Use of polygonatum polysaccharide in preparation of products for preventing and / or treating lung injury

Abstract

The present application relates to the application of Huangjing polysaccharide in the preparation of products for preventing and / or treating lung injury, and belongs to the technical field of traditional Chinese medicines. The application of Huangjing polysaccharide in the preparation of products for preventing and / or treating lung injury, wherein the products are drugs or health products. The present application is based on the traditional Chinese medicine theory of treating disease before it occurs, comprehensively analyzes the clinical application characteristics of traditional Chinese medicines in treating lung injury, selects yin-nourishing and qi-tonifying traditional Chinese medicines that enter the lung meridian as the research object, and uses the constructed in-vivo lung injury chemical prevention evaluation system to screen different lung protection activities of Huangjing polysaccharide. Experimental research shows that Huangjing polysaccharide can obviously improve lung injury and has a protective effect on the lung, especially for acute lung injury.

Description

Technical Field

[0001] This invention relates to the application of Polygonatum polysaccharide in the preparation of products for the prevention and / or treatment of lung injury, and belongs to the field of traditional Chinese medicine technology. Background Technology

[0002] Lung injury is a group of diseases involving multiple pathological mechanisms, including acute lung injury, meconium aspiration syndrome, pneumoconiosis, radiation-induced lung injury, and mechanical ventilation-related lung injury. From the perspective of traditional Chinese medicine, the lungs are considered a delicate organ, externally connected to the skin and hair, internally connecting to all the blood vessels, governing respiration, and regulating the body's functions. Lung qi is easily deficient, intolerant to cold and heat, and easily damaged by external factors. Its physiological functions are easily impaired by the invasion of internal and external pathogenic factors. From a pathological perspective, lung injury is mostly caused by external pathogenic factors or internal factors. External pathogenic factors include wind-heat, damp-heat, and phlegm, while internal factors are mostly related to insufficient qi and blood, and imbalance of yin and yang. The pathological nature is mainly a mixture of deficiency and excess, with heat toxicity being the core pathogenesis throughout the entire process of lung injury, while phlegm and blood stasis play important roles at different stages. Therefore, tonifying qi and nourishing yin, and promoting blood circulation and removing blood stasis are important principles throughout treatment. Clinically, corresponding treatments should also follow the basic principles of nourishing yin and moistening the lungs, tonifying qi and removing blood stasis, and clearing heat and detoxifying.

[0003] Currently, treatments for lung injury mainly include interventions such as glucocorticoids, bronchodilators, and mechanical ventilation. While these are effective, long-term use can lead to immunosuppression, secondary infections, and metabolic disorders. Traditional Chinese medicine (TCM) compound preparations have shown advantages in treating lung injury, including multiple targets and fewer side effects. Herbs with qi-tonifying and blood-activating effects, such as Astragalus membranaceus and Salvia miltiorrhiza, are often used in these formulas. However, existing TCM preparations generally suffer from limitations such as complex components, inconsistent quality control standards, and unclear efficacy bases. Therefore, developing a drug and / or health product with clearly defined components, high safety, good prognosis, and the ability to both treat and prevent lung injury is of significant clinical importance. Summary of the Invention

[0004] To address the aforementioned problems in existing technologies, this invention provides the application of Polygonatum polysaccharides in the preparation of products for the prevention and / or treatment of lung injury. Experimental studies have shown that Polygonatum polysaccharides can significantly improve lung injury and have a protective effect on the lungs, especially in cases of acute lung injury.

[0005] To achieve the above-mentioned objectives, the present invention adopts the following technical solution:

[0006] The application of Polygonatum polysaccharide in the preparation of products for the prevention and / or treatment of lung injury, wherein the products are pharmaceuticals or health products.

[0007] Furthermore, the lung injury is one or more of the following: acute lung injury, meconium aspiration syndrome, pneumoconiosis, radiation-induced lung injury, or mechanical ventilation-related lung injury.

[0008] Preferably, the lung injury is acute lung injury.

[0009] Furthermore, the monosaccharide composition of the Polygonatum polysaccharide includes fructose, rhamnose, galactose, glucose, mannose, and xylose; the contents of fructose, rhamnose, galactose, glucose, mannose, and xylose are 0.24~0.50 mg / g, 0.79~1.49 mg / g, 0.75~1.52 mg / g, 0.23~0.72 mg / g, 3.79~5.28 mg / g, and 0.10~0.78 mg / g, respectively.

[0010] In the above technical solution, the Polygonatum polysaccharide is obtained by extraction using conventional extraction methods followed by extraction and column chromatography separation.

[0011] Furthermore, the conventional extraction method is one or more of ultrasonic extraction, decoction, heating and reflux extraction, percolation extraction, or microwave extraction, wherein the extraction solvent is water; the number of extractions is 3-5 times; the extraction time for each extraction is 60-120 min; and the volume-to-mass ratio of the extraction solvent to Polygonatum is 2-5 L : 1 kg.

[0012] Furthermore, the conventional separation method is one or both of D101 macroporous resin adsorption method or AB-8 macroporous resin adsorption method, wherein the elution solvent is one or more of water or ethanol solution with a volume fraction of 30%-90%.

[0013] The Polygonatum polysaccharide of the present invention is preferably prepared by the following method: (1) Weigh out the Polygonatum according to the selected weight proportions, add water, soak, decoct, filter, and obtain filtrate and dregs; (2) Precipitate the filtrate obtained in step (1) with alcohol, and collect the supernatant and precipitate; (3) Disperse the supernatant obtained in step (2) with water, extract with ethyl acetate and n-butanol, and collect the n-butanol layer sample; (4) Pass the n-butanol layer sample obtained in step (3) through D101 macroporous resin and elute it sequentially with pure water, 50% ethanol aqueous solution and 90% ethanol aqueous solution, and collect the water elution portion. (5) Combine the water-washed portion obtained in step (4) and the precipitate obtained in step (2), remove the protein, and dry to obtain Polygonatum polysaccharide.

[0014] Furthermore, in step (1), the soaking time is 60-120 min.

[0015] Furthermore, in step (1), the simmering time is 60-120 min.

[0016] Furthermore, in step (1), the boiling process is carried out 3-5 times.

[0017] Furthermore, in step (2), the alcohol precipitation is performed 3-5 times.

[0018] Furthermore, in step (3), ethyl acetate and n-butanol are used for extraction 3-5 times each.

[0019] Furthermore, in step (5), the deproteinization is performed 1-5 times.

[0020] Furthermore, deproteinization can be carried out using conventional methods known in the art, such as one or more of the Sevage method, TCA method, or enzymatic hydrolysis method.

[0021] Furthermore, the methods for analyzing the composition and determining the content of polysaccharides are one or more of the following: high performance liquid chromatography, gas chromatography, ion chromatography, and thin-layer chromatography.

[0022] The beneficial effects of this invention are as follows: This invention provides the application of Polygonatum polysaccharide in the preparation of products for the prevention and / or treatment of lung injury. Preferably, the lung injury includes, but is not limited to, acute lung injury, meconium aspiration syndrome, pneumoconiosis, radiation-induced lung injury, and mechanical ventilation-related lung injury. Based on the traditional Chinese medicine theory of "prevention of disease," this invention comprehensively analyzes the clinical application characteristics of traditional Chinese medicine in treating lung injury, selects yin-nourishing and qi-tonifying traditional Chinese medicines that enter the lung meridian as research objects, and uses a constructed in vivo chemopreventive effect evaluation system for lung injury to screen different lung-protective activities of Polygonatum polysaccharide. Attached Figure Description

[0023] Figure 1 The figure shows the effect of Polygonatum polysaccharide obtained in Example 1 on inflammatory factors and antioxidant enzymes in ALI mice.

[0024] Figure 2 The figure shows the effect of Polygonatum polysaccharide obtained in Example 1 on inflammatory factors and antioxidant enzymes in NPs mice.

[0025] Figure 3 The figure shows the effect of Polygonatum polysaccharide obtained in Example 2 on inflammatory factors and antioxidant enzymes in ALI mice.

[0026] Figure 4 The figure shows the effect of Polygonatum polysaccharide obtained in Example 2 on inflammatory factors and antioxidant enzymes in NPs mice.

[0027] Figure 5 The figure shows the effect of Polygonatum polysaccharide obtained in Example 3 on inflammatory factors and antioxidant enzymes in ALI mice.

[0028] Figure 6 The figure shows the effect of Polygonatum polysaccharide obtained in Example 3 on inflammatory factors and antioxidant enzymes in NPs mice. Detailed Implementation

[0029] The following non-limiting embodiments are intended to enable those skilled in the art to more fully understand the invention, but do not limit the invention in any way.

[0030] Unless otherwise specified, the experimental methods described in the following examples are conventional methods; the reagents and materials described are commercially available unless otherwise specified.

[0031] For experiments not specifically described in the examples, the procedures or conditions should be followed according to the conventional experimental procedures described in the literature in this field. Reagents or instruments whose manufacturers are not specified are all commercially available conventional reagent products.

[0032] All pharmaceutical materials passed quality inspection and met the standards of the Pharmacopoeia of the People's Republic of China.

[0033] Example 1: Polysaccharide Extraction and Separation (1) Take 9 g of Polygonatum and add water, and extract by ultrasonication 3 times. For the first time, add 100 mL of water, soak for 120 min, and sonicate for 120 min. For the second time, add 80 mL of water and sonicate for 90 min. For the third time, add 50 mL of water and sonicate for 60 min. Pass through a 100-mesh sieve, combine the filtrates, and obtain an aqueous solution.

[0034] (2) The aqueous decoction was concentrated to obtain a crude polysaccharide solution with a concentration of 10%. The solution was rapidly stirred and slowly precipitated with 80% ethanol solution. The process was repeated three times, and the precipitate and supernatant were collected. The supernatant was evaporated to dryness, dispersed in water, and extracted three times with ethyl acetate and n-butanol, respectively, to obtain an aqueous layer, an ethyl acetate layer, and a n-butanol layer. The n-butanol extract layer was selected by TLC for separation with D101 macroporous resin. The extract was eluted with water, 50% ethanol / water, and 90% ethanol / water in sequence to obtain three fractions with different polarities. The water-eluted fraction was examined by silica gel TLC and combined with the precipitate collected by the alcohol precipitation. The solvent was evaporated and the solution was redissolved with distilled water to obtain a polysaccharide solution, which was pre-cooled in an ice bath for later use. Protein removal was performed using the TCA method. 10% TCA was slowly added dropwise to the polysaccharide solution while stirring slowly until the final TCA concentration reached 5%. The solution was then incubated on ice for 60 min, centrifuged at 10,000 rpm for 20 min, and the supernatant polysaccharide solution was collected. 1 mol / L NaOH was slowly added dropwise to adjust the pH to 7.0. The solvent was evaporated, and the solution was dried to obtain Polygonatum polysaccharide.

[0035] (3) The polysaccharide composition and content of the obtained Polygonatum polysaccharides were analyzed by high performance liquid chromatography. After hydrolysis of the sample, derivatization was performed using PMP. After sample processing, the contents of each monosaccharide were obtained by the instrument operation. The results are shown in Table 1.

[0036] Table 1. Monosaccharide composition analysis and content determination of Polygonatum polysaccharides

[0037] Example 2 Polysaccharide Extraction and Separation (1) Add water to 12 g of Polygonatum and heat and reflux to extract 3 times. Add 100 mL of water for the first time, soak for 120 min, and heat and reflux for 120 min. Add 80 mL of water for the second time, and heat and reflux for 90 min. Add 50 mL of water for the third time, and heat and reflux for 60 min. Pass through a 100-mesh sieve, combine the filtrates, and obtain the water extract.

[0038] (2) The aqueous decoction was concentrated to obtain a 10% crude polysaccharide solution. The solution was rapidly stirred and slowly precipitated with 80% ethanol solution. This process was repeated three times, and the precipitate and supernatant were collected. The supernatant was evaporated of solvent, dispersed in water, and extracted three times with ethyl acetate and n-butanol, respectively, to obtain an aqueous layer, an ethyl acetate layer, and a n-butanol layer. The n-butanol extract layer was selected by TLC for separation using AB-8 macroporous resin. Elution was performed sequentially with water, 50% ethanol / water, and 90% ethanol / water to obtain three fractions of different polarities. The water-eluted fraction was examined by silica gel TLC and combined with the precipitate collected by the alcohol precipitation. After evaporation of solvent, the fraction was reconstituted with pH 7.0 phosphate buffer to obtain a polysaccharide solution, which was then placed in a 50°C water bath for later use. Protein was removed using enzymatic hydrolysis. Papain was added at 3% of the polysaccharide mass, and the reaction was stirred at a constant temperature for 3 h. The mixture was then immediately transferred to a 100°C water bath for 10 min to inactivate the enzyme, followed by cooling to room temperature. Centrifuge at 5000 rpm for 20 min, collect the upper polysaccharide aqueous solution, evaporate the solvent, and dry to obtain Polygonatum polysaccharide.

[0039] (3) The polysaccharide composition and content of the obtained Polygonatum polysaccharides were analyzed by ion chromatography. After hydrolysis of the sample, the contents of each monosaccharide were obtained by the instrument, and the results are shown in Table 2.

[0040] Table 2. Monosaccharide composition analysis and content determination of Polygonatum polysaccharides

[0041] Example 3 Polysaccharide Extraction and Separation (1) Add 15 g of Polygonatum to water and decoct 3 times. The first time, add 150 mL of water, soak for 120 min, and decoct for 120 min. The second time, add 80 mL of water and decoct for 90 min. The third time, add 100 mL of water and decoct for 60 min. Pass through a 100-mesh sieve, combine the filtrates, and obtain the decoction.

[0042] (2) The aqueous decoction was concentrated to obtain a crude polysaccharide solution with a concentration of 10%. The solution was rapidly stirred and slowly precipitated with 80% ethanol solution. The process was repeated three times, and the precipitate and supernatant were collected. The solvent in the supernatant was evaporated, and after dispersion in water, it was extracted three times with ethyl acetate and n-butanol to obtain an aqueous layer, an ethyl acetate layer, and a n-butanol layer. The n-butanol extract was selected by TLC and separated with D101 macroporous resin. The extract was eluted with water, 50% ethanol / water, and 90% ethanol / water in sequence to obtain three fractions with different polarities. The water-eluted fraction was examined by silica gel TLC and combined with the precipitate collected by the alcohol precipitation. After evaporation of the solvent, the precipitate was redissolved with distilled water to obtain a polysaccharide solution for later use. Protein removal was performed using the Sevage method. Chloroform and n-butanol were mixed at a ratio of 5:1, and then the polysaccharide solution was mixed with Sevage reagent at a ratio of 4:1. The mixture was shaken thoroughly for 20 min, allowed to stand, and centrifuged at 5000 rpm for 20 min. The supernatant polysaccharide aqueous solution was collected, and the process was repeated 5 times. The polysaccharide aqueous solutions were combined, the solvent was evaporated, and the mixture was dried to obtain Polygonatum polysaccharide.

[0043] (3) Gas chromatography was used to analyze the polysaccharide composition and determine the content of the obtained Polygonatum polysaccharides. After hydrolysis of the sample, acetylation was performed using acetic anhydride-pyridine (1:1). After sample processing, the contents of each monosaccharide were obtained by gas chromatography. The results are shown in Table 3.

[0044] Table 3. Monosaccharide composition analysis and content determination of Polygonatum polysaccharides

[0045] Protective effect of Polygonatum polysaccharide obtained in Example 1 against lung injury induced by nano-polystyrene microplastics (NPs) and LPS in mice. 1. Laboratory animals C57BL / 6 mice (purchased from Liaoning Changsheng Biotechnology Co., Ltd.) 2. Experimental reagents

[0046] 3. Experimental instruments

[0047] 4. Animal Experiment Procedure (1) LPS acute lung injury model established by non-invasive tracheal instillation in mice C57BL / 6 mice were housed in an SPF-grade environment and randomly assigned to four groups: a control group, an LPS group, a dexamethasone (Dex) group, and a polysaccharide (PP) group (n=8). Mice were numbered by marking their tails. After numbering, the environment was disinfected with 0.4% iodine solution. The mice's condition and weight were recorded daily. The mice were then fed according to standard care to allow them to acclimatize to their new environment.

[0048] Administer the medication via gavage for 7 days prior to modeling.

[0049] Anesthesia was administered via intraperitoneal injection of 0.5 ml / 100g of 20% urethane solution. Anesthesia was confirmed approximately 5 minutes later by clamping the toes with a foot clamp. If the foot clamping reflex persisted, a supplemental dose of urethane could be given. A 22g mouse endotracheal cannula was used, inserted into the trachea as a protective sheath. A 20 μL pipette was inserted through the sheath to administer the infusion. After anesthetizing the mouse, it was placed supine on a homemade operating table. The tongue was pulled out with forceps, and the oral cavity was wiped with a cotton swab. A direct flashlight was used to locate the trachea, which opens and closes in the throat. LPS solution was drawn up, and the endotracheal cannula was gently inserted into the trachea for infusion at a dose of 5 mg / kg. A few seconds were allowed for aspiration. After the infusion was complete, if any liquid remained, 200 μL of air was injected using an empty pipette to help all the LPS solution enter the trachea and reach the lungs. After removing the intubation tube, keep the mouse upright, gently press on the chest cavity, and roll it from side to side to ensure the LPS solution is evenly distributed in the lungs. Model establishment is complete after 6 hours of infusion. The mouse is then euthanized. Samples are collected.

[0050] (2) Establishment of NPs lung injury model by non-invasive tracheal instillation in mice C57BL / 6 mice were housed in an SPF-grade environment and randomly assigned to four groups: a control group, an NPs group, a dexamethasone (Dex) group, and a polysaccharide (PP) group (n=8). Mice were numbered by marking their tails. After numbering, the mice were disinfected with 0.4% iodine solution. The mice's condition and weight were recorded daily. The mice were then fed according to standard care to allow them to acclimatize to their new environment.

[0051] The intubation procedure is the same as (1). Aspirate the NPs suspension and gently insert the endotracheal cannula into the trachea for infusion. The infusion dose is 250 mg / kg. Wait a few seconds for the liquid to be inhaled. After the infusion is complete, if there is any remaining liquid, use an air pipette to inject 200 μL of air to help the NPs suspension enter the trachea and reach the lungs. After removing the cannula, keep the mouse upright, gently press on the mouse's chest cavity, and rotate it from side to side to ensure the NPs are evenly distributed in the mouse's lungs. Infuse once every other day for a total of 15 infusions. Administer the medication by gavage for 14 days. Sacrifice the mouse. Take samples.

[0052] 5. TNF-α in serum and bronchoalveolar lavage fluid α and IL-1 β Horizontal detection (1) Mouse tumor necrosis factor α (TNF- α ELISA kit a. Tissue sample preparation: Take an appropriate amount of tissue and homogenize it on ice with pre-cooled PBS in a ratio of 1:9 (tissue:PBS = 1:9). Centrifuge at 5000 g for 5 min at 4℃ and take the supernatant as the sample to be tested. b. Prepare the washing buffer, diluent, biotinylated antibody working solution, and HRP working solution according to the kit instructions; c. Prepare standards: Dilute the 20 ng / ml standard with 1× diluent to 1000 pg / ml, 500 pg / ml, 250 pg / ml, 125 pg / ml, 62.5 pg / ml, 31.25 pg / ml and 15.625 pg / ml, and add them to a 96-well plate for testing; d. Sample incubation: Add 100 μL of the prepared sample to each well of a 96-well plate and incubate at 37°C in the dark for 1.5 h. After incubation, add 1× wash buffer, gently shake for 30 s, and repeat 3 times. e. Antibody incubation: Add biotinylated antibody working solution and incubate at 37°C in the dark for 1 hour. After incubation, wash 4 times; f. Enzyme-labeled incubation: Add 1×SA-HRP working solution, incubate at 37℃ in the dark for 30 min, and wash 4 times after incubation. g. Substrate color development: Add color development solutions A and B sequentially, and react at 37°C in the dark for 15 min; h. Termination of reaction: After the color development is complete, add the stop solution and measure the absorbance at 450 nm using an ELISA reader.

[0053] i. A standard curve was constructed based on the absorbance of the diluted standard solution at 450 nm, which is y = 49.371x. 2 + 475.74x -20.534, R² = 0.9988, the calculated TNF-α α The concentration.

[0054] (2) Mouse interleukin-1 β (IL-1 β ELISA kit a. Tissue sample preparation: with TNF-α α The kit prepares samples simultaneously; b. Prepare the washing buffer, diluent, biotinylated antibody working solution, and HRP working solution according to the kit instructions; c. Prepare standards: Dilute the 30 ng / ml standard with 1× diluent to obtain concentrations of 1500 pg / ml, 750 pg / ml, 375 pg / ml, 187.5 pg / ml, 93.75 pg / ml, 46.88 pg / ml and 23.44 pg / ml, and add them to a 96-well plate for testing; e. The remaining steps are related to TNF-α α The reagent kits are the same.

[0055] f. A standard curve was constructed based on the absorbance of the diluted standard solution at 450 nm, as shown in the figure: y = 244.28 x 2 -313.75x +181.92, R² = 0.9961, IL-1 is calculated. β The concentration.

[0056] 6. Detection of MDA and SOD levels in lung tissue (1) MDA reagent kit a. Tissue sample preparation: Take 50 mg of tissue sample and homogenize it on ice at a ratio of 10% of tissue weight to homogenate. Centrifuge at 12000 g for 10 min at 4℃, and take the supernatant as the sample to be tested. Quantify the BCA protein concentration to facilitate subsequent calculation of MDA content per unit weight of tissue protein. b. For the mixing ratio of the MDA detection working solution and standard solution, please refer to the instruction manual for details; c. Set up the reaction system according to the kit instructions: e. After mixing thoroughly, heat in a metal bath for 15 minutes, and press the centrifuge tube cap tightly with a heavy object to prevent the liquid from boiling over and splashing. f. After heating, cool on ice, then centrifuge at 1000 g / min for 10 min. Pipette 200 μL of supernatant from each sample into a 96-well plate and measure at 532 nm. g. Dilute the standard to 1, 2, 5, 10, 20, and 50 μM with distilled water, add 0.2 ml of MDA detection working solution, and measure the absorbance at 530 and 450 nm to prepare a standard curve, such as y = 99.439x - 3.2508, R² = 0.9945. Calculate the molar concentration of MDA.

[0057] (2) SOD kit (WST-8 method) a. Tissue sample preparation: Take 50 mg of tissue sample, add SOD sample preparation solution at a ratio of 10 mg tissue to 100 μL, and homogenize on ice. Centrifuge at 12000 g for 5 min at 4℃, and take the supernatant as the sample to be tested; b. Prepare the WST-8 / enzyme working solution and reaction initiation working solution according to the kit instructions; c. Set up the reaction system according to the kit instructions, add the reagents in the required order, vortex to homogenize, and incubate at 37°C for 30 min. Measure at 450 and 600 nm, and subtract the absorbance of the 600 nm reference wavelength from the absorbance at 450 nm for the actual absorbance value.

[0058] d. Calculation of total SOD activity in the sample: e. Inhibition rate % = [(A (control 1) - A (control 2)) - (A (sample) - A (control3))] / (A (control 1) - A (control 2)) × 100% f. When the inhibition percentage is 50%, the SOD enzyme activity in the reaction system is defined as one enzyme activity unit.

[0059] g. SOD enzyme activity = inhibition rate / (1 - inhibition rate) × 100% h. Finally, based on the sample protein concentration and dilution factor, convert the SOD activity units to U / mg protein.

[0060] 7. Experimental Results (1) The lung-protective activity of Polygonatum polysaccharide was evaluated using an LPS-induced mouse acute lung injury model. The results are shown in […]. Figure 1 .

[0061] Compared with the control group (Con), the LPS (5 mg / kg) model group mice had a higher concentration of the pro-inflammatory factor TNF-α in their bronchoalveolar lavage fluid (BALF). α IL-1 β The levels of LPS were significantly increased (P<0.0001; P<0.0001), the content of malondialdehyde (MDA) in lung tissue was significantly increased (P<0.0001), and the activity of superoxide dismutase (SOD) was significantly decreased (P<0.001), indicating that LPS successfully induced lung inflammation and oxidative stress damage in mice.

[0062] Compared with the LPS model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1 β The levels of polysaccharides in lung tissue decreased significantly (P<0.0001; P<0.05), MDA content decreased significantly (P<0.0001), and SOD activity increased significantly (P<0.001). These results suggest that Polygonatum polysaccharides can significantly inhibit LPS-induced lung inflammation and oxidative stress in mice, thus exerting a lung-protective effect.

[0063] (2) The activity of Polygonatum polysaccharide in treating lung injury was evaluated using an NPs-induced mouse model of acute lung injury. The results are shown in […]. Figure 2 .

[0064] Compared with the control group (Con), the NPs (250 mg / kg) model group mice had TNF-α in their BALF. α IL-1 β The levels were significantly increased (P<0.0001); the MDA content in lung tissue was significantly increased (P<0.0001); and the SOD activity was significantly decreased (P<0.0001), indicating that NPs successfully induced lung inflammation and oxidative stress damage in mice.

[0065] Compared with the NPs model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1 β The levels of polysaccharides significantly decreased (P<0.05; P<0.05), the MDA content in lung tissue significantly decreased (P<0.01), and the SOD activity significantly increased (P<0.05). These results suggest that, at the administered concentration, Polygonatum polysaccharides can significantly reverse NPs-induced lung inflammation and oxidative stress in mice, thus exerting a therapeutic effect on lung injury.

[0066] The protective effect of Polygonatum polysaccharide obtained in Example 2 against lung injury in mice induced by nano-polystyrene microplastics and LPS. 1. The experimental animals, reagents, instruments and methods are the same as those described in Example 1 regarding the protective effect of Polygonatum polysaccharide on lung injury in mice induced by nano-polystyrene microplastics (NPs) and LPS.

[0067] 2. Experimental Results (1) The lung-protective activity of Polygonatum polysaccharide was evaluated using an LPS-induced mouse acute lung injury model. The results are shown in […]. Figure 3 .

[0068] Compared with the control group (Con), the LPS (5 mg / kg) model group mice had a higher concentration of the pro-inflammatory factor TNF-α in their bronchoalveolar lavage fluid (BALF). α IL-1 β The levels of LPS were significantly increased (P<0.0001; P<0.0001), the content of malondialdehyde (MDA) in lung tissue was significantly increased (P<0.0001), and the activity of superoxide dismutase (SOD) was significantly decreased (P<0.001), indicating that LPS successfully induced lung inflammation and oxidative stress damage in mice.

[0069] Compared with the LPS model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1β The levels of polysaccharides in lung tissue decreased significantly (P<0.001; P<0.01), MDA content decreased significantly (P<0.01), and SOD activity increased significantly (P<0.01). These results suggest that Polygonatum polysaccharides can significantly inhibit LPS-induced lung inflammation and oxidative stress in mice, thus exerting a lung-protective effect.

[0070] (2) The activity of Polygonatum polysaccharide in treating lung injury was evaluated using an NPs-induced mouse model of acute lung injury. The results are shown in […]. Figure 4 .

[0071] Compared with the control group (Con), the NPs (250 mg / kg) model group mice had TNF-α in their BALF. α IL-1 β The levels were significantly increased (P<0.0001); the MDA content in lung tissue was significantly increased (P<0.0001); and the SOD activity was significantly decreased (P<0.0001), indicating that NPs successfully induced lung inflammation and oxidative stress damage in mice.

[0072] Compared with the NPs model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1 β The levels of polysaccharides in lung tissue decreased significantly (P<0.01; P<0.01), MDA content decreased significantly (P<0.001), and SOD activity increased significantly (P<0.01). These results suggest that, at the administered concentration, Polygonatum polysaccharides can significantly reverse NPs-induced lung inflammation and oxidative stress in mice, thus exerting a therapeutic effect on lung injury.

[0073] The protective effect of Polygonatum polysaccharide obtained in Example 3 against mouse damage induced by nano-polystyrene microplastics and LPS. 1. The experimental animals, reagents, instruments and methods are the same as those described in Example 1 regarding the protective effect of Polygonatum polysaccharide on lung injury in mice induced by nano-polystyrene microplastics (NPs) and LPS.

[0074] 2. Experimental Results (1) The lung-protective activity of Polygonatum polysaccharide was evaluated using an LPS-induced mouse acute lung injury model. The results are shown in […]. Figure 5 Compared with the control group (Con), the LPS (5 mg / kg) model group mice had higher levels of the pro-inflammatory factor TNF-α in their bronchoalveolar lavage fluid (BALF). α IL-1 βThe levels of LPS were significantly increased (P<0.0001; P<0.0001), the content of malondialdehyde (MDA) in lung tissue was significantly increased (P<0.0001), and the activity of superoxide dismutase (SOD) was significantly decreased (P<0.001), indicating that LPS successfully induced lung inflammation and oxidative stress damage in mice.

[0075] Compared with the LPS model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1 β The levels of polysaccharides in lung tissue decreased significantly (P<0.0001; P<0.001), MDA content decreased significantly (P<0.0001), and SOD activity increased significantly (P<0.01). These results suggest that Polygonatum polysaccharides can significantly inhibit LPS-induced lung inflammation and oxidative stress in mice, thus exerting a lung-protective effect.

[0076] (2) The activity of Polygonatum polysaccharide in treating lung injury was evaluated using an NPs-induced mouse model of acute lung injury. The results are shown in […]. Figure 6 .

[0077] Compared with the control group (Con), the NPs (250 mg / kg) model group mice had TNF-α in their BALF. α IL-1 β The levels were significantly increased (P<0.0001); the MDA content in lung tissue was significantly increased (P<0.0001); and the SOD activity was significantly decreased (P<0.0001), indicating that NPs successfully induced lung inflammation and oxidative stress damage in mice.

[0078] Compared with the NPs model group, intervention with Polygonatum polysaccharide (PP, 105 mg / kg) reduced TNF-α levels in mouse BALF. α IL-1 β The levels of polysaccharides in lung tissue were significantly reduced (P<0.001; P<0.001), MDA content was significantly decreased (P<0.001), and SOD activity was significantly increased (P<0.001). These results suggest that, at the administered concentration, Polygonatum polysaccharides can significantly reverse NPs-induced lung inflammation and oxidative stress in mice, thus exerting a therapeutic effect on lung injury.

Claims

1. The application of Polygonatum polysaccharide in the preparation of products for the prevention and / or treatment of lung injury, characterized in that: The product in question is a medicine or health supplement.

2. The application according to claim 1, characterized in that: The lung injury is one or more of the following: acute lung injury, meconium aspiration syndrome, pneumoconiosis, radiation-induced lung injury, or mechanical ventilation-related lung injury.

3. The application according to claim 1, characterized in that: The monosaccharide composition of the Polygonatum polysaccharide includes fructose, rhamnose, galactose, glucose, mannose, and xylose; the contents of fructose, rhamnose, galactose, glucose, mannose, and xylose are 0.24~0.50 mg / g, 0.79~1.49 mg / g, 0.75~1.52 mg / g, 0.23~0.72 mg / g, 3.79~5.28 mg / g, and 0.10~0.78 mg / g, respectively.

4. The application according to claim 1, characterized in that: The polysaccharide from Polygonatum was prepared by first extracting Polygonatum using conventional extraction methods, followed by separation using extraction and column chromatography.

5. The application according to claim 4, characterized in that: The conventional extraction method is one or more of the following: ultrasonic extraction, decoction, heating and reflux extraction, percolation extraction, or microwave extraction. The extraction solvent is water; the number of extractions is 3-5 times; the extraction time for each extraction is 60-120 min; and the volume-to-mass ratio of the extraction solvent to Polygonatum is 2-5 L : 1 kg.

6. The application according to claim 4, characterized in that: The column chromatography method is one or both of D101 macroporous resin adsorption method and AB-8 macroporous resin adsorption method, wherein the elution solvent is one or more of water or ethanol solution with a volume fraction of 30%-90%.

7. The application according to claim 4, characterized in that: The Polygonatum polysaccharide was prepared by the following method: (1) Weigh out the Polygonatum according to the selected weight proportions, add water, soak, decoct, filter, and obtain filtrate and dregs; (2) Precipitate the filtrate obtained in step (1) with alcohol, and collect the supernatant and precipitate; (3) Disperse the supernatant obtained in step (2) with water, extract with ethyl acetate and n-butanol, and collect the n-butanol layer sample; (4) Pass the n-butanol layer sample obtained in step (3) through D101 macroporous resin and elute it sequentially with pure water, 50% ethanol aqueous solution and 90% ethanol aqueous solution, and collect the water elution portion. (5) Combine the water-washed portion obtained in step (4) and the precipitate obtained in step (2), remove the protein, and dry to obtain Polygonatum polysaccharide.

8. The application according to claim 7, characterized in that: In step (1), the soaking time is 60-120 min; the decocting time is 60-120 min; and the number of times water is added for decocting is 3-5 times.

9. The application according to claim 7, characterized in that: In step (2), the alcohol precipitation is performed 3-5 times; in step (3), ethyl acetate and n-butanol are used for extraction 3-5 times each; in step (5), the protein is removed 1-5 times, and each time it is left to stand overnight at 4°C.

10. The application according to claim 7, characterized in that: In step (5), the deproteinization method is one or more of the Sevage method, TCA method, or enzymatic hydrolysis method.

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