Application of abrus herb extract in preparation of medicine for preventing and treating lung injury caused by cigarette smoke

Abstract

The invention discloses an application of an abrus cantoniensis hance extract in preparation of a medicine for preventing and treating lung injury caused by cigarette smog, the abrus cantoniensis hance extract comprises total saponins, polysaccharides, total flavonoids and total alkaloids, and the survival rate of lung cells injured by cigarette smog induction can be increased; the composition can improve the levels of SOD, GSH, GSH-Px and CAT in normal or injured human bronchial epithelial cells (HBE) and reduce the level of IL-6 in the normal HBE, has a lung injury protection effect, and has important significance in prevention and treatment of respiratory system chronic diseases caused by smoke.

Description

Technical Field

[0001] The invention relates to the fields of medical technology and food, and in particular to the use of a smilax china or smilax china extract in the preparation of a herb for preventing, improving and treating lung damage and pulmonary fibrosis caused by cigarette smoke. Background Art

[0002] Chronic obstructive pulmonary disease (COPD) is a disease characterized by long-term irreversible airway restriction and persistent respiratory symptoms. Smoking is the main risk factor for COPD. Its course of development is closely related to mechanisms such as inflammatory response, oxidative stress, proteases / antiproteases, apoptosis and airway remodeling. It has now become the third leading cause of death in the world. There is still no specific treatment for COPD. It mainly focuses on smoking cessation, inhaled bronchodilators, oxygen control and mechanical ventilation when necessary. Conventional Western medicine has significant toxic and side effects and cannot reverse the progressive decline in lung function. Many research results have shown that Chinese medicine has a good effect in improving the lung function of COPD patients, alleviating clinical symptoms, and improving the quality of life of patients, and has better safety.

[0003] As a traditional Chinese medicine for both medicine and food, the whole plant is used as medicine, which has the effects of removing dampness and jaundice, clearing away heat and detoxifying. It is often used to treat damp-heat jaundice, rib discomfort, stomach pain, mastitis and other diseases. It is the main ingredient of the Chinese patent medicine chicken bone grass capsule and one of the main ingredients of Guangdong herbal tea. The "Lingnan Herbal Medicine Records" records: "Chicken bone grass can soothe the liver, harmonize the spleen, and heal fractures"; the "National Compilation of Chinese Herbal Medicines" shows that "Chicken bone grass can treat acute and chronic hepatitis, ascites due to liver cirrhosis, stomachache, rheumatic bone pain, snake bites, and can be used as a refreshing drink in summer." Modern research on chicken bone grass shows that chicken bone grass has antioxidant, anti-cell proliferation, anti-cancer and immunomodulatory effects.

[0004] Our previous studies have shown that radix serrata contains soya saponin Bb, abrin, schaftoside, abrinine, erythrine and other ingredients. Studies have shown that soya saponins (A1, A2 and I) in radix serrata can inhibit the release of prostaglandin E2 (PGE2), an inflammatory marker induced by LPS, which is similar to the NF-κB inhibitor (BAY117082). It can also inhibit the expression of cyclooxygenase 2 (COX-2) in a dose-dependent manner by inhibiting the activation of NF-κB. Soya saponins can inhibit the activation of PI3K / Akt and the production of reactive oxygen species (ROS) induced by LPS, increase SOD activity and the GSH / GSSG ratio, and reduce the occurrence of inflammation.

[0005] The present invention prepares an extract from Aconitum carmichaelii or Aconitum pubescens, and verifies the protective effect and related mechanism of Aconitum carmichaelii extract on CSE-induced cell damage through a cigarette smoke (CSE)-induced cell damage model, suggesting that Aconitum carmichaelii extract has important potential in preventing and treating lung injury and pulmonary fibrosis, and has great social and economic value in view of the Healthy China strategy and the major national needs for the prevention and treatment of chronic diseases.

[0006] References

[0007] 1.Zhang Chengzhong,Bu Qitao,Li Chuyan,Lu Pengfei,Liu Chang,HuangBaokang.Simultaneous determination of abrine,hypaphorine,schaftoside andsoyasaponin Bb in rat plasma by UPLC-MS / MS and its application to apharmacokinetic study after oral administration of Abrus cantoniensis Hanceextract.Biomed Chromatogr.2023;37(10):e5696.

[0008] 2.Wang M, Chen

[0009] 3. Jiang Yuhang, Mei Xiaofeng, Jia Lidan, Tian Yange, Zhao Peng. Mechanism of airway epithelial barrier damage induced by cigarette smoke in mice with chronic obstructive pulmonary disease. Chin J Pathophysiol. 2022, 7: 1297-1303.

[0010] 4. Xiao Xiao, Yao Xiangcao, Yu Yaru, Xu Chongyuan, Huang Baokang. Resource survey and pharmacognosy identification of Herba Corydalis. Journal of Pharmaceutical Practice. 2019, 4: 318-321

[0011] 5. Xiao Xiao, Xu Zhongyuan, Yang Dejun, Huang Baokang. Comparative analysis of volatile oil and fatty acid composition of Herba Corydalis and Herba Corydalis, Journal of Pharmaceutical Practice, 2017, 35(1), 39-42. Summary of the invention

[0012] The purpose of the present invention is to provide a Chinese medicine extract for preventing and treating lung injury and pulmonary fibrosis in view of the shortcomings of current research technology for preventing and treating chronic lung injury. Another purpose is to provide the use of a herbaceous herb or a herbaceous herb extract in preventing and treating lung injury and pulmonary fibrosis.

[0013] To achieve the above purpose, the technical scheme adopted by the present invention is: using scutellaria baicalensis and scutellaria baicalensis as raw materials, extracting with ethanol solution and water to obtain the extract, which includes total saponin, total alkaloid, total flavonoid and total polysaccharide.

[0014] In order to achieve the above second purpose, the extracts of the Chinese medicinal and edible herbs, scutellaria baicalensis and scutellaria baicalensis, are used in the preparation of medicines or foods for preventing and treating lung injury and pulmonary fibrosis. The scutellaria baicalensis extract disclosed in the present invention can be used alone or in combination, in the form of medicine, or in the form of food or health food, including tablets, granules, capsules, pills, oral liquids, injections, etc., and are not limited to the above-mentioned pharmaceutical dosage forms, and can be herbal teas, brewed beverages, solid foods, etc., and are not limited to the above-mentioned forms.

[0015] When the Herba Corydalis extract disclosed by the invention is used in medicine or food, necessary excipients, fillers, diluents and other auxiliary materials that can be added to medicines or foods can be added. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 :Effects of Herba Corydalis extract on cell activity. C is the blank group, M is the model group, DEX is the dexamethasone control group, W1, W2, and W3 are three different concentrations of Herba Corydalis water extract groups, G1 and G2 are different concentrations of total saponin groups, and D1 and D are different concentrations of polysaccharide groups.

[0017] Figure 2 : Effects of Herba Corydalis extract on the levels of SOD, GSH and GSH-Px in normal HBE cells. C is the blank group, M is the model group, NAc is the n-acetylcysteine ​​positive control group, W is the Herba Corydalis water extract group, ZG is the total saponin group, and DT is the polysaccharide group.

[0018] Figure 3 :Effects of Herba Corydalis extract on the levels of SOD, GSH, GSH-Px and CAT in HBE cells damaged by CSE. C is the blank group, M is the model group, NAc is the n-acetylcysteine ​​positive control group, W is the Herba Corydalis water extract group, ZG is the total saponin group, and DT is the polysaccharide group.

[0019] Figure 4 :Effect of Aconitum carmichaelii on cellular IL-6 levels. DETAILED DESCRIPTION

[0020] The following examples are used to illustrate specific implementation methods, which further illustrate and supplement the content of the present invention. However, the scope of the above subject matter of the present invention is not limited to the following examples. All technologies and applications realized based on the above content of the present invention belong to the scope of the present invention.

[0021] Example 1 - Preparation of Polysaccharide from Glechoma longituba

[0022] The whole dried herb of Glechoma longituba was crushed, pretreated with 3% cellulase for 2 hours, and then extracted with hot water. 4 ) Orthogonal test was used to investigate temperature, extraction time, solid-liquid ratio, extraction times, and phenol-sulfuric acid method to determine the polysaccharide yield. The optimal extraction process was as follows: extraction temperature 100℃, extraction time 1.5h, solid-liquid ratio 1:25, extraction 3 times, and the extract was centrifuged (4000rpm, 15min) to remove insoluble impurities. Sevage method (chloroform: n-butanol = 4:1) was used to remove proteins. Activated carbon adsorption method (1% activated carbon, 60℃, 30min). 4 times the volume of anhydrous ethanol was added, and the mixture was allowed to stand at 4℃ for 12h, and the precipitate was collected by centrifugation. The polysaccharide of Glechoma longituba was obtained by freeze drying, and the polysaccharide extraction rate was 8.51%.

[0023] Example 2 - Preparation of Polysaccharide Capsules of Glechoma longituba for Lung Injury and Pulmonary Fibrosis

[0024] 50g of radix scutellariae polysaccharide, 30g of microcrystalline cellulose (MCC), 15g of lactose, 3g / tablet of sodium carboxymethyl starch (CMS-Na), 1g of magnesium stearate, and 1g of polyvinyl pyrrolidone (PVP K30). Pass radix scutellariae polysaccharide, MCC, lactose, and CMS-Na through an 80-mesh sieve and mix evenly. Wet granulate with an ethanol solution of PVP K30 (5% w / v) and pass through a 20-mesh sieve. Dry at 60°C until the moisture is ≤5%. Pass through a 20-mesh sieve to granulate. Add magnesium stearate and mix evenly. Use a single-punch tablet press to press tablets to prepare 1000 tablets, with a tablet weight of 100mg. Each tablet contains 50mg of radix scutellariae polysaccharide, and other requirements comply with the relevant provisions of the 2020 edition of the Pharmacopoeia of the People's Republic of China on capsules.

[0025] Example 3 - Preparation of scutellaria baicalensis saponin capsules for lung injury and pulmonary fibrosis

[0026] Take 5g of total saponins of Herba Corydalis, 20g of corn starch, and an appropriate amount of cyclodextrin, mix well, and put them into capsules to prepare 100 capsules, each containing 50mg of total saponins. Other requirements shall comply with the relevant provisions of the 2020 edition of the Pharmacopoeia of the People's Republic of China on capsules.

[0027] Example 4 - Preparation of scutellaria baicalensis saponin-polysaccharide capsules for lung injury and pulmonary fibrosis

[0028] 2.5 g of total saponins and polysaccharides of Herba Polygoni Multiflori were mixed evenly, and 1000 capsules were prepared according to the method of Example 3, each containing 25 mg of total saponins and 25 mg of polysaccharides. Other requirements were in accordance with the relevant provisions of the 2020 edition of the Pharmacopoeia of the People's Republic of China on capsules.

[0029] Example 5 - Preparation of Herba Glehniae Tea

[0030] Take 1kg of Chinese herbal medicine Herba Glechomae (or Herba Glechomae), cut into sections, add water and boil twice, each time with 8L of pure water, combine the decoction, concentrate into extract, take 100g of dextrin, 100g of soluble starch, appropriate amount of powdered sugar, mix well, make tea blocks, and package them in batches, with a filling volume of 5.0g / bag. Other requirements should meet the requirements for tea preparations.

[0031] Example 6 - Preparation of Herba Glehniae Oral Liquid / Mixture

[0032] Take the extract described in any one of Examples 1-8, add 2 times the amount of alcohol, stir and precipitate overnight. Take the supernatant, concentrate it to a thick extract, add appropriate pharmaceutical excipients, and prepare an oral liquid or mixture.

[0033] The above is only a preferred embodiment of the present invention. It should be pointed out that ordinary technicians in this technical field can make several improvements and supplements without departing from the method of the present invention. These improvements and supplements should also be regarded as the scope of protection of the present invention.

[0034] Example 7-Effect of Herba Corydalis Extract on Lung Injury Induced by Cigarette Smoke:

[0035] 1. Preparation of Herba Corydalis Extract

[0036] (1) Aqueous extract of scutellaria baicalensis

[0037] Take the whole dried plant of Acanthopanax gracilis, chop it into 2 cm long sections, soak it in 10 times the volume of water for 1 hour, boil it for 2 hours, separate the filtrate, boil it in 5 times the volume of water for 1 hour, combine the two filtrates, concentrate it to the equivalent of the original drug concentration of 3g / ml, package it, and obtain Acanthopanax gracilis water extract, which is stored at 4°C for later use.

[0038] (2) Total alkaloids from scutellaria baicalensis

[0039] Take the powder of Acanthopanax leaf (passed through No. 4 sieve), add 70% ethanol in a solid-liquid ratio (g:ml) of 1:9, and perform ultrasonic extraction at 70°C for 30 minutes, twice in total. After combining the filtrate, add petroleum ether to remove the pigment, extract with chloroform twice, take the chloroform layer and concentrate it to a brown extract. The total alkaloids of Acanthopanax were obtained and stored at -20°C for later use.

[0040] (3) Total flavonoids from Aconitum

[0041] Take the dried scutellaria powder (passed through a No. 4 sieve), defat it with petroleum ether for 1.5 hours, and then dry it in an electric constant temperature blast drying oven at 80°C. Take the defatted scutellaria powder, add 10 times the amount of 80% ethanol, and ultrasonically extract it at 70°C for 2.5 hours, twice in total. After combining the filtrates, concentrate them, and dry them in an electric constant temperature blast drying oven to obtain the total flavonoids of scutellaria, which are then stored at -20°C for future use

[10] .

[0042] (4) Total saponins of Acanthopanax granatum

[0043] The powder of Acanthopanax salsa was extracted with ethanol, extracted with n-butanol, separated and purified with macroporous adsorption resin, and then freeze-dried and stored at -20°C for future use.

[0044] (5) Polysaccharide from Acanthopanax gracilis

[0045] Take the powder of scutellaria baicalensis (pass through a No. 4 sieve), add 10 times the amount of water, place in a 65°C constant temperature water bath for 3 hours, repeat 3 times, combine the 3 filtrates, and concentrate. Add 3 times the volume of 95% ethanol, stand in the backlight for 24 hours, filter, take out the precipitate, wash it twice with anhydrous ethanol and petroleum ether in turn, and obtain scutellaria baicalensis polysaccharide, which is stored at -20°C for later use

[0046] 2. Preparation of cigarette smoke extract (CSE) and establishment of CSE-induced cell injury model

[0047] (1) Preparation of CSE

[0048] According to the aforementioned reference 2 Method: To prepare CSE, add 50 ml of culture medium as the absorption liquid into a 75 ml large tube, connect a cigarette at one end of the tube and a 50 ml syringe at the other end. After the cigarette is lit, control the number of puffs per cigarette to 15 times (a total of 20 cigarettes), draw smoke at a constant speed until the cigarette is burned out, gently shake the glass bottle to fully dissolve the smoke, transfer the collected liquid into a centrifuge tube, and filter it with a 0.22 μm pore filter to obtain 100% CSE stock solution. The stock solution is divided and frozen in a -80°C refrigerator, and diluted with the corresponding volume of culture medium before use. The prepared CSE of different concentrations is used within 30 minutes.

[0049] (2) Establishment of CSE-induced cell injury model

[0050] The HBE cells used in this experiment were inoculated in 96-well plates and treated with different concentrations of CSE. Based on preliminary experiments in a wide concentration and long time range, the concentration gradient and action time were clarified. The cell viability after the action of CSE was detected by the CCK-8 method, and the CSE concentration with a cell survival rate of about 50% and the appropriate treatment time were selected as the conditions for establishing the cell injury model.

[0051] 3. Study on the protective activity of Herba Corydalis extract against CSE cell damage

[0052] The cells were seeded in 96-well plates, and 100nM / ml dexamethasone was used as a positive control. The cells were pretreated with an appropriate concentration of extract for 24 hours, and then the HBE cells were treated with 1.52% CSE for 24 hours to establish a CSE cell injury model. The cell viability was detected by CCK-8 method to evaluate the effect of the Herba Corydalis extract on cell viability. The cell survival rate of the model group treated with CSE was significantly different from that of the blank control group (p < 0.0001); the positive control group was significantly different from the model group (P < 0.0001). Herba Corydalis water extract, Herba Corydalis total saponins and Herba Corydalis polysaccharides all have a protective effect on CSE-induced HBE cell damage (p < 0.01) ( Figure 1 ).

[0053] 4. Effect of Herba Corydalis Extract on Oxidative Stress Response of CSE-Injured Cells

[0054] The water extract of Herba Corydalis can effectively increase the levels of SOD, GSH, GSH-Px and CAT, and has the best effect on GSH, which is significantly different from the Control group (P < 0.001); total saponins can increase the levels of GSH, GSH-Px and CAT in normal HBE cells; polysaccharides can also increase the levels of SOD, GSH-Px and CAT ( Figure 2 ).

[0055] After pretreatment of HBE cells with 1.52% CSE for 24 hours, the CSE-HBE cell injury model was established, and the effect of 1.52% CSE on the oxidative stress response of CSE-injured cells was detected. The results showed that the water extract and polysaccharide had the best effect on improving the SOD level in CSE-injured HBE cells (P < 0.0001, vs model group). 1.52% CSE was used to treat HBE cells with 1.52% CSE for 24 hours. The polysaccharide and saponin of 1.52% CSE increased the levels of GSH and GSH-Px in HBE-injured cells (P < 0.01, vs model group).

[0056] (P<0.05)( Figure 3 ).

[0057] 5. Effect of Herba Corydalis Extract on the Levels of Inflammatory Factors in CSE-Injured Cells

[0058] The cells were inoculated in 96-well plates, and blank group, model group, dexamethasone positive control group, and scutellaria baicalensis extract group were set up. After adding the corresponding drug pretreatment, except for the blank group, the cells were treated under appropriate modeling conditions, and the supernatant culture fluid of each group was collected. The level of IL-6 in the supernatant was detected by ELISA to evaluate the effect of scutellaria baicalensis extract on the level of inflammatory factors in CSE-damaged cells. The results showed that scutellaria baicalensis water extract, total saponins and polysaccharide cells could significantly reduce the level of IL-6 in HBE cells (P<0.0001)( Figure 4 ).

Claims

1. Application of Herba Corydalis extract in the preparation of drugs for preventing and treating lung damage caused by cigarette smoke.

2. The use according to claim 1, characterized in that: The extract of scutellaria baicalensis is water extract of scutellaria baicalensis, total flavonoids of scutellaria baicalensis, total saponins of scutellaria baicalensis or polysaccharide of scutellaria baicalensis.

3. The use according to claim 1 or 2, characterized in that: The extract of Acanthopanax ovale can improve the survival rate of damaged lung cells induced by cigarette smoke, increase the levels of intracellular SOD, GSH, GSH-Px, and CAT, and reduce the content of IL-6 in normal bronchial cells of normal people.

4. The use according to claim 1 or 2, characterized in that: The extract of Aconitum carmichaelii is made into tablets, capsules, granules, oral liquids, drop pills, pills, ointments, mixtures, gels, chewable tablets, lozenges, chewing gums or teas.

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