Traditional Chinese medicine fermentation composition helpful for improving sleep and having effects of treating lung cancer and pulmonary nodule and application of traditional Chinese medicine fermentation composition

Abstract

The invention discloses a traditional Chinese medicine fermentation composition helpful for improving sleep and having effects of treating lung cancer and pulmonary nodule and application thereof, the traditional Chinese medicine fermentation composition comprises astragalus membranaceus, codonopsis pilosula, radix ophiopogonis, gastrodia elata, rehmannia glutinosa, turmeric, oyster, sweet almond, bunge cherry seed, exocarpium citri grandis, sea-buckthorn, angelica sinensis, peach kernel, lotus leaf, jasmine, chrysanthemum and lucid ganoderma, the exocarpium and the sea-buckthorn are fermented by bifidobacterium; the traditional Chinese medicine fermented composition disclosed by the invention is prepared by a grouped fermentation method, can effectively improve the sleep disorder symptom of a mouse, and is proved to be capable of inhibiting the proliferation of lung cancer cells and inhibiting the growth of tumor cells by carrying out efficacy evaluation research on pulmonary nodules and lung cancer; the pharmacological basis and reference are provided for the clinical application of the traditional Chinese medicine composition in the blood stasis ointment.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine and biological fermentation preparations, and particularly relates to a traditional Chinese medicine fermentation composition prepared by a fermentation method, which has the effects of improving sleep, treating lung cancer and lung nodules. The traditional Chinese medicine fermentation composition combines traditional Chinese medicine ingredients with modern biotechnology, and through the fermentation of Lactobacillus lactis and Bifidobacterium, improves the drug efficacy and enhances the bioavailability. It can not only effectively relieve sleep disorders, but also inhibit the proliferation of lung cancer cells, and has important clinical application prospects. Background Art

[0002] In traditional Chinese medicine, blood stasis constitution refers to the unsmooth flow of qi and blood, resulting in blood stasis in the body, often manifested as symptoms such as pain, mass, tumor, etc.

[0003] The active ingredients in many traditional blood stasis quality plasters are directly added to the plaster without any treatment, which may be restricted by biological barriers such as gastrointestinal absorption and metabolism, resulting in low bioavailability of the drug; and the existing blood stasis quality plasters mainly focus on promoting blood circulation to remove blood stasis and relieving pain. Although they have certain therapeutic effects, they are often insufficient for the comprehensive conditioning of complex symptoms (such as sleep disorders, low immune function, etc.). There is also a lack of clear evidence for the efficacy of some products in treating tumor-related diseases such as lung cancer, and it is difficult to effectively meet the multi-dimensional needs in cancer treatment. On the other hand, many traditional blood stasis quality plasters may contain more crude drug ingredients or high-concentration herbal extracts. Although they can be absorbed locally or orally, due to the limitations of the plaster form, their absorption speed is slow and the action is relatively slow. For symptoms that need to be relieved urgently, such as insomnia, tumor-related symptoms, etc., they may not take effect in a short time. Finally, most of the existing blood stasis quality plasters on the market are formulated based on traditional Chinese medicine theory, lacking systematic pharmacological and clinical verification. Although some products are effective, it is often difficult to provide sufficient scientific evidence to prove their efficacy in clinical practice, and it is difficult to obtain the recognition of the modern medical community. Moreover, the efficacy varies greatly among individuals and lacks standardization.

[0004] As one of the malignant tumors with the highest incidence and mortality worldwide, lung cancer has no obvious early symptoms and is often accompanied by discomfort such as insomnia and fatigue, affecting the quality of life of patients. Therefore, finding drugs that can both condition the constitution and have anti-tumor effects has become a research hotspot in medical research.

[0005] Compared with the existing blood stasis constitution ointments on the market, the traditional Chinese medicine fermentation composition of the present invention, as the active ingredient of the blood stasis constitution ointment, can improve the bioavailability of the drug efficacy, promote the transformation of active ingredients and enhance their pharmacological effects through the introduction of modern fermentation technology. It can also reduce the toxic and side effects of some medicinal materials, expand the treatment field, improve the diversity of curative effects, and has more definite pharmacological basis support. Its comprehensive effects can not only improve the blood stasis constitution and relieve sleep disorders, but also play a significant role in anti-tumor, immune regulation and other aspects, solving the deficiencies of existing products such as lack of multiple curative effects, difficult to absorb, low safety and lack of pharmacological basis, and having greater clinical application prospects. Summary of the Invention

[0006] The purpose of the present invention is to overcome the deficiencies of the prior art and provide a traditional Chinese medicine fermentation composition and its application that are helpful for improving sleep and have the effects of treating lung cancer and lung nodules, so as to solve the problems raised in the above technical background.

[0007] To achieve the above purpose, the present invention is realized through the following technical solutions:

[0008] In the first aspect, the present invention provides a traditional Chinese medicine fermentation composition that is helpful for improving sleep and has the effects of treating lung cancer and lung nodules, which contains the following raw materials by weight: 5-8 parts of Astragalus membranaceus, 4-6 parts of Codonopsis pilosula, 6-8 parts of Ophiopogon japonicus, 6-8 parts of Gastrodia elata, 5-8 parts of Rehmannia glutinosa, 5-7 parts of Curcuma longa, 5-7 parts of Oyster, 6-8 parts of Sweet Almond, 6-9 parts of Prunus davidiana, 8-12 parts of Exocarpium Citri Grandis, 4-7 parts of Hippohgae rhamnoides, 6-9 parts of Angelica sinensis, 5-8 parts of Peach Kernel, 3-6 parts of Lotus Leaf, 4-7 parts of Jasmine, 3-5 parts of Chrysanthemum, 2-5 parts of Ganoderma lucidum; among them, Astragalus membranaceus and Codonopsis pilosula are fermented by Lactobacillus, and Exocarpium Citri Grandis and Hippohgae rhamnoides are fermented by Bifidobacterium.

[0009] In the above technical solution, it contains the following raw materials by weight: 6 parts of Astragalus membranaceus, 5 parts of Codonopsis pilosula, 7 parts of Ophiopogon japonicus, 7 parts of Gastrodia elata, 7 parts of Rehmannia glutinosa, 6 parts of Curcuma longa, 6 parts of Oyster, 7 parts of Sweet Almond, 6 parts of Prunus davidiana, 10 parts of Exocarpium Citri Grandis, 7 parts of Hippohgae rhamnoides, 6 parts of Angelica sinensis, 5 parts of Peach Kernel, 4 parts of Lotus Leaf, 5 parts of Jasmine, 4 parts of Chrysanthemum, 3 parts of Ganoderma lucidum.

[0010] In the above technical solution, after mixing the Chinese medicinal materials except Astragalus membranaceus, Codonopsis pilosula, Exocarpium Citri Grandis and Hippohgae rhamnoides in proportion, add 8-10 times the amount of distilled water, and carry out extraction 3 times. After extraction, combine the filtrates and concentrate to 1 / 5 of the filtrate to obtain the Chinese medicinal material extract.

[0011] In the above technical solution, the dosage of the Lactobacillus bacterial liquid is 0.6% of the total mass of Astragalus membranaceus and Codonopsis pilosula, and the dosage of the Bifidobacterium bacterial liquid is 0.6% of the total mass of Exocarpium Citri Grandis and Hippohgae rhamnoides. The concentration of the strains contained in the Lactobacillus bacterial liquid is 10 8 ~10 9CFU / ml, the concentration of the strains contained in the Bifidobacterium bacterium solution is 10 8 ~10 9 CFU / ml.

[0012] In the above technical solution, Lactobacillus acidophilus CICC 6243 is selected as the Lactobacillus, and Bifidobacterium longum CICC 24632 is selected as the Bifidobacterium.

[0013] In a second aspect, the present invention also provides a preparation method of a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules, including the following steps:

[0014] (1) Weigh 5 - 8 parts of Astragalus membranaceus, 4 - 6 parts of Codonopsis pilosula, 6 - 8 parts of Ophiopogon japonicus, 6 - 8 parts of Gastrodia elata, 5 - 8 parts of Rehmannia glutinosa, 5 - 7 parts of Curcuma longa, 5 - 7 parts of Oyster, 6 - 8 parts of Sweet Almond, 6 - 9 parts of Prunus davidiana, 8 - 12 parts of Citrus reticulata Blanco cv. Tomentosa, 4 - 7 parts of Hippohgae rhamnoides L., 6 - 9 parts of Angelica sinensis, 5 - 8 parts of Prunus persica, 3 - 6 parts of Lotus leaf, 4 - 7 parts of Jasminum sambac, 3 - 5 parts of Chrysanthemum morifolium, 2 - 5 parts of Ganoderma lucidum by weight, wash, dry and then crush them into 80 - mesh powder respectively;

[0015] (2) Mix 6 - 8 parts of Ophiopogon japonicus, 6 - 8 parts of Gastrodia elata, 5 - 8 parts of Rehmannia glutinosa, 5 - 7 parts of Curcuma longa, 5 - 7 parts of Oyster, 6 - 8 parts of Sweet Almond, 6 - 9 parts of Prunus davidiana, 6 - 9 parts of Angelica sinensis, 5 - 8 parts of Prunus persica, 3 - 6 parts of Lotus leaf, 4 - 7 parts of Jasminum sambac, 3 - 5 parts of Chrysanthemum morifolium, 2 - 5 parts of Ganoderma lucidum, add 8 - 10 times the amount of distilled water after mixing, and carry out extraction 3 times, extracting for 60h, 40min, 40min respectively. After the extraction is completed, combine the filtrates and concentrate to 1 / 5 of the filtrate to obtain the traditional Chinese medicine extract.

[0016] (3) On the basis of the traditional Chinese medicine extract obtained in step (2), Astragalus membranaceus, Codonopsis pilosula, Citrus reticulata Blanco cv. Tomentosa, and Hippohgae rhamnoides L. are treated by a group fermentation method. Specifically, prepare Lactobacillus acidophilus bacterium solution with a concentration of 1×10 8 ~10 9 CFU / ml and Bifidobacterium bacterium solution with a concentration of 1×10 8 ~1~10 9 CFU / mL. The selected strain of Lactobacillus acidophilus is Lactobacillus acidophilus CICC 6243 (purchased from China Center for Industrial Culture Collection), and the selected strain of Bifidobacterium is Bifidobacterium longum CICC 24632 (China Center for Industrial Culture Collection). The freeze - dried strains of Lactobacillus acidophilus CICC 6243 and Bifidobacterium longum CICC 24632 are dissolved in sterile water, and then the freeze - dried strain powder is allowed to revive by adding MRS liquid medium and RCM liquid medium respectively and then continue to be cultured into the bacterium solution for fermentation, and adjusted to make the fermentation concentration 1×10 8 ~10 9 CFU / mL.

[0017] MRS liquid medium: 10.0 g of casein peptone, 10.0 g of beef extract powder, 5.0 g of yeast extract powder, 20.0 g of glucose, 1.0 g of Tween 80, 5.0 g of sodium acetate, 2.0 g of ammonium citrate, 2.0 g of K2HPO4, 0.2 g of MgSO4·7H2O, 0.05 g of MnSO4·H2O, 1000.0 mL of distilled water, pH 6.2 - 6.5; after preparation, sterilize at 121 °C for 15 minutes.

[0018] RCM liquid medium: 10.0 g of beef extract powder, 5.0 g of peptone, 3.0 g of yeast extract powder, 5.0 g of glucose, 1.0 g of soluble starch, 5.0 g of NaCl, 3.0 g of NaAc, 0.5 g of L-cysteine hydrochloride, 1000.0 mL of distilled water, pH 6.8; after preparation, sterilize at 121 °C for 15 minutes.

[0019] (4) After mixing astragalus and codonopsis pilosula, add the bacterial liquid of Lactobacillus acidophilus CICC 6243 to ferment to obtain the astragalus / codonopsis pilosula ferment, fermentation process:

[0020] S1. Mix the powders of astragalus and codonopsis pilosula obtained in step (1) in proportion.

[0021] S2. Add the prepared bacterial liquid of Lactobacillus acidophilus CICC 6243 and MRS liquid medium to the mixture of astragalus and codonopsis pilosula and stir well. The addition amount of the Lactobacillus acidophilus bacterial liquid is 0.6% of the total mass of astragalus and codonopsis pilosula, and the addition amount of MRS liquid medium is 2 times the total mass of astragalus and codonopsis pilosula.

[0022] S3. Transfer the mixture added with the Lactobacillus acidophilus bacterial liquid to a sterile fermentation container, ensure anaerobic conditions in the container to support the growth of Lactobacillus acidophilus CICC 6243.

[0023] S4. Ferment at a constant temperature of 37 °C for 48 hours. After fermentation, perform water extraction on the obtained ferment to obtain the filtrate and concentrate it to 1 / 5 of the filtrate, that is, obtain the astragalus / codonopsis pilosula ferment extraction solution. Among them, the water extraction method of the ferment is: add 8 times the amount of distilled water to the ferment, perform extraction 3 times, extract for 60 min, 40 min, and 40 min respectively, combine the filtrates after extraction, and concentrate to 1 / 5 of the filtrate to obtain the astragalus / codonopsis pilosula ferment water extract;

[0024] (5) After mixing tangerine peel and seabuckthorn, add the bacterial liquid of Bifidobacterium longum CICC 24632 to ferment to obtain the tangerine peel / seabuckthorn ferment, and the dosage of the bifidobacterium is 0.6% of the total mass of tangerine peel and seabuckthorn. Fermentation process:

[0025] S1. Mix the powder of tangerine peel and seabuckthorn obtained in step (1) in proportion.

[0026] S2. Add the prepared Bifidobacterium longum CICC 24632 bacterial liquid and RCM liquid medium to the mixture of tangerine peel and seabuckthorn, and stir well. The dosage of the Bifidobacterium longum bacterial liquid is 0.6% of the total mass of tangerine peel and seabuckthorn, and the addition amount of the RCM liquid medium is 2 times the total mass of tangerine peel and seabuckthorn.

[0027] S3. Transfer the mixture added with the Bifidobacterium longum bacterial liquid to a sterile fermentation container, and ensure anaerobic conditions in the container to support the fermentation of Bifidobacterium.

[0028] S4. Ferment at 37°C for 48 hours;

[0029] S5. After fermentation, the obtained fermented product is subjected to water extraction to obtain a filtrate and concentrated to 1 / 5 of the filtrate, that is, the tangerine peel / seabuckthorn fermented water extract is obtained; for subsequent production use. Among them, the water extraction method of the fermented product is: add 8 times the amount of distilled water to the fermented product, extract 3 times, extract for 60 min, 40 min, and 40 min respectively, combine the filtrates after extraction, and concentrate to 1 / 5 of the filtrate to obtain the tangerine peel / seabuckthorn fermented water extract;

[0030] (6) Mix the traditional Chinese medicine extract obtained in step (2), the Astragalus membranaceus / Codonopsis pilosula fermented extract obtained in step (4), and the tangerine peel / seabuckthorn fermented extract obtained in step (5) evenly according to the mass ratio of 50:15:15, and freeze-dry to obtain the traditional Chinese medicine fermentation composition.

[0031] In the third aspect, the present invention provides the application of the above-mentioned traditional Chinese medicine fermentation composition or the traditional Chinese medicine fermentation composition prepared by the above-mentioned preparation method in the preparation of products for improving sleep and having the effects of treating lung cancer and lung nodules.

[0032] In the above technical solution, the traditional Chinese medicine fermentation composition is used to prepare Xuezhi Yu paste.

[0033] In the above technical solution, excipients are added to the traditional Chinese medicine fermentation composition. The addition amount of the excipients is 20% of the total mass of Xuezhi Yu paste. The specific excipient formula is as follows (calculated based on the proportion of Xuezhi Yu paste): tortoise shell glue accounts for 3%, turtle shell glue accounts for 3%, donkey-hide gelatin accounts for 5%, xylitol accounts for 3%, xanthan gum accounts for 1%, and isomaltooligosaccharide accounts for 5%. The purpose is to enhance the lubricity, stability and absorbability of the paste, and at the same time improve the transmission and comfort of the drug effect. Gradually boil under heating conditions until a suitable paste is formed, that is, Xuezhi Yu paste is obtained. During the boiling process, it is necessary to continuously stir to ensure the uniformity and viscosity of the ointment, and finally obtain an ideal paste.

[0034] Based on the traditional Chinese medicine production, the traditional Chinese medicine fermentation composition of the present invention has the following innovation points:

[0035] 1. Application of Group Fermentation Technology

[0036] The present invention adopts the method of group fermentation. Especially during the fermentation of Astragalus membranaceus and Codonopsis pilosula with Lactobacillus, and the fermentation of Exocarpium Citri Grandis and Hippohgae rhamnoides with Bifidobacterium, the fermentation of different strains is utilized to improve the biological activity and absorption rate of traditional Chinese medicines. Through the fermentation of Lactobacillus and Bifidobacterium, the biological transformation of active ingredients in the medicinal materials, such as saponins and polysaccharides, is effectively enhanced, the medicinal efficacy is strengthened, and at the same time, it also has the effects of improving sleep and anti-lung cancer.

[0037] 2. Combination of Carefully Selected Medicinal Materials

[0038] The traditional Chinese medicine fermentation composition of the present invention selects traditional Chinese medicines with multiple pharmacological effects, such as Astragalus membranaceus, Codonopsis pilosula, Ophiopogon japonicus, Gastrodia elata, Rehmannia glutinosa, Curcuma longa, Oyster, Sweet Almond, Prunus davidiana, Exocarpium Citri Grandis, Hippohgae rhamnoides, Ganoderma lucidum, etc. Especially for the fermented medicinal materials, not only the medicinal efficacy of traditional medicinal materials is enhanced, but also their bioavailability and therapeutic effect in the body are improved. For example, Ganoderma lucidum not only helps to improve sleep quality, but also has multiple effects such as anti-tumor, immunomodulation, and lowering blood pressure and blood sugar to protect cardiovascular health, forming a multi-faceted and comprehensive therapeutic effect.

[0039] Compared with the prior art, the beneficial effects of the present invention are:

[0040] 1. Improve sleep disorder symptoms

[0041] According to experimental data, the traditional Chinese medicine fermentation composition of the present invention can significantly improve the sleep disorder symptoms of mice and has strong sedative and tranquilizing effects.

[0042] 2. Anti-lung cancer effect

[0043] After being fermented, the traditional Chinese medicine fermentation composition can effectively inhibit the proliferation of lung cancer cells and the growth of tumor cells. Experiments show that these components play a good anti-cancer effect by regulating immune function and inhibiting the proliferation of tumor cells.

[0044] The traditional Chinese medicine fermentation composition of the present invention is prepared by the method of group fermentation, in which Astragalus membranaceus and Codonopsis pilosula are fermented by Lactobacillus, and Exocarpium Citri Grandis and Hippohgae rhamnoides are fermented by Bifidobacterium. Acting synergistically with other traditional Chinese medicine components, it shows a significant anti-cancer effect in pharmacological research, can effectively inhibit the proliferation of lung nodules and lung cancer cells, and has a certain improvement effect on the Lewis lung cancer model of mice, further providing a scientific basis for the clinical application of the traditional Chinese medicine fermentation composition. At the same time, the traditional Chinese medicine fermentation composition of the present invention can also effectively relieve the sleep disorder symptoms of mice and improve sleep quality, providing a new idea for the treatment of sleep-related diseases. Brief Description of the Drawings

[0045] Figure 1 Bar chart of the effect of the traditional Chinese medicine fermentation composition in the present invention on the serum index CAT of mice with Lewis lung cancer;

[0046] Figure 2 Bar chart of the effect of the traditional Chinese medicine fermentation composition in the present invention on the serum index PFK of mice with Lewis lung cancer;

[0047] Figure 3 Bar chart of the effect of the traditional Chinese medicine fermentation composition on the sleep duration extension of mice;

[0048] Figure 4 Bar chart of the effect of the traditional Chinese medicine fermentation composition on the sleep latency of mice. Detailed implementation mode

[0049] The following specific examples illustrate the implementation mode of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific implementation modes. Various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that, without conflict, the following embodiments and the features in the embodiments can be combined with each other.

[0050] Example 1 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effects of treating lung cancer and lung nodules. The composition is 6 parts of Astragalus membranaceus, 5 parts of Codonopsis pilosula, 7 parts of Ophiopogon japonicus, 7 parts of Gastrodia elata, 7 parts of Rehmannia glutinosa, 6 parts of Curcuma longa, 6 parts of Oyster, 7 parts of Sweet almond, 6 parts of Prunus davidiana, 10 parts of Citrus reticulata Blanco cv. Tomentosa, 7 parts of Hippohgae rhamnoides, 6 parts of Angelica sinensis, 5 parts of Prunus persica, 4 parts of Lotus leaf, 5 parts of Jasminum sambac, 4 parts of Chrysanthemum morifolium, and 3 parts of Ganoderma lucidum. Among them, Astragalus membranaceus and Codonopsis pilosula are fermented by Lactobacillus, and Citrus reticulata Blanco cv. Tomentosa and Hippohgae rhamnoides are fermented by Bifidobacterium. The Lactobacillus used is Lactobacillus acidophilus CICC 6243, and the Bifidobacterium used is Bifidobacterium longum CICC 24632. Both Lactobacillus acidophilus CICC 6243 and Bifidobacterium longum CICC 24632 are purchased from the China Center for Industrial Culture Collection of Microorganisms. The dosage of Lactobacillus acidophilus CICC 6243 is 0.6% of the total mass of Astragalus membranaceus and Codonopsis pilosula, and the dosage of Bifidobacterium longum CICC 24632 is 0.6% of the total mass of Citrus reticulata Blanco cv. Tomentosa and Hippohgae rhamnoides. The freeze-dried strain powders of Lactobacillus acidophilus CICC 6243 and Bifidobacterium longum CICC 24632 are dissolved in sterile water, and after conventional activation, they are respectively added to MRS liquid medium and RCM liquid medium to be cultured into fermentation bacterial liquids, and the fermentation concentrations of the two bacteria are both adjusted to 1×10 9 CFU / mL.

[0051] Preparation method of the traditional Chinese medicine fermentation composition in this example:

[0052] (1) Weigh all the traditional Chinese medicine raw materials according to the formula of this embodiment, wash and dry them respectively, and then pulverize them into 80-mesh powder;

[0053] (2) Mix the powders of Ophiopogon japonicus, Gastrodia elata, Rehmannia glutinosa, Curcuma longa, Oyster, Sweet Almond, Prunus japonica, Angelica sinensis, Prunus persica, Lotus Leaf, Jasmine, Chrysanthemum, Ganoderma lucidum. After mixing, add 8 times the amount of distilled water and perform extraction 3 times, with extraction times of 60 h, 40 min, and 40 min respectively. After the extraction is completed, combine the filtrates and concentrate them to 1 / 5 of the original volume to obtain the traditional Chinese medicine extract;

[0054] (3) Prepare Lactobacillus acidophilus CICC 6243 bacterial liquid with a concentration of 1×10 9 CFU / ml and Bifidobacterium longum CICC 24632 bacterial liquid with a concentration of 1×10 9 CFU / ml;

[0055] (4) After mixing Astragalus membranaceus and Codonopsis pilosula, add Lactobacillus acidophilus CICC 6243 bacterial liquid to ferment to obtain Astragalus membranaceus / Codonopsis pilosula fermentation extract. The dosage of the lactobacillus is 0.6% of the total mass of Astragalus membranaceus and Codonopsis pilosula. The fermentation process is as follows:

[0056] S1. Mix the powders of Astragalus membranaceus and Codonopsis pilosula obtained in step (1) according to the ratio;

[0057] S2. Add the prepared Lactobacillus acidophilus CICC 6243 bacterial liquid and MRS liquid medium to the mixture of Astragalus membranaceus and Codonopsis pilosula and stir well. The dosage of the bifidobacterium bacterial liquid is 0.6% of the total mass of Astragalus membranaceus and Codonopsis pilosula, and the addition amount of MRS liquid medium is 2 times the total mass of Astragalus membranaceus and Codonopsis pilosula;

[0058] S3. Transfer the mixture added with Lactobacillus acidophilus bacterial liquid to a sterile fermentation container to ensure anaerobic conditions in the container to support the growth of Lactobacillus acidophilus CICC 6243;

[0059] S4. Ferment at a constant temperature of 37°C for 48 hours. After the fermentation is completed, perform water extraction on the obtained fermented product, take the filtrate and concentrate it to 1 / 5 of the filtrate to obtain Astragalus membranaceus / Codonopsis pilosula fermentation extract;

[0060] (5) After mixing Exocarpium Citri Grandis and Hippohgae rhamnoides, add Bifidobacterium longum CICC 24632 bacterial liquid to ferment to obtain Exocarpium Citri Grandis / Hippohgae rhamnoides fermentation extract. The dosage of the Bifidobacterium longum is 0.6% of the total mass of Exocarpium Citri Grandis and Hippohgae rhamnoides. The fermentation process is as follows:

[0061] S1. Mix the powders of Exocarpium Citri Grandis and Hippohgae rhamnoides obtained in step (1) according to the ratio;

[0062] S2. Add the prepared Bifidobacterium longum CICC 24632 bacterial liquid and RCM liquid medium to the mixture of Exocarpium Citri Grandis and Hippohgae rhamnoides L., and stir well. The dosage of the Bifidobacterium longum bacterial liquid is 0.6% of the total mass of Exocarpium Citri Grandis and Hippohgae rhamnoides L., and the added amount of the RCM liquid medium is 2 times the total mass of Exocarpium Citri Grandis and Hippohgae rhamnoides L.;

[0063] S3. Transfer the mixture added with the Bifidobacterium longum bacterial liquid to a sterile fermentation container, and ensure anaerobic conditions in the container to support the fermentation of Bifidobacterium longum CICC 24632.

[0064] S4. Conduct fermentation at 37 °C for 48 hours;

[0065] S5. After the fermentation ends, subject the obtained fermented product to water extraction, take the filtrate and concentrate it to 1 / 5 of the filtrate, and thus obtain the water extract of fermented Exocarpium Citri Grandis / Hippohgae rhamnoides L.;

[0066] (6) Mix the Chinese medicinal material extract obtained in step (2), the fermented extract of Astragalus membranaceus / Codonopsis pilosula obtained in step (4), and the fermented extract of Exocarpium Citri Grandis / Hippohgae rhamnoides L. obtained in step (5) evenly according to the mass ratio of 50:15:15, and thus obtain the Chinese medicinal fermentation composition.

[0067] Example 2 of the present application provides a Chinese medicinal fermentation composition that helps improve sleep and has the effects of treating lung cancer and lung nodules. The composition is: 7 parts of Astragalus membranaceus, 6 parts of Codonopsis pilosula, 6 parts of Ophiopogon japonicus, 8 parts of Gastrodia elata, 5 parts of Rehmannia glutinosa, 5 parts of Curcuma longa, 7 parts of Oyster, 6 parts of Sweet Almond, 9 parts of Prunus davidiana, 8 parts of Exocarpium Citri Grandis, 5 parts of Hippohgae rhamnoides L., 6 parts of Angelica sinensis, 7 parts of Prunus persica, 6 parts of Lotus Leaf, 5 parts of Jasmine, 5 parts of Chrysanthemum, 2 parts of Ganoderma lucidum. The type and dosage of the bacterial liquid are the same as those in Example 1, and the preparation method is the same as that in Example 1.

[0068] Example 3 of the present application provides a Chinese medicinal fermentation composition that helps improve sleep and has the effects of treating lung cancer and lung nodules. The formula is: 8 parts of Astragalus membranaceus, 4 parts of Codonopsis pilosula, 8 parts of Ophiopogon japonicus, 6 parts of Gastrodia elata, 8 parts of Rehmannia glutinosa, 7 parts of Curcuma longa, 5 parts of Oyster, 8 parts of Sweet Almond, 6 parts of Prunus davidiana, 9 parts of Exocarpium Citri Grandis, 4 parts of Hippohgae rhamnoides L., 7 parts of Angelica sinensis, 5 parts of Prunus persica, 5 parts of Lotus Leaf, 6 parts of Jasmine, 4 parts of Chrysanthemum, 4 parts of Ganoderma lucidum. The type and dosage of the bacterial liquid are the same as those in Example 1, and the preparation method is the same as that in Example 1.

[0069] Example 4 of the present application provides a Chinese medicinal fermentation composition that helps improve sleep and has the effects of treating lung cancer and lung nodules. The formula is: 5 parts of Astragalus membranaceus, 5 parts of Codonopsis pilosula, 8 parts of Ophiopogon japonicus, 6 parts of Gastrodia elata, 6 parts of Rehmannia glutinosa, 7 parts of Curcuma longa, 7 parts of Oyster, 6 parts of Sweet Almond, 7 parts of Prunus davidiana, 12 parts of Exocarpium Citri Grandis, 7 parts of Hippohgae rhamnoides L., 8 parts of Angelica sinensis, 6 parts of Prunus persica, 3 parts of Lotus Leaf, 4 parts of Jasmine, 3 parts of Chrysanthemum, 5 parts of Ganoderma lucidum. The type and dosage of the bacterial liquid are the same as those in Example 1, and the preparation method is the same as that in Example 1.

[0070] Example 5 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. The formula is as follows: 6 parts of Astragalus membranaceus, 4 parts of Codonopsis pilosula, 7 parts of Ophiopogon japonicus, 7 parts of Gastrodia elata, 6 parts of Rehmannia glutinosa, 6 parts of Curcuma longa, 6 parts of Oyster, 7 parts of Sweet Almond, 8 parts of Prunus davidiana var. ansu, 11 parts of Citrus reticulata Blanco cv. Tomentosa, 6 parts of Hippohgae rhamnoides L., 6 parts of Angelica sinensis, 8 parts of Prunus persica, 4 parts of Lotus leaf, 7 parts of Jasmine, 4 parts of Chrysanthemum morifolium Ramat., 3 parts of Ganoderma lucidum. The type and dosage of the bacterial liquid are the same as those in Example 1, and the preparation method is the same as that in Example 1.

[0071] Comparative Example 1 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Astragalus membranaceus is not used.

[0072] Comparative Example 2 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Codonopsis pilosula is not used.

[0073] Comparative Example 3 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Citrus reticulata Blanco cv. Tomentosa is not used.

[0074] Comparative Example 4 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Hippohgae rhamnoides L. is not used.

[0075] Comparative Example 5 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Lactobacillus acidophilus CICC 6243 is not used, and Astragalus membranaceus and Codonopsis pilosula are treated by direct fermentation.

[0076] Comparative Example 6 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Bifidobacterium longum CICC 24632 is not used, and Citrus reticulata Blanco cv. Tomentosa and Hippohgae rhamnoides L. are treated by direct fermentation.

[0077] Comparative Example 7 of the present application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. It is similar to Example 1, except that Lactobacillus acidophilus CICC 6243 is replaced with other Lactobacillus acidophilus. The other Lactobacillus is Lactobacillus acidophilus CICC 22162, purchased from China Center for Industrial Culture Collection.

[0078] Comparative Example 8 of this application provides a traditional Chinese medicine fermentation composition that helps improve sleep and has the effect of treating lung cancer and lung nodules. Similar to Example 1, the difference is that Bifidobacterium longum CICC 24632 is replaced with other Bifidobacterium. The other Bifidobacterium is Bifidobacterium longum CICC 25033, which is purchased from the China Center for Industrial Culture Collection Management.

[0079] I. Efficacy evaluation study of traditional Chinese medicine fermentation composition on lung nodules and lung cancer

[0080] 1.1 Effects on the proliferation of lung nodules and lung cancer cells

[0081] According to the provided experimental content, the following is a summary of the grouping objects, drugs used in the grouping experiments, specific experimental methods, and detection indicators of the experimental results for each experiment:

[0082] (1). Experimental materials:

[0083] 1. Test drugs: Traditional Chinese medicine fermentation compositions obtained from Examples 1 - 5 and Comparative Examples 1 - 8;

[0084] 2. Instruments: BioTek Synergy H1 multi-functional microplate reader, QX200 TM Droplet Digital TM PCR System

[0085] (2). Experimental methods and summary:

[0086] 1. Effects on the proliferation of lung nodules and lung cancer cells

[0087] Grouping objects: Lung nodule and lung cancer cells (A549, Lewis cells) of humans and mice treated with traditional Chinese medicine fermentation compositions from Examples 1 - 5 and Comparative Examples 1 - 8 (at concentrations of 25 μg / mL and 100 μg / mL, diluted with deionized water), and negative control group (A549, Lewis cells only with culture medium).

[0088] Drugs: Traditional Chinese medicine fermentation composition (treated at concentrations of 25 μg / mL and 100 μg / mL)

[0089] Experimental methods:

[0090] Culture A549 cells and Lewis cells using RPMI medium containing 10% FBS, and culture them under the conditions of 37°C and 5% CO2.

[0091] When the cells grow to 70% - 80%, passage them with trypsin.

[0092] Cells in the logarithmic growth phase were inoculated into 96-well plates and incubated for 12 hours. After the cells adhered to the plate, they were treated with media containing different concentrations of the traditional Chinese medicine fermented composition for 24 hours.

[0093] A negative control group (A549 and Lewis cells treated only with media) was set up.

[0094] The above same experiment was repeated with the traditional Chinese medicine fermented compositions of Examples 1 - 5 and Comparative Examples 1 - 8, and each sample was repeated 5 times.

[0095] Experimental detection indicators:

[0096] The cell proliferation was detected by the CCK-8 method, and the specific process was as follows:

[0097] 1. Cell culture: The cells to be detected were inoculated in 96-well plates and cultured under appropriate conditions.

[0098] 2. Adding CCK-8 reagent: When the cells grew to an appropriate density, CCK-8 reagent was added. The reagent reacted with the metabolites of living cells to generate an orange-yellow product.

[0099] 3. Incubation: Continue to culture for a period of time (2 hours) to allow the reaction to proceed fully.

[0100] 4. Measuring absorbance: The OD value of each well was measured using a microplate reader at a wavelength of 450 nm.

[0101] 5. Data analysis: The cell proliferation was evaluated by measuring the OD values of different treatment groups.

[0102] Calculation formula for cell proliferation rate (cell viability):

[0103] Cell proliferation rate (%) = {((OD value of experimental group - OD value of blank group) / (OD value of negative control group - OD value of blank group))} × 100

[0104] Where:

[0105] 6. OD value of experimental group: The absorbance value of the treatment group.

[0106] 7. OD value of negative control group: The absorbance value of the group of cells without treatment or under normal culture.

[0107] 8. OD value of blank group: The absorbance value of the medium without cells.

[0108] The results are shown in Table 1 and Table 2. The traditional Chinese medicine fermentation composition has obvious inhibitory effects on A549 and Lewis cells at 25 μg / mL and 100 μg / mL for 24 hours (P < 0.05); compared with Comparative Examples 1-8, the cell proliferation rates of Examples 1-5 are significantly reduced, indicating that the traditional Chinese medicine fermentation composition of the present invention, through the method of grouped fermentation, the astragalus membranaceus and codonopsis pilosula are fermented by lactobacillus, and the citrus reticulata blanco and seabuckthorn are fermented by bifidobacterium and then act synergistically with other traditional Chinese medicine components, can significantly inhibit the proliferation of lung nodule lung cancer cells.

[0109] Table 1 Cell viability of A549 cells for 24 hours (%)

[0110] Group 25 ug / mL(%) 100 ug / mL(%) Example 1 53.47±6.00 8.69±0.75 Example 2 54.12±5.87 8.92±0.64 Example 3 54.37±5.93 9.66±0.70 Example 4 53.23±5.85 8.23±0.72 Example 5 51.85±5.79 8.18±0.98 Comparative Example 1 75.41±6.20 26.36±2.64 Comparative Example 2 77.28±6.10 27.11±2.05 Comparative Example 3 76.84±6.15 26.62±2.12 Comparative Example 4 74.71±6.12 25.38±1.78 Comparative Example 5 76.56±6.14 26.54±2.10 Comparative Example 6 77.79±6.18 27.23±2.24 Comparative Example 7 71.02±6.22 24.27±2.15 Comparative Example 8 72.92±6.19 24.71±2.12

[0111] Table 2 Cell viability of Lewis cells for 24 hours (%)

[0112]

[0113]

[0114] 1.2. Effect on the proliferation of lung nodule lung cancer cells (serum pharmacology)

[0115] Grouping objects: Sprague-Dawley rats (5 in the negative control group, 5 * 13 in the traditional Chinese medicine fermentation composition group, 5 in the cyclophosphamide control group)

[0116] Drugs for gavage in each group:

[0117] Negative control group: normal saline;

[0118] Traditional Chinese medicine fermentation compositions of Examples 1-5, 80 mg / kg;

[0119] Traditional Chinese medicine fermentation compositions of Comparative Examples 1-8, 80 mg / kg;

[0120] Positive control group: cyclophosphamide 100 mg / kg;

[0121] Experimental method: The rats were gavaged with drugs according to the above grouping for 7 days, once a day. Blood was taken 1 hour after the last gavage to separate serum. The separated serum was used to treat A549 cells, and 28 groups were set:

[0122] Negative control group: 10% normal culture (normal saline) with serum + RPMI-1640 medium

[0123] Positive control group: 10% cyclophosphamide serum + RPMI-1640 medium

[0124] Examples 1-5, Comparative Examples 1-8: Chinese medicine fermentation composition group: 10% composition-containing drug serum + RPMI-1640 medium. Each group of cells was treated for 24 hours and 48 hours, and then detected.

[0125] Experimental detection indicators:

[0126] The process of detecting cell proliferation by CCK-8 method is as follows:

[0127] 1. Cell culture: Inoculate the cells to be detected in a 96-well plate and culture them under appropriate conditions.

[0128] 2. Add CCK-8 reagent: When the cells grow to an appropriate density, add CCK-8 reagent. The reagent reacts with the metabolites of living cells to produce an orange-yellow product.

[0129] 3. Incubation: Continue to culture for a period of time (2 hours) to allow the reaction to proceed fully.

[0130] 4. Measure absorbance: Use an enzyme-linked immunosorbent assay (ELISA) reader to measure the OD value of each well at a wavelength of 450 nm.

[0131] 5. Data analysis: Evaluate cell proliferation by measuring the OD values at different time points or different treatment groups.

[0132] Calculation formula for cell proliferation rate (cell viability):

[0133] Cell proliferation rate (%) = {((OD value of experimental group - OD value of blank group) / (OD value of negative control group - OD value of blank group))} × 100

[0134] Where:

[0135] 6. OD value of experimental group: Absorbance value of the treatment group.

[0136] 7. OD value of negative control group: Absorbance value of the cell group cultured normally (with physiological saline).

[0137] 8. OD value of blank group: Absorbance value of the medium without cells.

[0138] The results are shown in Table 3 and Table 4. The data at 24 h and 48 h show that the Chinese medicine fermentation compositions of Examples 1-5 and Comparative Examples 1-8, as well as cyclophosphamide, can significantly inhibit the activity of A549 cells; compared with Comparative Examples 1-8, the cell proliferation rate of Examples 1-5 is significantly reduced, indicating that the Chinese medicine fermentation composition of the present invention, through the method of grouped fermentation, the fermentation of Astragalus membranaceus and Codonopsis pilosula by Lactobacillus, and the fermentation of Citrus reticulata Blanco and Hippohgae rhamnoides L. by Bifidobacterium, and the synergistic effect with other Chinese medicine components, can significantly inhibit the proliferation of lung nodule lung cancer cells.

[0139] Table 3 A549 cell viability at 24 h (%)

[0140] Group Cell Proliferation Rate(%) Example 1 84.08±4.72 Example 2 84.34±5.12 Example 3 85.16±5.32 Example 4 83.58±4.92 Example 5 83.46±5.42 Comparative Example 1 92.71±6.38 Comparative Example 2 94.17±6.05 Comparative Example 3 93.85±6.34 Comparative Example 4 92.28±6.17 Comparative Example 5 93.56±5.32 Comparative Example 6 94.63±6.56 Comparative Example 7 91.33±6.74 Comparative Example 8 91.48±6.18 Positive Drug Group 93.18±2.29

[0141] Table 4 A549 cell viability at 48 h (%)

[0142] Group Cell Proliferation Rate(%) Example 1 57.37±5.12 Example 2 58.15±4.85 Example 3 58.91±5.05 Example 4 56.44±4.70 Example 5 56.08±3.95 Comparative Example 1 81.19±5.20 Comparative Example 2 83.59±5.60 Comparative Example 3 82.82±4.90 Comparative Example 4 80.27±3.88 Comparative Example 5 81.84±5.15 Comparative Example 6 84.23±4.20 Comparative Example 7 79.04±3.95 Comparative Example 8 79.64±4.60 Positive Drug Group 78.32±8.15

[0143] 1.3 Pharmacodynamic evaluation of traditional Chinese medicine fermentation composition on murine lung cancer

[0144] The Lewis lung cancer mouse model was used to evaluate the remission effect of traditional Chinese medicine fermentation composition on lung cancer by detecting multiple serum biochemical indexes.

[0145] 1.3.1 Experimental animals, grouping and model construction: Eighty male C57BL / 6 mice, weighing 20±2 g, were used in the experiment. The animals were housed in an environment with a room temperature of 25±1 °C, a relative humidity of 60%, and a normal 12-hour day-night cycle. They had free access to food and water. The animals were randomly divided into 16 groups, namely the blank group, the model group, the positive drug cyclophosphamide Cytoxan (100 mg / kg, 0.2 mL / 10 g) group, and the traditional Chinese medicine fermentation composition (150 mg / kg, 0.2 mL / 10 g) group (divided into Examples 1-5 and Comparative Examples 1-8, a total of 13 groups), with 5 animals in each group.

[0146] Modeling method: After anesthetizing the C57 mice, place them in the right lateral position on the operating table. Shave the hair on the left axilla of the mice and disinfect it with 75% ethanol. Make a 5-mm incision at about 1.5 cm above the upper edge of the costal arch in the left anterior axillary line. Separate the skin and subcutaneous tissue to expose the chest wall until the pink lung lobe can be seen moving up and down with breathing. Mix 50 ul of PBS suspension containing tumor cells (10 6 cells per mouse) and 50 ul of Matrigel matrix and place it on ice. Vertically inject it into the left lung with a microinjector, with the injection depth of about 3 mm. Stop the needle for 20 seconds after injection, and suture the incision with 1-2 stitches after removing the needle.

[0147] Cell line used: Murine LLC (Lewis)-luc cells (murine lung cancer cells - luciferase-labeled), catalog number LZQ0009, Shanghai Zhongqiao Xinzhou Co., Ltd., cultured in DMEM medium containing 10% fetal bovine serum.

[0148] Drug administration method: After successful modeling, the traditional Chinese medicine fermentation composition was administered by gavage at 150 mg / kg, 0.2 mL / 10 g once a day; the positive group was intraperitoneally injected with cyclophosphamide at 100 mg / kg, 0.2 mL / 10 g every other day; the model group was given an equal volume of distilled water, and the drug administration period was 14 days. Blank group: No modeling, no drug administration.

[0149] 1.3.2. Serum biochemical index detection: 24 hours after the last administration, blood was collected by enucleating the eyeballs. After standing at 4°C for 30 minutes, it was centrifuged at 3500 r / min for 10 minutes, and the supernatant was stored at -80°C for later measurement. The serum biochemical indexes detected in this project include: catalase (CAT), phosphofructokinase (PFK).

[0150] Table 5: Changes in serum CAT index

[0151] Group Serum CAT (U / mL) Blank Group 85.12±7.42 Model Group 50.32±5.08 CTX Group 80.67±7.16 Example 1 75.45±6.25 Example 2 73.58±7.13 Example 3 72.22±6.03 Example 4 72.87±6.59 Example 5 73.78±6.04 Comparative Example 1 59.46±5.17 Comparative Example 2 60.32±5.56 Comparative Example 3 58.97±5.29 Comparative Example 4 60.11±5.18 Comparative Example 5 55.06±5.12 Comparative Example 6 54.89±5.23 Comparative Example 7 58.72±5.14 Comparative Example 8 58.84±5.20

[0152] Table 6: Serum PFK enzyme activity

[0153] Group PFK Enzyme Activity (U / mL) Blank Group 12.15±0.98 Model Group 22.08±1.67 CTX Group 11.05±0.85 Example 1 11.52±0.86 Example 2 11.19±0.93 Example 3 10.86±0.79 Example 4 11.37±1.08 Example 5 10.93±0.84 Comparative Example 1 15.92±1.32 Comparative Example 2 16.17±1.18 Comparative Example 3 15.58±1.24 Comparative Example 4 15.81±1.12 Comparative Example 5 16.03±1.20 Comparative Example 6 16.45±1.25 Comparative Example 7 15.89±1.19 Comparative Example 8 15.62±1.16

[0154] From Table 5, Table 6 and Figure 1 、 Figure 2 it can be seen that compared with the blank group (Control), the levels of catalase (CAT) in the serum of model (Model) mice were significantly decreased; the levels of phosphofructokinase (PFK) were significantly increased, indicating that the model was successfully established; compared with Comparative Examples 1-8, the content of CAT was significantly increased and the content of PFK was significantly decreased in the traditional Chinese medicine fermentation composition of Examples 1-5. It is indicated that the traditional Chinese medicine fermentation composition, prepared by the method of grouped fermentation, in which Astragalus membranaceus and Codonopsis pilosula are fermented by Lactobacillus, and Citrus reticulata Blanco cv. Tomentosa and Hippohgae rhamnoides are fermented by Bifidobacterium, synergistically acts with other traditional Chinese medicine components and has a certain improvement effect on Lewis lung cancer in mice.

[0155] II. Therapeutic effect of traditional Chinese medicine fermentation composition on sleep disorder in mice

[0156] This experiment aimed to study the therapeutic effect of traditional Chinese medicine fermentation composition on sleep disorder in mice, and mainly evaluated its effects on sleep latency (time to fall asleep), sleep duration and sleep depth.

[0157] 1. Experimental subjects and grouping

[0158] The experimental subjects were male SPF-grade ICR mice aged 8-10 weeks, weighing 20-25 grams. After 7 days of adaptive feeding before the experiment, the mice were randomly divided into 14 groups, with 10 mice in each group, and there was no statistical difference among the groups of mice; each group was as follows:

[0159] Normal control group: normal saline, 0.1 mL / 10 g;

[0160] Examples 1-5: The traditional Chinese medicine fermentation composition obtained in Examples 1-5 (diluted with deionized water at 150 mg / kg, 0.1 mL / 10 g);

[0161] Comparative Examples 1 - 8: The traditional Chinese medicine fermentation composition obtained in Comparative Examples 1 - 8 (diluted with deionized water at 150 mg / kg, 0.1 mL / 10 g).

[0162] 2. Experimental methods

[0163] Each group of mice was administered the drug once in the morning and once in the evening every day, with a gavage volume of 0.1 ml / 10 g each time, and continuously gavaged for 14 days. The following experiments were carried out after the last gavage:

[0164] 2.1 Direct sleep experiment in mice

[0165] The direct sleep experiment in mice was carried out according to the "Technical Specification for Inspection and Evaluation of Health Foods" (2003 Edition). Whether sleep occurred after the last administration was observed. Sleep was indicated by the disappearance of the righting reflex in mice. When the mouse was in the dorsal recumbent position and could not right itself for more than 60 s, it was considered that the righting reflex had disappeared and the mouse had entered sleep. The recovery of the righting reflex indicated that the animal was awake. The time from the disappearance of the righting reflex to its recovery was the direct sleep duration. The number of sleeping animals and the direct sleep time of each group of mice were recorded, and the results are shown in Table 7.

[0166] 2.2 Experiment on prolonging the sleep time of pentobarbital sodium in mice

[0167] The experiment on prolonging the sleep time of pentobarbital sodium was carried out according to the "Technical Specification for Inspection and Evaluation of Health Foods" (2003 Edition). 20 minutes after the last gavage, each group of mice was intraperitoneally injected with a pentobarbital sodium solution (35 mg / kg·BW), and the injection volume was 0.1 mL / 10 g. Sleep was indicated by the disappearance of the righting reflex in mice. When the mouse was in the dorsal recumbent position and could not right itself for more than 60 s, it was considered that the righting reflex had disappeared and the mouse had entered sleep. The recovery of the righting reflex indicated that the animal was awake. The time from the disappearance of the righting reflex to its recovery after injecting pentobarbital sodium was recorded as the prolonged sleep duration, and the results are shown in Table 8.

[0168] 2.3 Sleep latency experiment of sodium barbital in mice

[0169] The sleep latency experiment of sodium barbital was carried out according to the "Technical Specification for Inspection and Evaluation of Health Foods" (2003 Edition). 20 minutes after the last gavage, each group of mice was intraperitoneally injected with sodium barbital (200 mg / kg·BW), and the injection volume was 0.1 ml / 10 g. Using the disappearance of the reflex as an indicator, the sleep latency of each group was recorded, and the results are shown in Table 9.

[0170] Table 7: Number of animals falling asleep and direct sleep duration (min)

[0171] Group Number of Animals / piece Number of Animals Falling Asleep / piece Direct Sleep Duration (min) Control Group 10 0 - Example 1 10 0 - Example 2 10 0 - Example 3 10 0 - Example 4 10 0 - Example 5 10 0 - Comparative Example 1 10 0 - Comparative Example 2 10 0 - Comparative Example 3 10 0 - Comparative Example 4 10 0 - Comparative Example 5 10 0 - Comparative Example 6 10 0 - Comparative Example 7 10 0 - Comparative Example 8 10 0 -

[0172] Table 8: Prolonged sleep duration (min)

[0173]

[0174]

[0175] Table 9: Sleep latency (min)

[0176] Group Sleep Latency (min) Control Group 15.58±1.50 Example 1 7.82±1.25 Example 2 9.14±1.21 Example 3 8.89±1.32 Example 4 7.54±1.28 Example 5 8.22±1.36 Comparative Example 1 13.51±1.17 Comparative Example 2 12.69±1.29 Comparative Example 3 13.48±1.33 Comparative Example 4 12.81±1.18 Comparative Example 5 13.62±1.30 Comparative Example 6 13.78±1.35 Comparative Example 7 12.23±1.26 Comparative Example 8 13.14±1.21

[0177] As can be seen from Table 7, the number of mice that fell asleep in Examples 1-5 and Comparative Examples 1-8 was 0. Compared with the control group, there was no statistically significant difference in Examples 1-5 and Comparative Examples 1-8, indicating that the fermented traditional Chinese medicine composition has no effect on direct sleep in mice. From Tables 8-9, Figure 3 - 4 it can be seen that compared with the control group, the sleep time of the mice in Examples 1-5 and Comparative Examples 1-8 was significantly prolonged, the sleep latency was significantly shortened, and the mice fell asleep faster. Compared with Comparative Examples 1-8, the sleep time of the mice in Examples 1-5 was significantly prolonged, the sleep latency was significantly shortened, and the mice fell asleep faster. This shows that the traditional Chinese medicine fermentation composition can effectively shorten the sleep onset time of mice, prolong the sleep duration, and has the effect of improving sleep.

[0178] The above-described embodiments merely represent specific embodiments of the present invention, and the description thereof is relatively specific and detailed, but it should not be construed as a limitation on the scope of the invention patent. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention.

Claims

1. A traditional Chinese medicine fermentation composition that helps improve sleep and has the effects of treating lung cancer and lung nodules, characterized in that: The raw materials of the blood stasis paste include, by weight: 5-8 parts of astragalus, 4-6 parts of codonopsis, 6-8 parts of ophiopogon, 6-8 parts of gastrodia, 5-8 parts of rehmannia, 5-7 parts of turmeric, 5-7 parts of oyster, 6-8 parts of sweet almond, 6-9 parts of prunus mume, 8-12 parts of tangerine peel, 4-7 parts of sea buckthorn, 6-9 parts of angelica, 5-8 parts of peach kernel, 3-6 parts of lotus leaf, 4-7 parts of jasmine, 3-5 parts of chrysanthemum, and 2-5 parts of ganoderma; among them, astragalus and codonopsis are fermented with lactobacillus, and tangerine peel and sea buckthorn are fermented with bifidobacterium.

2. The traditional Chinese medicine fermentation composition according to claim 1, which is helpful for improving sleep and has the functions of treating lung cancer and lung nodules, is characterized in that: The raw materials include 6 parts of astragalus, 5 parts of codonopsis, 7 parts of ophiopogon, 7 parts of gastrodia, 7 parts of rehmannia, 6 parts of turmeric, 6 parts of oyster, 7 parts of sweet almond, 6 parts of prunes, 10 parts of orange peel, 7 parts of sea buckthorn, 6 parts of angelica, 5 parts of peach kernel, 4 parts of lotus leaf, 5 parts of jasmine, 4 parts of chrysanthemum, and 3 parts of ganoderma.

3. The traditional Chinese medicine fermentation composition according to claim 1, which is helpful for improving sleep and has the functions of treating lung cancer and lung nodules, is characterized in that: After mixing the Chinese medicinal materials except astragalus, codonopsis, tangerine peel and seabuckthorn in proportion, add 8-10 times the amount of distilled water, and perform extraction three times. After the extraction, combine the filtrates and concentrate them to 1 / 5 of the filtrate to obtain the Chinese medicinal material extract.

4. The traditional Chinese medicine fermentation composition according to claim 3, which is helpful for improving sleep and has the functions of treating lung cancer and lung nodules, is characterized in that: The lactobacillus selected was Lactobacillus acidophilus CICC 6243, and the bifidobacterium selected was Bifidobacterium longum CICC24632.

5. The traditional Chinese medicine fermentation composition according to claim 4, which is helpful for improving sleep and has the functions of treating lung cancer and pulmonary nodules, is characterized in that: The concentration of the strains contained in the Lactobacillus bacterial solution is 10 8 ~10 9 CFU / ml, and the concentration of the strains contained in the Bifidobacterium bacterial solution is 10 8 ~10 9 CFU / ml.

6. The traditional Chinese medicine fermentation composition according to claim 5, which is helpful for improving sleep and has the functions of treating lung cancer and lung nodules, is characterized in that: The dosage of the lactobacillus liquid is 0.6% of the total weight of astragalus and codonopsis, and the dosage of the bifidobacterium liquid is 0.6% of the total weight of tangerine peel and seabuckthorn.

7. The preparation method of the traditional Chinese medicine fermentation composition according to any one of claims 1-6, which is helpful for improving sleep and has the functions of treating lung cancer and lung nodules, is characterized in that: The following steps are involved: (1) Weigh 5-8 parts of Astragalus, 4-6 parts of Codonopsis, 6-8 parts of Radix Ophiopogonis, 6-8 parts of Gastrodia, 5-8 parts of Rehmannia, 5-7 parts of Curcuma, 5-7 parts of Oyster, 6-8 parts of Apricot, 6-9 parts of Prunus Amygdalus, 8-12 parts of Citrus aurantium, 4-7 parts of Hippophae rhamnoides, 6-9 parts of Angelica sinensis, 5-8 parts of Peach Kernel, 3-6 parts of Lotus Leaf, 4-7 parts of Jasmine, 3-5 parts of Chrysanthemum, and 2-5 parts of Ganoderma Lucidum, wash and dry them respectively, and then grind them into 80 mesh powder; (2) Mix 6-8 parts of Ophiopogon japonicus, 6-8 parts of Gastrodia elata, 5-8 parts of Rehmannia root, 5-7 parts of Curcuma longa, 5-7 parts of Oyster, 6-8 parts of Apricot kernel, 6-9 parts of Prunus armeniaca, 6-9 parts of Angelica sinensis, 5-8 parts of Peach kernel, 3-6 parts of Lotus leaf, 4-7 parts of Jasmine, 3-5 parts of Chrysanthemum, and 2-5 parts of Ganoderma lucidum, add 8-10 times the amount of distilled water after mixing, perform extraction 3 times, combine the filtrates after extraction, and concentrate to 1 / 5 of the filtrate to obtain a Chinese herbal medicine extract; (3) Prepare Lactobacillus acidophilus CICC 6243 bacterial liquid with a concentration of 1×10 8 ~10 9 CFU / ml and Bifidobacterium longum CICC 24632 bacterial liquid with a concentration of 1×10 8 ~10 9 CFU / ml; (4) After the astragalus and codonopsis are mixed, a Lactobacillus acidophilus CICC 6243 bacterial solution is added to ferment to obtain an astragalus / codonopsis fermented extract. The fermentation process is as follows: S1, mixing the powders of Astragalus and Codonopsis obtained in step (1) in proportion to obtain a mixture; S2, adding the prepared Lactobacillus acidophilus CICC 6243 bacterial solution and MRS liquid culture medium to the mixture of Astragalus and Codonopsis pilosula and stirring well, wherein the amount of Lactobacillus acidophilus CICC 6243 bacterial solution added is 0.6% of the total mass of Astragalus and Codonopsis pilosula; S3, transferring the mixture with the Lactobacillus acidophilus bacterial solution into a sterile fermentation container, and ensuring anaerobic conditions in the container to support the growth of Lactobacillus acidophilus CICC 6243; S4. Incubate at a constant temperature of 37 °C for 48 hours. After the fermentation is completed, subject the obtained fermented product to water extraction to obtain a filtrate and concentrate it to 1 / 5 of the filtrate, thereby obtaining the Astragalus membranaceus / Codonopsis pilosula fermented extract. (5) After mixing tangerine peel and seabuckthorn, add the liquid culture of Bifidobacterium longum CICC 24632 and ferment to obtain the tangerine peel / seabuckthorn fermented extract. The fermentation process is as follows: S1. Mix the powders of tangerine peel and seabuckthorn obtained in step (1) in proportion. S2. Add the prepared liquid culture of Bifidobacterium longum CICC 24632 and RCM liquid medium to the mixture of tangerine peel and seabuckthorn, and stir well. The dosage of the Bifidobacterium longum liquid culture is 0.6% of the total mass of tangerine peel and seabuckthorn. S3. Transfer the mixture added with the Bifidobacterium longum liquid culture to a sterile fermentation container, and ensure anaerobic conditions in the container to support the fermentation of Bifidobacterium longum CICC 24632. S4. Ferment at 37 °C for 48 hours. S5. After the fermentation is completed, subject the obtained fermented product to water extraction to obtain a filtrate and concentrate it to 1 / 5 of the filtrate, thereby obtaining the tangerine peel / seabuckthorn fermented water extract. (6) Mix the Chinese herbal medicine extract obtained in step (2), the Astragalus membranaceus / Codonopsis pilosula fermented extract obtained in step (4), and the tangerine peel / seabuckthorn fermented extract obtained in step (5) evenly according to the mass ratio of 50:15:15, thereby obtaining the Chinese herbal medicine fermentation composition.

8. Use of the Chinese herbal medicine fermentation composition described in claims 1-6 or the Chinese herbal medicine fermentation composition prepared by the preparation method described in claim 7 in the preparation of a product for improving sleep and having the effects of treating lung cancer and lung nodules.

9. The application according to claim 8, wherein This Chinese herbal medicine fermentation composition is used for the preparation of Xuezhi Yu paste.

10. The application according to claim 9, characterized in that The preparation method of Xuezhi Yu paste is as follows: Add excipients to this Chinese herbal medicine fermentation composition, and the addition amount of the excipients is 20% of the total mass of Xuezhi Yu paste; among them: tortoise shell glue accounts for 3%, turtle shell glue accounts for 3%, donkey-hide gelatin accounts for 5%, xylitol accounts for 3%, xanthan gum accounts for 1%, and isomaltooligosaccharide accounts for 5%. Gradually boil under heating conditions until a paste is formed, thereby obtaining Xuezhi Yu paste.

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