Compound traditional Chinese medicine preparation fermentation liquor as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of biological medicine, and particularly relates to a composite traditional Chinese medicine preparation fermentation broth and a preparation method and application thereof.The preparation method of the composite traditional Chinese medicine preparation fermentation broth comprises the following steps that astragalus membranaceus powder, angelica sinensis powder, red ginseng powder and liquorice powder are mixed and then dissolved in water, cellulase and pectinase are added to destroy cell walls, then enzyme deactivation is conducted, and fermentation broth is obtained; sterilizing to obtain a compound traditional Chinese medicine preparation fermentation culture medium; the method comprises the following steps: mixing a seed solution of lactobacillus plantarum GMNL-662 and a seed solution of lactobacillus plantarum R1012 to obtain a compound strain seed solution; inoculating the compound strain seed solution into a compound traditional Chinese medicine preparation fermentation culture medium for fermentation culture to obtain a fermentation solution; crushing the fermentation liquor, collecting supernate, extruding and filtering the supernate, and collecting filtrate, so as to obtain the composite traditional Chinese medicine preparation fermentation liquor. According to the method, the content of crude polysaccharide is increased by adopting compound strain fermentation and combining with a mechanical extrusion mode. According to the method disclosed by the invention, the concentration of the crude polysaccharide is increased from 3.4 mg / mL to 5.1 mg / mL.

Description

Technical Field

[0001] The invention belongs to the technical field of biomedicine, and specifically relates to a compound Chinese medicine preparation fermentation liquid and a preparation method and application thereof. Background Art

[0002] Astragalus, with a sweet taste and slightly warm nature, has the effects of replenishing qi and raising yang, consolidating the exterior and stopping sweating, promoting diuresis and reducing swelling, and detoxifying and promoting muscle regeneration. In clinical practice, Astragalus is often used in combination with other Chinese medicines, such as Atractylodes macrocephala, Salvia miltiorrhiza, Angelica sinensis, Stephania tetrandra, Cinnamomum cassia, Cornus officinalis, Leonurus japonicus, Bletilla striata, etc., which enhances the clinical efficacy of Astragalus and expands the scope of use of Astragalus. The combination of Astragalus is often recorded in ancient medical books. The Synopsis of the Golden Chamber records the Huangqi Jianzhong Decoction, which is mainly used to treat Qi deficiency and internal cold syndrome, and is composed of Astragalus, jujube, white peony and other Chinese medicines; The Neiwaishang Bianhuo Lun records the Danggui Buxue Decoction composed of Astragalus and Angelica sinensis, which can be used to replenish qi and produce blood; the Buzhong Yiqi Decoction composed of Atractylodes macrocephala, ginseng and Astragalus has the effect of replenishing the middle and raising yang; the Guipi Decoction in the Jisheng Fang is composed of ginseng, Astragalus, licorice and Atractylodes macrocephala, which can achieve the effect of producing blood by replenishing the spleen and replenishing qi. In terms of effective ingredients, astragalus contains astragalus polysaccharides, astragalus flavonoids, a variety of trace elements and essential amino acids for the human body, and is often used for the prevention and treatment of immune diseases, blood diseases, endocrine diseases, cardiovascular diseases, etc. Astragalus polysaccharides are often used in combination with Lycium barbarum polysaccharides, Angelica sinensis polysaccharides and Codonopsis pilosula polysaccharides, etc., and have the health care function of enhancing immunity; Astragalus polysaccharides are used in combination with Poria cocos polysaccharides, puerarin, bitter melon saponins and yam to achieve the effect of regulating blood sugar; Astragalus flavonoids are used in combination with Rhodiola rosea extract, Ganoderma lucidum triterpenoids, Epimedium total flavonoids and Hawthorn flavonoids, and are often used in functional products for antioxidants and anti-aging; Rehmannia glutinosa polysaccharides, jujube polysaccharides, and proteins and amino acids in donkey-hide gelatin are also combined with astragalus polysaccharides to develop related products; In addition, astragalus polysaccharides are also widely developed and applied in radiation protection, anti-tumor, liver protection and other aspects.

[0003] As the active ingredient in traditional Chinese medicine, crude polysaccharides have broad application prospects in the fields of medicine and health care products. However, the current preparation of crude polysaccharides faces the problem of low yield, which seriously restricts the large-scale production and application of crude polysaccharides.

[0004] In the existing microbial fermentation technology, the efficiency of microorganisms in utilizing Chinese medicine raw materials is generally low. On the one hand, the complex chemical components in Chinese medicine may inhibit the growth and metabolic activity of microorganisms, making it impossible for microorganisms to fully exert their ability to convert crude polysaccharides. On the other hand, the growth characteristics and metabolic regulation mechanisms of microorganisms themselves also limit the yield of crude polysaccharides.

[0005] Due to the low yield of crude polysaccharides prepared by microbial fermentation, the production cost remains high, making crude polysaccharide-related products expensive and difficult to meet market demand. This not only hinders the widespread application of crude polysaccharides in the medical field, but also limits its large-scale promotion in the health care product industry. Therefore, it is urgent to develop a method that can significantly improve the yield of crude polysaccharides prepared by microbial fermentation. Summary of the invention

[0006] In order to solve the problem of low yield of crude polysaccharide prepared by microbial fermentation, the present invention provides a method for preparing a composite Chinese medicine preparation fermentation broth.

[0007] The technical solution adopted by the present invention is: The present invention provides a method for preparing a composite Chinese medicine preparation fermentation broth, comprising the following steps: Astragalus powder, angelica powder, red ginseng powder and liquorice powder are mixed and dissolved in water, cellulase and pectinase are added to destroy cell walls, then the enzymes are inactivated and sterilized to obtain a composite Chinese medicine preparation fermentation medium; seed liquid of Lactobacillus plantarum GMNL-662 is prepared; seed liquid of Lactobacillus plantarum R1012 is prepared; the seed liquid of Lactobacillus plantarum GMNL-662 and the seed liquid of Lactobacillus plantarum R1012 are mixed to obtain a composite strain seed liquid; the composite strain seed liquid is inoculated into a composite Chinese medicine preparation fermentation medium for fermentation and culture to obtain a fermentation liquid; the fermentation liquid is crushed to collect a supernatant, the supernatant is squeezed and filtered, and the filtrate is collected to obtain the composite Chinese medicine preparation fermentation liquid.

[0008] Preferably, the mass ratio of the astragalus powder, angelica powder, red ginseng powder and licorice powder is 5:1:2:2.

[0009] Preferably, the pore size of the filter membrane used in the extrusion filtration is 0.22 μm.

[0010] Preferably, the volume ratio of the seed solution of Lactobacillus plantarum GMNL-662 to the seed solution of Lactobacillus plantarum R1012 is 1:1, and the inoculation amount of the composite strain seed solution is 4% by volume.

[0011] Preferably, the number of viable Lactobacillus plantarum GMNL-662 in the seed solution of Lactobacillus plantarum GMNL-662 is 1×10 8 CFU / mL; the number of viable Lactobacillus plantarum R1012 in the seed solution of Lactobacillus plantarum R1012 is 1×10 8 CFU / mL.

[0012] Preferably, the fermentation culture condition is static culture at 37° C. for 48 hours.

[0013] The present invention also provides a composite Chinese medicine preparation fermentation liquid obtained according to the preparation method.

[0014] The present invention also provides an application of the compound Chinese medicine preparation fermentation broth, wherein the compound Chinese medicine preparation fermentation broth is used for preparing an antioxidant.

[0015] Preferably, the antioxidant further comprises a pharmaceutically acceptable excipient.

[0016] Preferably, the pharmaceutically acceptable excipient is ethanol.

[0017] Compared with the prior art, the present invention has the following beneficial effects: The present invention provides a method for preparing a composite Chinese medicine preparation fermentation broth, comprising the following steps: After mixing astragalus powder, angelica powder, red ginseng powder and liquorice powder, dissolve in water, add cellulase and pectinase to destroy cell wall, then kill enzyme, sterilize, obtain composite Chinese medicine preparation fermentation medium; prepare seed liquid of plant lactobacillus GMNL-662; prepare seed liquid of plant lactobacillus R1012; mix seed liquid of plant lactobacillus GMNL-662 and seed liquid of plant lactobacillus R1012 to obtain composite strain seed liquid; inoculate composite strain seed liquid into composite Chinese medicine preparation fermentation medium for fermentation and culture to obtain fermentation liquid; collect supernatant after fermentation liquid is crushed, and squeeze and filter supernatant, collect filtrate, and obtain composite Chinese medicine preparation fermentation liquid. The present invention adopts composite strain fermentation, and combines the content of crude polysaccharide in the way of mechanical extrusion. By the method described in the present invention, the concentration of crude polysaccharide is increased from 3.4mg / mL to 5.1mg / mL.

[0018] As probiotics, Lactobacillus plantarum GMNL-662 and Lactobacillus plantarum R1012 can spontaneously generate bacterial exovesicles during the fermentation process. The present invention collects the supernatant of the fermentation liquid co-fermented by Lactobacillus plantarum GMNL-662 and Lactobacillus plantarum R1012, and prepares the composite Chinese medicine preparation fermentation liquid containing bacterial exovesicles by mechanical extrusion. The bacterial exovesicles can wrap the crude polysaccharide in the fermentation liquid, improve the absorption and utilization of the crude polysaccharide, and enhance its anti-inflammatory effect.

[0019] The present invention uses astragalus, angelica, red ginseng and licorice as substrates for cultivation. On the one hand, probiotics can produce bacterial exovesicles during the growth process, and at the same time, the growth metabolism of probiotics can transform the polysaccharides in astragalus, angelica, red ginseng and licorice to produce secondary metabolites with high physiological activity such as astragalus polysaccharides, angelica polysaccharides, and saponins. During the mechanical extrusion process, the bacterial exovesicles can enrich and encapsulate the crude polysaccharides, which can be applied in the field of food, beverage and cosmetics functional raw materials. This method simplifies the process of engineering bacterial exovesicles to encapsulate crude polysaccharides, and can effectively improve the problem of absorption and utilization.

[0020] Bacterial extracellular vesicles act as carriers to encapsulate crude polysaccharides, which can effectively improve their antioxidant activity. In addition, after being taken by humans, they can be precisely targeted and absorbed by small intestinal epithelial cells, thereby improving the bioavailability of crude polysaccharides.

[0021] The present invention combines probiotic fermentation technology with bacterial exosome preparation technology, effectively solving the problem that probiotics only function as fermentation agents. In the present invention, the bacterial exosome encapsulates crude polysaccharides to form a nanoemulsion system, which can significantly improve the bioavailability of crude polysaccharides. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 These are the characterization results of bacterial extracellular vesicles. A and B are bacterial extracellular vesicles under different fields of view.

[0023] Figure 2 This is the particle size distribution of bacterial extracellular vesicles.

[0024] Figure 3 The concentration of crude polysaccharide in the fermentation broth of the composite Chinese medicine preparation prepared in Example 1 and Comparative Example 1.

[0025] Figure 4 To simulate the characterization results of bacterial extracellular vesicle structure after in vitro digestion, A is the transmission electron microscopy photograph of the blank control group; B is the transmission electron microscopy photograph after simulated digestion with artificial saliva, artificial gastric juice and artificial intestinal juice. DETAILED DESCRIPTION

[0026] The present invention is further described below by specific examples, but the scope of the present invention is not limited thereto. The details and forms of the technical solution of the present invention may be modified or replaced without departing from the spirit and scope of the present invention, but these modifications or replacements all fall within the protection scope of the present invention.

[0027] The inventive concept of the present invention is as follows: Due to the low yield of crude polysaccharides prepared by microbial fermentation, the production cost remains high, making crude polysaccharide-related products expensive and difficult to meet market demand. This not only hinders the widespread application of crude polysaccharides in the medical field, but also limits its large-scale promotion in the health care product industry.

[0028] Based on this, the present invention provides a method for preparing a composite Chinese medicine preparation fermentation broth, comprising the following steps: Astragalus powder, angelica powder, red ginseng powder and liquorice powder are mixed and dissolved in water, cellulase and pectinase are added to destroy cell walls, then the enzymes are inactivated and sterilized to obtain a composite Chinese medicine preparation fermentation medium; seed liquid of Lactobacillus plantarum GMNL-662 is prepared; seed liquid of Lactobacillus plantarum R1012 is prepared; the seed liquid of Lactobacillus plantarum GMNL-662 and the seed liquid of Lactobacillus plantarum R1012 are mixed to obtain a composite strain seed liquid; the composite strain seed liquid is inoculated into a composite Chinese medicine preparation fermentation medium for fermentation and culture to obtain a fermentation liquid; the fermentation liquid is crushed to collect a supernatant, the supernatant is squeezed and filtered, and the filtrate is collected to obtain the composite Chinese medicine preparation fermentation liquid.

[0029] As probiotics, Lactobacillus plantarum GMNL-662 and Lactobacillus plantarum R1012 can spontaneously produce BEVs, which are spherical structures with lipid bilayers and are generally 20nm~400nm in diameter. As small molecule active substances derived from bacteria, BEVs have the advantages of stable structure and long effective circulation time. They can encapsulate a variety of bioactive molecules, including proteins, lipids, nucleic acids and metabolites. A large number of studies have shown that BEVs have great potential in immune response and antioxidant effects, and have great application potential as new postbiotics in functional foods, disease treatment substitutes and immunomodulatory adjuvants. In addition, BEVs are also ideal carriers for nano-delivery systems of bioactive compounds because they have good immunological and pharmacokinetic properties and the ability to penetrate physiological barriers. Therefore, high-value functional ingredients can be loaded through technical means to prepare delivery systems. After taking related products in the human body, precise targeted nutritional intervention effects can be achieved, and they can be used as an important carrier for constructing food nutritional factor delivery systems.

[0030] In order to enable those skilled in the art to better understand the technical solution of the present invention and implement it, the present invention is further described below in conjunction with specific embodiments. In the description of the present invention, unless otherwise specified, the reagents used are all commercially available, and the methods used are all conventional techniques in the art.

[0031] The abbreviations of the present invention are shown in Table 1.

[0032] Table 1 Abbreviations The strains and manufacturer information used in the present invention are shown in Table 2.

[0033] Table 2 Strains and manufacturer information The cellulase and pectinase used in the present invention were purchased from Novozymes, the enzymatic activity of the cellulase was 700U / mL, and the enzymatic activity of the pectinase was 10000U / mL; red ginseng powder was purchased from Jilin Hongjiu Biotechnology Co., Ltd.; astragalus powder, angelica powder and licorice powder were all purchased from Bozhou Qingyan Pharmaceutical Co., Ltd.; artificial saliva, artificial gastric juice and artificial intestinal juice were purchased from Shanghai Xinfan Biotechnology Co., Ltd.

[0034] In the present invention, the detection method of crude polysaccharide is as follows: Place 2 μL of the fermentation liquid of the composite Chinese medicine preparation in a 20 mL stoppered test tube, add distilled water to 1 mL, add 1 mL of 5% phenol solution, and then quickly add 5 mL of 98% concentrated sulfuric acid. Use a vortex oscillator to fully mix the reaction solution, let it stand for 10 minutes, and then place it in a 30°C water bath to react for 20 minutes, and measure the absorbance at 490 nm.

[0035] Example 1 A method for preparing a composite Chinese medicine preparation fermentation broth is as follows: S1. Dissolve astragalus powder, angelica powder, red ginseng powder and liquorice powder in water, add cellulase and pectinase to destroy cell walls, then inactivate the enzymes and sterilize to obtain a fermentation medium for a composite Chinese medicine preparation.

[0036] The composite Chinese medicine powder was obtained by mixing astragalus powder, angelica powder, red ginseng powder and licorice powder in a mass ratio of 5:1:2:2, and the composite Chinese medicine powder was mixed with distilled water in a mass ratio of 1:12, and cellulase and pectinase were added; the enzymatic hydrolysis was performed at a speed of 100 rpm / min for 3 hours; the enzyme was inactivated at 85°C for 10 minutes, and centrifuged at 4200 rpm / min for 15 minutes, and the supernatant was collected and sterilized at 121°C for 20 minutes to obtain the composite Chinese medicine fermentation medium. Among them, the amount of cellulase and pectinase added was: 0.05 mL of cellulase and 0.05 mL of pectinase were added to every 1 g of the composite Chinese medicine powder.

[0037] S2. Prepare seed liquid of Lactobacillus plantarum GMNL-662 and seed liquid of Lactobacillus plantarum R1012 respectively, mix the seed liquid of Lactobacillus plantarum GMNL-662 and the seed liquid of Lactobacillus plantarum R1012 to obtain composite strain seed liquid.

[0038] Lactobacillus plantarum GMNL-662 was dissolved in sterile water to obtain a bacterial solution of Lactobacillus plantarum GMNL-662. The bacterial solution of Lactobacillus plantarum GMNL-662 was gently streaked on an MRS solid medium using an inoculation loop, and cultured at 37°C until a single colony of Lactobacillus plantarum GMNL-662 appeared. A single colony of Lactobacillus plantarum GMNL-662 was picked up into a sterilized test tube containing 5 mL of MRS liquid medium using an inoculation loop, and the strain was activated and cultured at 37°C for 18 hours to obtain a seed solution of Lactobacillus plantarum GMNL-662. The number of viable bacteria of Lactobacillus plantarum GMNL-662 in the seed solution of Lactobacillus plantarum GMNL-662 was 1×10 8 CFU / mL.

[0039] Lactobacillus plantarum R1012 was dissolved in sterile water to obtain a bacterial solution of Lactobacillus plantarum R1012. The bacterial solution of Lactobacillus plantarum R1012 was gently streaked on an MRS solid medium using an inoculation loop, and cultured at 37°C until a single colony of Lactobacillus plantarum R1012 appeared. A single colony of Lactobacillus plantarum R1012 was picked up into a sterilized test tube containing 5 mL of MRS liquid medium using an inoculation loop, and the strain was activated. The culture was statically cultured at 37°C for 18 hours to obtain a seed solution of Lactobacillus plantarum R1012, wherein the number of viable Lactobacillus plantarum R1012 in the seed solution of Lactobacillus plantarum R1012 was 1×10 8 CFU / mL.

[0040] The seed liquid of Lactobacillus plantarum GMNL-662 and the seed liquid of Lactobacillus plantarum R1012 were mixed in a volume ratio of 1:1 to obtain a composite strain seed liquid.

[0041] S3. Inoculate the composite strain seed liquid into the composite Chinese medicine preparation fermentation medium for fermentation and culture to obtain a fermentation liquid.

[0042] The composite strain seed liquid was inoculated into the composite Chinese medicine preparation fermentation medium at an inoculum volume fraction of 4%, and cultured statically in a 37°C incubator for 48 hours. After the fermentation was completed, the fermentation liquid was collected.

[0043] S4, crushing the fermentation liquid, collecting the supernatant, squeezing and filtering the supernatant, and collecting the filtrate to obtain the composite Chinese medicine preparation fermentation liquid.

[0044] After the fermentation broth was ultrasonically disrupted, it was centrifuged at 4000 rpm / min for 10 min at 4°C to remove cell debris, and the supernatant was retained. The supernatant was manually squeezed and filtered using a needle filter to pass through a 0.22 μm filter membrane, and the filtrate was collected to finally obtain the composite Chinese medicine preparation fermentation broth.

[0045] Comparative Example 1 A method for preparing a composite Chinese medicine preparation fermentation broth is as follows: S1-S3 are exactly the same as those in Example 1, and the supernatant in S4 is the fermentation broth of the composite Chinese medicine preparation.

[0046] The fermentation broth of the composite Chinese medicine preparation prepared in Example 1 was characterized as follows: The bacterial extracellular vesicles in the fermentation broth of the composite Chinese medicine preparation prepared in Example 1 were observed, and the bacterial extracellular vesicles with obvious Tyndall effect were observed by transmission electron microscopy, and the laser particle size distribution and protein concentration were measured.

[0047] Transmission electron microscopy results Figure 1 The particle size analysis results are shown in Figure 2 As shown. Figure 1 It can be seen that bacterial extracellular vesicles are round structures. Figure 2 It can be seen that the average particle size of bacterial extracellular vesicles is 149.8nm. In addition, the vesicle concentration and protein concentration of bacterial extracellular vesicles were also measured. The test results showed that the concentration of bacterial extracellular vesicles was 8.1×10 9 Particles / mL, and the protein concentration was 10.77 mg / mL.

[0048] The crude polysaccharides in the fermentation broth of the composite Chinese medicine preparations prepared in Example 1 and Comparative Example 1 were determined as follows: Depend on Figure 3 It can be seen that the crude polysaccharide concentration in the fermentation broth of the composite Chinese medicine preparation prepared in Comparative Example 1 is 3.4 mg / mL; while the crude polysaccharide concentration in the fermentation broth of the composite Chinese medicine preparation prepared in Example 1 is increased to 5.1 mg / mL.

[0049] In addition, an equal volume of 20% SDS was added to the fermentation broth of the composite Chinese medicine preparation prepared in Example 1 for demulsification. After demulsification, the crude polysaccharide concentration reached 7.5 mg / mL, indicating that the bacterial extracellular vesicles can effectively encapsulate the crude polysaccharide.

[0050] Example 2 An application of a compound Chinese medicine preparation fermentation liquid is as follows: 1. Determine the DPPH free radical scavenging ability.

[0051] The fermentation broth of the composite Chinese medicine preparation prepared in Example 1, the fermentation medium of the composite Chinese medicine preparation prepared in S1 in Example 1, and the fermentation broth prepared in S3 in Example 1 were tested.

[0052] The DPPH free radical scavenging ability detection kit was purchased from Beijing Solebow Technology Co., Ltd. according to the instructions of the DPPH free radical scavenging ability detection kit.

[0053] The results showed that the DPPH radical scavenging rates of the fermentation broth, the composite Chinese medicine preparation fermentation medium and the composite Chinese medicine preparation fermentation broth were 15.34%, 13.28% and 35.89%, respectively, indicating that the antioxidant activity was improved after fermentation. Therefore, the composite Chinese medicine preparation fermentation broth of the present invention can be prepared into an antioxidant by adding ethanol.

[0054] 2. Simulate in vitro digestion.

[0055] The fermentation liquid of the composite Chinese medicine preparation prepared in Example 1 was reacted in artificial saliva, artificial gastric juice and artificial intestinal juice in turn to simulate the digestion process, the method is as follows: a. Take 500 μL of the fermentation liquid of the compound Chinese medicine preparation, add 500 μL of artificial saliva kept in a 37°C water bath, mix thoroughly, and place in a 37°C water bath for 5 minutes.

[0056] b. Add 1000 μL of artificial gastric juice kept warm in a 37°C water bath to the fermentation broth of the composite Chinese medicine preparation after digestion with artificial saliva, mix thoroughly, and place in a 37°C water bath for 2 hours.

[0057] c. Continue to add 2000 μL of artificial intestinal fluid kept in a 37°C water bath, mix thoroughly, and place in a 37°C water bath for 1 hour.

[0058] d. After the previous step, place in a 90℃ water bath for 5 min.

[0059] e. Centrifuge at 8000 rpm for 10 min, collect the supernatant and perform detection.

[0060] Depend on Figure 4 It can be seen that after the fermentation broth of the compound Chinese medicine preparation was digested by artificial saliva, artificial gastric juice and artificial intestinal juice in turn, the complete vesicle structure was still retained, indicating that it can resist digestion by gastrointestinal juice. The bacterial extracellular vesicle structure in the fermentation broth of the compound Chinese medicine preparation is stable and has a good ability to retain its contents.

[0061] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0062] The above-mentioned embodiments only express several implementation methods of the present invention, and the description is relatively specific and detailed, but it cannot be understood as limiting the scope of the invention patent. It should be pointed out that for ordinary technicians in this field, several modifications and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention.

Claims

1. A method for preparing a composite Chinese medicine preparation fermentation broth, characterized in that: The following steps are involved: Mixing astragalus powder, angelica powder, red ginseng powder and liquorice powder and dissolving them in water, adding cellulase and pectinase to destroy cell walls, then inactivating the enzymes and sterilizing to obtain a composite Chinese medicine preparation fermentation medium; preparing a seed solution of Lactobacillus plantarum GMNL-662; preparing a seed solution of Lactobacillus plantarum R1012; Mixing the seed liquid of Lactobacillus plantarum GMNL-662 and the seed liquid of Lactobacillus plantarum R1012 to obtain a composite strain seed liquid; Inoculating the composite strain seed liquid into the composite traditional Chinese medicine preparation fermentation medium for fermentation and culturing to obtain a fermentation liquid; The fermentation liquid is crushed and the supernatant is collected. The supernatant is squeezed and filtered, and the filtrate is collected to obtain the composite Chinese medicine preparation fermentation liquid.

2. The preparation method according to claim 1, characterized in that The mass ratio of the astragalus powder, angelica powder, red ginseng powder and liquorice powder is 5:1:2:

2.

3. The preparation method according to claim 1, characterized in that: The pore size of the filter membrane used in the extrusion filtration is 0.22 μm.

4. The preparation method according to claim 1, characterized in that: The volume ratio of the seed liquid of Lactobacillus plantarum GMNL-662 to the seed liquid of Lactobacillus plantarum R1012 is 1:1, and the inoculation amount of the composite strain seed liquid is 4% by volume.

5. The preparation method according to claim 1, characterized in that: The number of viable bacteria of Lactobacillus plantarum GMNL-662 in the seed liquid of Lactobacillus plantarum GMNL-662 is 1×10 8 CFU / mL; The number of viable bacteria of Lactobacillus plantarum R1012 in the seed liquid of Lactobacillus plantarum R1012 is 1×10 8 CFU / mL.

6. The preparation method according to claim 1, characterized in that: The fermentation culture conditions are static culture at 37° C. for 48 hours.

7. A composite Chinese medicine preparation fermentation broth obtained according to the preparation method of claim 1.

8. The use of the fermentation liquid of the composite Chinese medicine preparation as claimed in claim 7, characterized in that: The composite Chinese medicine preparation fermentation liquid is used for preparing antioxidants.

9. The use according to claim 8, characterized in that The antioxidant also includes pharmaceutically acceptable excipients.

10. The use according to claim 9, characterized in that The pharmaceutically acceptable excipient is ethanol.

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