEG 4~6K-PCL 8~12K.

[0013] Furthermore, the concentration of the nanomicelle type chemical fiber nebulized inhaler is 10 μg / ml to 10 mg / ml.

[0014] The second objective of this invention is to provide a method for preparing the aforementioned nano-micelle type chemical fiber nebulized inhalant, comprising the following steps: (1) Preparation of chemical fiber extract; (2) Nano micelles were prepared by combining the extract of chemical fiber with polyethylene glycol-polycaprolactone copolymer through a nano micelle assembly process; (3) Prepare nanomicelles into lyophilized powder form; (4) Dissolve the lyophilized powder in sterile deionized water to prepare a nebulized inhalant.

[0015] Furthermore, the preparation of the chemical fiber extract in step (1) includes: (a) Soaking the Chinese herbal raw material as described in claim 1 in clean water; (b) Combine the decoctions after boiling and concentrate them; (c) Ultrasonic oscillation extraction; (d) Filter to obtain the extract.

[0016] Furthermore, the decoction process described in step (b) involves two decoctions, each lasting 40 to 60 minutes, and the combined decoctions are concentrated to 80 to 120 ml.

[0017] Furthermore, the assembly of nanomicelles in step (2) includes: (a) Freeze-dry the extract of the chemical fiber into freeze-dried powder; (b) Dissolve the lyophilized powder and the polyethylene glycol-polycaprolactone copolymer in acetone at a mass-to-volume ratio of 1:(8~12); (c) Add the above solution dropwise to an equal volume of deionized water and use ultrasonic oscillation to promote assembly; (d) Preliminary removal of acetone by rotary evaporation; (e) Dialysis to remove free substances; (f) Add trehalose as a freeze-drying protectant.

[0018] Furthermore, the freeze-drying process described in step (a) includes: pre-freezing at -70~-90℃ for 22~26 hours, then freeze-drying at -30~-40℃ and 80~120Pa for 10~14 hours, and then freeze-drying at 24~26℃ and 8~12Pa for 6~10 hours.

[0019] Furthermore, after pre-freezing the dialyzed nanomicelles in step (f), they are freeze-dried at -30 to -40°C and 18 to 22 Pa for 22 to 26 hours, and then freeze-dried at 24 to 26°C and 4 to 6 Pa for 10 to 14 hours to obtain freeze-dried powder.

[0020] The third objective of this invention is to provide the application of the aforementioned nanomicelle-type chemical fiber nebulized inhaler in the preparation of products for treating lung diseases.

[0021] The main innovative points of this invention are as follows: 1. Formula of Huaxianfang: Huaxianfang is an experienced formula of Professor Lin Bing, a famous traditional Chinese medicine doctor in Sichuan Province and affiliated hospital of Chengdu University of Traditional Chinese Medicine. The formula is based on the understanding of RILI and related pulmonary diseases by physicians throughout history and combined with Professor Lin Bing’s decades of clinical experience and understanding of the disease. It has been used in clinical practice for more than ten years with good results.

[0022] 2. Chemical fiber nanomicelles and their preparation process: This is the core of the present invention. The chemical fiber traditional Chinese medicine formula is optimized, a suitable nanomaterial loading and modification process is designed, and a vacuum freeze-drying technology is adopted that can completely evaporate water and acetone while maintaining the stability of its nanostructure. This improves the water solubility and bioavailability of the active ingredients of the traditional Chinese medicine compound, while reducing the economic and time costs of using traditional Chinese medicine. Furthermore, the conditions of the present invention are mild and easy to transform and develop.

[0023] 3. Application areas: The chemical fiber nebulized inhaler developed with this technology can be used in the early stages of patients receiving thoracic radiotherapy to prevent and treat the onset and progression of RILI. After drug group dosage adjustment, it can also be used to improve symptoms in patients with advanced RILI.

[0024] Compared with the prior art, the present invention has the following advantages: 1. The traditional Chinese medicine formula of the chemical fiber formula has been optimized: Although the traditional chemical fiber formula contains multiple effects such as clearing heat and resolving phlegm, invigorating qi and nourishing yin, and promoting blood circulation and removing blood stasis, its composition is complex, containing nearly 20 Chinese herbs, resulting in high costs. Furthermore, clinical use often involves adjustments to the formula, making it difficult to determine the specific composition and hindering industrial production. This technology optimizes the chemical fiber formula, clearly defining the key active ingredient groups for radiation-induced lung injury based on the original formula: honeysuckle, scutellaria, bamboo shavings, and coltsfoot flower for clearing heat and resolving phlegm; and astragalus, salvia miltiorrhiza, safflower, and notoginseng powder for invigorating qi and promoting blood circulation. The advantages of this optimized formula are its simple composition, specific ingredients, and strong targeting. Different types of inhalers can be prepared by adjusting the dosage ratio of the heat-clearing and phlegm-resolving group or the qi-invigorating and blood-activating group, applicable to different stages of radiation-induced lung injury.

[0025] 2. The extract of the chemical fiber formula has a complex composition, mainly containing triterpenoids and steroid-like components with large molecular sizes and strong hydrophobicity. Therefore, PCL with a molecular weight of 10KD is selected to form a stable hydrophobic core, which is beneficial for the co-loading of multiple components and increases drug loading. Simultaneously, PEG with a molecular weight of 5KD is selected to form a stable hydration layer without interfering with the formation of self-packed components, while controlling the particle size. Under this technology, the active ingredients of the chemical fiber formula can be encapsulated within the material, forming spherical or near-spherical nanoparticles that are stably dissolved in water in a micelle state. This technology can enhance the water solubility of the active ingredients in the chemical fiber formula, increase bioavailability, and further improve efficacy.

[0026] 3. Based on vacuum freeze-drying, the concentration of the nano-micelle-type chemical fiber nebulized inhaler can be flexibly controlled. Considering that vacuum freeze-drying can damage the already formed nanoparticle structure, this technology uses a programmed cooling method during pre-freezing to reduce ice crystal formation, while adding a certain concentration of trehalose to maintain structural stability. This freeze-dried powder is prepared from an aqueous solution; the acetone used in the production process is completely volatile, resulting in high water solubility. It can be directly dissolved in sterile deionized water and shaken to mix, eliminating the need for secondary sonication, making it convenient to use. This technology also improves the chemical fiber formulation, reducing the time and cost of decoction for patients and avoiding the impact of the odor and taste of traditional Chinese medicine on patient compliance.

[0027] 4. Based on nanomicelle-type chemical fiber nebulized inhalers, new strategies can be provided for the early prevention and treatment of clinical RILI, reduce lung tissue damage caused by chest radiotherapy, inhibit the progression of RILI, avoid the occurrence of RIPF, and may increase the chest radiotherapy dose for patients, thereby improving the treatment effect. It has great translational application value.

[0028] 5. The nano-modification of the main body and chemical fiber of this invention is based on the self-assembly characteristics of PCL-PEG. It does not require high temperature, high pressure and catalyst, and can be carried out at room temperature. The preparation conditions are low, the preparation process is simple, the cost is low, and it is easy to realize industrial production, which has good economic benefits. Attached Figure Description

[0029] Figure 1 This is a flowchart illustrating the preparation process of NHXF in Example 1 of the present invention.

[0030] Figure 2 This demonstrates that NHXF at different concentrations can exhibit a significant Tyndall effect in Example 1 of the present invention.

[0031] Figure 3 This is a comparison of the particle size of NHXF and chemical fiber extract (HXF) in Example 1 of the present invention.

[0032] Figure 4 This is a comparison of the zeta potentials of NHXF and HXF in Example 1 of the present invention.

[0033] Figure 5 This is a comparison of the electron microscopy morphology of particles in NHXF and HXF in Example 1 of the present invention.

[0034] Figure 6 The UV absorption spectra of NHXF, HXF, and nanomaterials in Example 1 of this invention are shown.

[0035] Figure 7 This is a chromatogram of the anion and cation peaks of NHXF detected by HPLC-MS in Example 1 of the present invention.

[0036] Figure 8 The comparison results between the three key components identified in NHXF in Example 1 of this invention and the database are shown.

[0037] Figure 9 This is a comparison of the solubility concentrations of the main indicator components of HXF and NHXF in Example 2 of the present invention.

[0038] Figure 10 This is a comparison of the signal intensity distribution of various organs in the body after nebulized inhalation of NHXF with free drugs in Example 2 of the present invention.

[0039] Figure 11 The results of CCK-8 incubation with different concentrations of NHXF in Example 3 of this invention are shown.

[0040] Figure 12 Examples of ROS fluorescence intensity after different treatments in Example 3 of this invention.

[0041] Figure 13The above are example scatter plots and statistical analysis bar charts for apoptosis detection after different treatments in Example 3 of the present invention.

[0042] Figure 14 The images shown are examples of HE staining in different groups in Example 3 of this invention. Scale bar = 50 μm.

[0043] Figure 15 The levels of three inflammatory factors, TNF-α, IL-6, and IL-1β, in serum and bronchoalveolar lavage fluid in Example 3 of this invention. Detailed Implementation

[0044] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention. The reagents, products, and instruments used in the following examples are all commercially available, and the methods used in the examples, unless otherwise specified, are consistent with conventionally used methods.

[0045] The technical solution of the present invention will be further described in detail below with reference to the embodiments.

[0046] Example 1: Preparation and Characterization of NHXF 1. Improved herbal formula for chemical fiber treatment: Honeysuckle 15g, Scutellaria baicalensis 10g, Bambusa textilis 15g, Coltsfoot flower 15g, Astragalus membranaceus 20g, Salvia miltiorrhiza 20g, Safflower 5g, Panax notoginseng powder 5g, Licorice 9g.

[0047] 2. Preparation of nanomicelle-type chemical fiber nebulized inhaler (NHXF): 2.1 Preparation of chemical fiber extract using standardized traditional Chinese medicine decoction process: (1) All of the above-mentioned drugs must be purchased from qualified hospitals or pharmacies, and the quality and weight of the drugs must be guaranteed to be good. (2) Soak the above-mentioned drugs in 500ml of water for 30 minutes; (3) Use non-metallic inert utensils (ceramic pot, earthenware pot, beaker, etc.) to decoct the above-mentioned Chinese medicine. After boiling, continue to decoct for 30 minutes. (4) Pour out the first decoction, add 500ml of water again and boil for 30 minutes; (5) Mix the two decoctions and concentrate the liquid to 100ml; (6) Further extract components by ultrasonic oscillation for 60 min; (7) Filter the chemical fiber extract with filter paper to remove non-drug precipitates and obtain the chemical fiber extract.

[0048] 2.2 Preparation of chemical fiber square nanomicelles: (1) The filtered chemical fiber extract was pre-frozen at -80℃ for 24 hours; (2) The freeze-dried powder was prepared using a vacuum freeze dryer. The freeze-drying parameters were: freeze-drying at -35℃ and 100Pa for 12 hours to remove free water, and freeze-drying at 25℃ and 10Pa for 8 hours to remove bound water. Equal masses of the freeze-dried powder of chemical fiber and PEG-PCL powder (molecular weight: PEG 5K-PCL 10K) were weighed and dissolved in acetone at a mass-volume ratio of 1:10. (3) While stirring, slowly add the above liquid to deionized water of the same volume as acetone, and sonicate for 60 min to assist in the efficient assembly of nanomicelles; (4) Evaporate acetone for 30 minutes in a water bath at 37°C and a vacuum of 200 mbar to initially volatilize it; (5) Dialyze in deionized water using a 15KD dialysis membrane for 12 hours, changing the deionized water medium every two hours to remove empty carriers and free drugs; (6) Add trehalose, a freeze-drying protectant, at a mass-volume ratio of 3% to stabilize the nanostructure; (7) After dialysis, the drug was placed at -80°C and slowly cooled (1°C per minute) for 24 hours using a programmed cooling box containing isopropanol for pre-freezing treatment. (8) Use a freeze dryer to prepare freeze-dried powder from the dialyzed micelles. The freeze-drying parameters are: freeze-drying at -35℃ and 20Pa for 24h to remove free water and completely evaporate acetone, freeze-drying at 25℃ and 5Pa for 12h to remove bound water and bound acetone. The freeze-dried powder should be stored at -80℃. 2.3 Preparation of nebulized inhalant: Weigh an appropriate amount of lyophilized fiber nanomicelle powder and dissolve it directly in sterile deionized water in an appropriate ratio. Depending on the requirements, different concentrations of 10 μg / ml, 50 μg / ml, 100 μg / ml up to 10 mg / ml of cellulose nanomicelle nebulized inhalant can be prepared for basic experiments or clinical trials. In basic experiments, a concentration of 100 μg / ml can be selected for cell experiments, and a concentration of 50 μg / ml can be selected for mouse experiments. There is no evidence for the concentration in clinical trials. Based on mouse experiments, it is speculated that 5 mg / ml or 10 mg / ml can be selected.

[0049] The preparation process flow chart of NHXF is as follows: Figure 1 As shown.

[0050] 3. Physicochemical characterization of nanomicelles in chemical fibers: 3.1 Tyndall effect: At room temperature, when laser light passes through NHXF of different concentrations, a clear straight beam is observed, exhibiting a significant Tyndall effect, indicating that the colloid has good stability. Figure 2 ).

[0051] 3.2 Particle Size: The particle size of the micelles was determined using a Malvern particle size analyzer under simulated body fluid conditions (37℃, pH=7.35). Compared to traditional chemical fiber extracts, the particle size of NHXF was significantly reduced, and the PDI value increased, indicating that the components in NHXF are dispersed in water as smaller and more uniform particles. Figure 3 ).

[0052] 3.3 Zeta potential: Under simulated body fluid conditions (37℃, pH=7.35), the absolute value of the zeta potential of NHXF was significantly increased compared to that of traditional chemical fiber extracts, with an average value of -23.4mV, indicating that its system is more stable and less prone to aggregation or precipitation. Figure 4 ).

[0053] 3.4 Transmission Electron Microscopy Images: NHXF was observed using a transmission electron microscope. It was found to be spherical with a particle diameter less than 200 nm, while the particles in the aqueous extract of the chemical fiber were irregular in shape and larger. Figure 5 ).

[0054] 3.5 UV Absorption Spectroscopy: UV-Vis analysis was performed on the extract of the chemical fiber compound and the chemical fiber nanomicelles. The results showed that the absorption spectra of the two were roughly the same, indicating that NHXF effectively retained the components of the chemical fiber compound extract and exhibited a good multi-component synergistic effect. Figure 6 ).

[0055] 3.6 Drug components: Based on high-performance chromatography-mass spectrometry (HPLC-MS) analysis and database comparison, 24 active components in NHXF were identified. Figure 7 (Table 1) The three main components present in higher amounts are L-amygin, hesperidin, and baicalin ( Figure 8 ).

[0056] Table 1 Main active ingredients of NHXF

[0057] Example 2: In vitro and in vivo bioavailability of NHXF 1. In vitro solubility of key indicator components: 1 mg of NHXF lyophilized powder and an equivalent dose of HXF lyophilized powder (0.5 mg, the lyophilized powder of the pure chemical fiber formula obtained in step 2.2-(2) of Example 1) were weighed and dispersed in 1 ml of PBS. The mixture was placed in a constant temperature shaker at 37 ℃ to simulate in vivo absorption. After shaking for 3 h, the mixture was centrifuged at 12,000 rpm for 10 min and the supernatant was collected. The absorbance was measured at the λmax wavelength using ultraviolet spectrophotometry, and the solubility of the main index components L-amygin, baicalin, and hesperidin in the aqueous phase was calculated according to the standard curve. The experimental results showed that, under the same conditions, the apparent solubility of the main index components L-amygin, baicalin, and hesperidin in the nano micelles of the chemical fiber formula was significantly higher than that of the traditional chemical fiber formula, indicating that NHXF can provide a higher concentration of effective ingredients for in vivo absorption. Figure 9 ).

[0058] 2. Drug utilization in vivo: NHXF was labeled with the fluorescent tag Cy5, with an equivalent dose of free Cy5 as a control. In vivo imaging was used to detect the fluorescence signal intensity in organs. The results showed that compared to free drug, NHXF had a significantly longer residence time and higher fluorescence intensity in the lungs, lasting for more than 10 hours, while free drug was rapidly metabolized and cleared from the lungs within 3 hours. Figure 10 The above results indicate that NHXF comprehensively improves its bioavailability by increasing the solubility of its active ingredient and the absorption concentration during its action time in the target organ, lung tissue.

[0059] Example 3: NHXF Effect Verification 1. Different concentrations of chemical fiber nanomicelles had no significant effect on the activity of rat lung epithelial cells RLE-6TN.

[0060] 1.1 Experimental Methods Cell line culture: Rat type II alveolar epithelial cells RLE-6TN (purchased from Baidi Biotechnology Co., Ltd., catalog number C5645) were cultured in DMEM medium supplemented with 10% fetal bovine serum and 1% penicillin G and streptomycin antibiotics in a cell culture incubator (CO2 concentration: 5%; temperature: 37℃). When the cell density increased to 80%-90%, the cells were passaged. After cell recovery, the cells were passaged three times for subsequent plate-coating experiments.

[0061] Cell intervention: cells were injected at a rate of 5 × 10⁻⁶. 3 The cells were seeded into 96-well plates at a concentration of / well. After cell adhesion, the appropriate concentration of chemical fiber nanomicelles was added, and the plates were placed in a cell culture incubator for 48 hours.

[0062] Cell viability assay: After cell intervention, the culture medium in the wells was aspirated, and CCK-8 working solution was prepared by diluting CCK-8 reagent with serum-free DMEM at a ratio of 1:10. 100 μl of CCK-8 working solution was added to each well, and the cells were incubated at 37°C in the dark for 1 h. The OD value at 450 nm was measured using a microplate reader, and cell viability was calculated according to the following formula: Cell viability = (OD value of treated wells - OD value of blank wells) / (OD value of control wells - OD value of blank wells) × 100% 1.2 Experimental Results and Conclusions The results showed that cell viability did not change significantly when the concentration of the chemical fiber nanomicelles increased from 0 μg / ml to 200 μg / ml, indicating that the chemical fiber nanomicelles had no significant effect on the activity of normal cells and had good safety. Figure 11 ).

[0063] 2. Chemical fiber nanomicelles can significantly inhibit oxidative stress in RLE-6TN induced by radiotherapy in lung epithelial cells.

[0064] 2.1 Experimental Methods 2.1.1 The cell culture conditions are the same as before.

[0065] 2.1.2 Grouping and Processing: (1) Control group (Ctrl): No radiation; no drug intervention; (2) Radiation group (IR): Radiation (biological irradiator, model: X-RAD 320 (PRECISION, USA), dose rate: 20 Gy); no drug intervention; (3) Traditional chemical fiber formula (IR+THXF): radiation; water extract of traditional chemical fiber formula intervention; (4) Low-dose group of chemical fiber nanomicelles (IR+NHXF 50μg / ml): radiation; intervention with 50μg / ml nanomicelle chemical fiber formula; (5) High-dose group of chemical fiber nanomicelles (IR+NHXF 100μg / ml): radiation; 100μg / ml nanomicelle chemical fiber intervention.

[0066] 2.1.3 Cell Intervention: After cell adhesion, the cells were placed at a rate of 2×10⁶ cells / year. 4 The cells were seeded into 24-well plates at the specified concentration. After cell adhesion, appropriate concentrations (50, 100 μg / ml) of chemical fiber nanomicelles were added, and the cells were cultured in a cell culture incubator for 48 h. The cells were then subjected to a single irradiation of 20 Gy using an irradiator, and ROS generation was detected immediately after irradiation.

[0067] 2.1.4 ROS (Reactive Oxygen Species) Detection: The culture medium in each well was aspirated, and 10 µM DCFH-DA probe working solution was added. The cells were incubated at 37°C in the dark for 60 minutes. After incubation, the cells were gently washed three times with PBS to remove excess probe that had not entered the cells, thus reducing background fluorescence interference. Changes in DCF fluorescence intensity were observed using a fluorescence microscope after washing.

[0068] 2.2 Experimental Results and Conclusions Cells showed a significant increase in ROS intensity after radiation exposure. Intervention with 100 μg / ml IR+NHXF significantly inhibited ROS fluorescence intensity, and the high-dose nanomicelle-type chemical fiber formula was significantly superior to the traditional decoction. This indicates that IR+NHXF can alleviate radiation-induced oxidative damage to lung epithelial cells, and its effect is significantly better than that of the traditional chemical fiber formula. Figure 12 ).

[0069] 3. Chemical fiber nanomicelles can significantly inhibit radiotherapy-induced apoptosis of lung epithelial cells RLE-6TN.

[0070] 3.1 Experimental Methods 3.1.1 Cell culture and grouping intervention conditions are the same as above. 3.1.2 Apoptosis detection of cells 72 h after irradiation: The Annexin V / PI kit was used for detection, and the steps are as follows: (1) Collect adherent and suspended detached cells using trypsin without EDTA; (2) After washing with PBS, add 100 μl of binding buffer to resuspend and mix well; (3) Add 5 μl Annexin V-FITC and incubate at room temperature in the dark for 5 min; (4) Add 5 μl of PI, mix well, and immediately add 400 μl of PBS; (5) After mixing, the samples were analyzed using a full-spectrum flow cytometer and Flowjo software. Annexin V was among the samples analyzed. + PI - Defined as early apoptotic cells, Annexin V + PI + Defined as late-stage apoptotic cells.

[0071] 3.2 Results and Conclusions Following radiation, the level of apoptosis in alveolar epithelial cells significantly increased. Compared to the radiation group, intervention with 100 μg / ml of nano-micelle-type chemical fiber significantly reduced the proportion of apoptotic cells, showing a significantly better effect than the chemical fiber extract and 50 μg / ml of nano-micelle-type chemical fiber. This indicates that the nano-micelle-type chemical fiber can alleviate radiation-induced epithelial cell apoptosis in lung epithelial cells, with a significantly better effect than the traditional chemical fiber formula, and 100 μg / ml is a more optimal dose than 50 μg / ml. Figure 13 ) 4. Nebulized inhalation of nanomicelle-type synthetic fiber formula can significantly alleviate the severity of radiation-induced lung injury in mice.

[0072] 4.1 Experimental Methods 4.1.1 Experimental Animals: Ten-week-old SPF-grade male C57BL / 6 mice (purchased from Beijing Huafukang Biotechnology Co., Ltd., product name C57BL / 6J Mice) were used. The mice were housed in an SPF-grade animal room with the ambient temperature controlled at 22±2℃ and the air humidity at 40%-70%. The experimental process followed the relevant regulations such as the "Regulations on the Management of Experimental Animals of the People's Republic of China" to strictly ensure animal welfare.

[0073] 4.1.2 Radiation: After anesthesia, mice were fixed and subjected to a single 20 Gy radiation to the right lung using a biological irradiator (model: X-RAD 320 (PRECISION, USA)) to establish a radiation-induced lung injury model.

[0074] 4.1.3 Grouping and Intervention: (1) Control group (Ctrl): No radiation + saline nebulized inhalation; (2) Drug control group (NHXF): Non-radioactive + high-dose NHXF (50μg / ml, 4ml / mouse) nebulized inhalation; (3) Radiation group (IR): Radiation + saline nebulization inhalation; (4) Chemical fiber extract atomization group (IR+HXF): radiation + chemical fiber extract atomization inhalation (50μg / ml, 4ml / mouse); (5) Low-dose group of chemical fiber nano-nebulizer (IR+NHXF-L): radiation + low-dose NHXF (20μg / ml, 4ml / mouse) nebulized inhalation; (6) High-dose group of chemical fiber nano-nebulizer (IR+NHXF-H): radiation + high-dose NHXF (50μg / ml, 4ml / mouse) nebulized inhalation; (7) Blank vector group (IR+PEG-PCL): Radiation + blank vector (50μg / ml, 4ml / mouse) nebulized inhalation.

[0075] 4.1.4 HE staining: After dewaxing and hydration, paraffin sections were stained with hematoxylin, rinsed with running water, and differentiated to regain blue color. Subsequently, cytoplasm was stained with eosin, dehydrated, cleared, and mounted. Tissue morphology was observed under a microscope.

[0076] 4.2 Results and Conclusions HE-stained lung tissue sections in the IR group showed severe alveolar structural damage, significant interstitial thickening, and extensive inflammatory cell infiltration. In contrast, the IR+NHXF-H group showed significantly reduced pathological changes and better preservation of alveolar structure, while both the IR+HXF and IR+NHXF-L groups still exhibited significant alveolar structural damage. The results indicate that nebulized inhalation of NHXF can significantly reduce radiation damage to lung tissue, with better efficacy than direct nebulization of the aqueous extract of the chemical fiber. Furthermore, 50 μg / ml is a more preferred dose concentration than 20 μg / ml. Figure 14 ).

[0077] 5. Nebulized inhalation of nanomicelle-type chemical fiber formula can significantly reduce the levels of local and systemic inflammatory factors in mice with radiation-induced lung injury.

[0078] 5.1 Experimental Methods: 5.1.1 Experimental animals and group interventions are the same as above. 5.2.2 The levels of inflammatory factors IL-6, IL-1β and TNF-α in bronchoalveolar lavage fluid and blood of mice were detected using an ELISA kit.

[0079] 5.2 Results and Conclusions: Detection of inflammatory cytokine levels in serum and bronchoalveolar lavage fluid supernatant showed that TNF-α, IL-6, and IL-1β levels were significantly elevated in the IR group; in contrast, the levels of these inflammatory factors were significantly decreased in the IR+NHXF-H group, approaching normal control levels. Relatively speaking, while the IR+HXF and IR+NHXF-L groups could inhibit the increase of inflammatory factors to some extent, the inhibitory effect was limited. The overall results showed a consistent trend, indicating that nebulized inhalation of NHXF can effectively reduce radiation-induced inflammatory responses, and its anti-inflammatory effect is superior to that of direct nebulization of the chemical fiber aqueous extract. Furthermore, at the set dosage, 50 μg / mL showed a more significant inhibitory effect than 20 μg / mL. Figure 15 ).

[0080] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A polyethylene glycol-polycaprolactone-loaded nanomicelle-type chemical fiber nebulizer, characterized in that, The nanomicelles are formed by nanomicelle assembly technology of chemical fiber extract and polyethylene glycol-polycaprolactone copolymer. The chemical fiber extract is prepared from the following Chinese herbal raw materials in the following weight ratio: honeysuckle 14-16g, scutellaria baicalensis 8-12g, bamboo shavings 14-16g, coltsfoot flower 14-16g, astragalus membranaceus 18-22g, salvia miltiorrhiza 18-22g, safflower 4-6g, notoginseng powder 4-6g, and licorice 8-10g.

2. The nano-micelle type chemical fiber square atomized inhalant according to claim 1, characterized in that, The molecular weight of the polyethylene glycol-polycaprolactone copolymer is PEG 4~6K-PCL 8~12K.

3. The nano-micelle type chemical fiber square atomized inhalant according to claim 2, characterized in that, The concentration of the nanomicelle type chemical fiber nebulized inhaler is 10 μg / ml to 10 mg / ml.

4. The preparation method of the nanomicelle-type chemical fiber square atomized inhalant according to any one of claims 1-3, characterized in that, Includes the following steps: (1) Preparation of chemical fiber extract; (2) Nano micelles were prepared by combining the extract of chemical fiber with polyethylene glycol-polycaprolactone copolymer through a nano micelle assembly process; (3) Prepare nanomicelles into lyophilized powder form; (4) Dissolve the lyophilized powder in sterile deionized water to prepare a nebulized inhalant.

5. The preparation method according to claim 4, characterized in that, The preparation of the chemical fiber extract in step (1) includes: (a) Soaking the Chinese herbal raw material as described in claim 1 in clean water; (b) Combine the decoctions after boiling and concentrate them; (c) Ultrasonic oscillation extraction; (d) Filter to obtain the extract.

6. The preparation method according to claim 5, characterized in that, The decoction process described in step (b) involves two decoctions, each lasting 40 to 60 minutes. The decoctions are then combined and concentrated to 80 to 120 ml.

7. The preparation method according to claim 4, characterized in that, The assembly of nanomicelles in step (2) includes: (a) Freeze-dry the extract of the chemical fiber into freeze-dried powder; (b) Dissolve the lyophilized powder and the polyethylene glycol-polycaprolactone copolymer in acetone at a mass-to-volume ratio of 1:(8~12); (c) Add the above solution dropwise to an equal volume of deionized water and use ultrasonic oscillation to promote assembly; (d) Preliminary removal of acetone by rotary evaporation; (e) Dialysis to remove free substances; (f) Add trehalose as a freeze-drying protectant.

8. The preparation method according to claim 7, characterized in that, The freeze-drying process described in step (a) includes: pre-freezing at -70~-90℃ for 22~26 hours, then freeze-drying at -30~-40℃ and 80~120Pa for 10~14 hours, and then freeze-drying at 24~26℃ and 8~12Pa for 6~10 hours.

9. The preparation method according to claim 7, characterized in that, After pre-freezing, the dialyzed nanomicelles in step (f) are freeze-dried at -30~-40℃ and 18~22Pa for 22~26 hours, and then freeze-dried at 24~26℃ and 4~6Pa for 10~14 hours to obtain freeze-dried powder.

10. The use of the nanomicelle type chemical fiber nebulized inhaler according to any one of claims 1 to 3 in the preparation of products for treating lung diseases.

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