CN111358814A - Ganoderma lucidum fruiting body composite probiotic preparation for enhancing organism immunity - Google Patents
Ganoderma lucidum fruiting body composite probiotic preparation for enhancing organism immunity Download PDFInfo
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- CN111358814A CN111358814A CN202010191175.0A CN202010191175A CN111358814A CN 111358814 A CN111358814 A CN 111358814A CN 202010191175 A CN202010191175 A CN 202010191175A CN 111358814 A CN111358814 A CN 111358814A
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- ganoderma lucidum
- fruiting body
- ganoderma
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/06—Fungi, e.g. yeasts
- A61K36/07—Basidiomycota, e.g. Cryptococcus
- A61K36/074—Ganoderma
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
- A61P37/04—Immunostimulants
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/10—Preparation or pretreatment of starting material
- A61K2236/19—Preparation or pretreatment of starting material involving fermentation using yeast, bacteria or both; enzymatic treatment
Abstract
The invention relates to the field of fermentation engineering, in particular to a ganoderma lucidum fruiting body composite probiotic preparation for enhancing organism immunity. The fermented product is obtained by co-fermenting the lucid ganoderma sporocarp extracting solution and the composite probiotic strain, wherein the composite probiotic is lactobacillus acidophilus and bifidobacterium breve, and the number ratio of the two viable bacteria is 1: 1. preparing Ganoderma encarpium powder into Ganoderma extractive solution with concentration of 8.8-33.2mg/ml, and adding into MRS culture medium containing 0.5% sugar. Then lactobacillus acidophilus and bifidobacterium breve are respectively inoculated into the culture medium in the concentration of 1 percent, and are cultured for 48 hours at the temperature of 37 ℃ for fermentation to obtain a fermentation product, which can effectively improve the phenomenon of low immunity and enhance the immunity of organisms. Meanwhile, the problem of low utilization rate of rare Chinese herbal medicine, namely ganoderma lucidum, can be improved.
Description
Technical Field
The invention relates to the field of fermentation engineering, in particular to a ganoderma lucidum fruiting body composite probiotic preparation for enhancing organism immunity.
Background
Ganoderma (scientific name: Ganoderma Lucidum Karst) has umbrella shape, pileus kidney shape, semicircular shape or nearly circular shape, and is fruiting body of Ganoderma Lucidum of Polyporaceae. Ganoderma has different curative effects on neurasthenia, hyperlipidemia, coronary heart disease, angina pectoris, arrhythmia, altitude sickness, hepatitis, hemorrhagic fever, dyspepsia, and tracheitis.
The Fruiting body is the spore-forming structure of higher fungi, i.e. fruit body, which is composed of organized mycelium, and is the main edible and medicinal part of edible fungi and medicinal fungi. The ganoderma lucidum fruiting body powder refers to powder of a ganoderma lucidum fruiting body which is crushed by an ultrafine crushing technology, is actually in a powdery form of the ganoderma lucidum fruiting body, and is easier to digest, absorb and utilize compared with the ganoderma lucidum fruiting body.
Probiotics is a kind of active microorganisms beneficial to a host, and is a general term for active beneficial microorganisms which are planted in the intestinal tract and the reproductive system of a human body and can generate definite health efficacy so as to improve the micro-ecological balance of the host and play beneficial roles. Researches prove that the probiotics can carry out biotransformation on the effective components of the traditional Chinese medicine, and convert macromolecular substances of the traditional Chinese medicine into micromolecular substances which can be directly absorbed by intestinal tracts of human bodies through microorganisms.
How to reasonably utilize the ganoderma lucidum sporocarp and the probiotics for co-fermentation to prepare the ganoderma lucidum sporocarp composite probiotic preparation for enhancing the immunity of the organism is a subject worthy of deep research, and no relevant literature report is found at present.
Disclosure of Invention
Aiming at the technical problems in the prior art, the invention aims to provide a ganoderma lucidum fruiting body composite probiotic preparation for enhancing the immunity of the organism, which can effectively improve the phenomenon of low immunity and enhance the immunity of the organism.
In order to achieve the purpose, the technical scheme adopted by the invention is as follows: a Ganoderma fruiting body compound probiotic preparation for enhancing immunity is prepared by fermenting Ganoderma fruiting body extractive solution and compound probiotic strain together to obtain fermented product, i.e. Ganoderma fruiting body compound probiotic preparation.
Preferably, the compound probiotics in the ganoderma lucidum fruiting body compound probiotic preparation for enhancing the immunity of the organism are lactobacillus acidophilus and bifidobacterium breve. Further preferably, the ratio of the number of the live bacteria of the lactobacillus acidophilus and the bifidobacterium breve in the composite probiotics is 1: 1.
the preparation method of the ganoderma lucidum fruiting body composite probiotic preparation for enhancing the immunity of organisms comprises the following steps:
1) preparing an MRS culture medium containing a ganoderma lucidum extracting solution:
preparing Ganoderma lucidum fruiting body powder into Ganoderma lucidum extractive solution, adding into MRS culture medium, and performing high pressure treatment to obtain MRS culture medium containing Ganoderma lucidum extractive solution;
2) inoculating and fermenting
Respectively inoculating Lactobacillus acidophilus and Bifidobacterium breve into the MRS culture medium containing Ganoderma extractive solution, and fermenting to obtain Ganoderma fruiting body compound probiotic preparation.
As a preferred technical scheme of the invention, the preparation method comprises the following steps:
preferably, the Ganoderma encarpium powder is prepared into Ganoderma extractive solution with concentration of 8.8-33.2mg/ml, and then added into MRS culture medium containing 0.5% sugar. Further preferably, the Ganoderma encarpium powder is first prepared into Ganoderma extract with concentration of 33.2mg/ml, and then added into MRS culture medium containing 0.5% sugar.
Preferably, in the step 2), lactobacillus acidophilus and bifidobacterium breve strains are respectively inoculated into the MRS culture medium containing the ganoderma lucidum extract at the concentration of 1 percent.
Preferably, the fermentation is carried out by culturing at 37 ℃ for 48h after the inoculation in the step 2).
Compared with the prior art, the invention has the beneficial effects that:
the lucid ganoderma sporocarp compound probiotic preparation disclosed by the invention is a fermented product obtained by co-fermenting lucid ganoderma sporocarp extracting solution and compound probiotic strains, and can effectively improve the phenomenon of low immunity and enhance the immunity of an organism. Meanwhile, the problem of low utilization rate of rare Chinese herbal medicine, namely ganoderma lucidum, can be improved.
Drawings
Fig. 1 is a graph of probiotic growth.
Figure 2 is a graph of total sugar content.
FIG. 3 is a graph comparing the components before and after fermentation.
FIG. 4 is a graph of the change in body weight of mice.
FIG. 5 is a colon HE stained pathological section.
FIG. 6 is a graph showing the results of blood small intestine tissue homogenate detection.
FIG. 7 is a PCR-denaturing gradient gel electrophoresis profile.
Detailed Description
The present invention will be described in further detail with reference to examples, but the present invention is not limited thereto.
Example 1
A preparation method of a ganoderma lucidum fruiting body composite probiotic preparation for enhancing organism immunity comprises the following steps:
1) and strain screening: detecting OD value to screen out strains capable of growing on Ganoderma culture medium, including Lactobacillus acidophilus, Bifidobacterium breve, 9218 Lactobacillus brevis, Lactobacillus casei, Bifidobacterium pseudocatenulatum, and Lactobacillus gasseri.
2) And compounding MRS culture medium containing three kinds of glossy ganoderma extractive liquid of low, medium and high concentration
Preparing Ganoderma encarpium powder with water into Ganoderma extractive solutions of three concentrations of 8.8mg/ml, 16.6mg/ml, and 33.2mg/ml, respectively, adding into MRS culture medium containing 0.5% sugar, and processing under high pressure to obtain MRS culture medium containing Ganoderma extractive solution.
3) Inoculating and fermenting
Inoculating the probiotic strains into MRS culture medium containing Ganoderma extractive solution at a concentration of 1%, and culturing at 37 deg.C for 48 hr to obtain various Ganoderma fruiting body probiotic preparation products.
Inoculating Lactobacillus acidophilus and Bifidobacterium breve into MRS culture medium containing Ganoderma extractive solution at concentration of 1% (viable count ratio of two probiotics is 1: 1), and culturing at 37 deg.C for 48 hr to obtain Ganoderma fruiting body composite probiotic preparation.
Next, the growth of each experimental microorganism group and the determination of total sugar content are observed and applied to the model animal model to observe the application effect of each experimental group to obtain the biological fermentation preparation.
Example 2
And (3) measuring the growth condition and the total sugar content of probiotics in the ganoderma lucidum sporocarp probiotic fermentation liquor: measuring an OD value at the position of 620nm and drawing a growth curve to obtain the growth condition of the probiotics; the decomposition rule of the saccharides in the fermentation liquor is known by adopting a phenol-concentrated sulfuric acid method, and then the proper concentration of the ganoderma lucidum culture medium is determined. And (3) carrying out mass spectrum analysis on the composite probiotic fermented preparation, and detecting whether component change is generated before and after fermentation. The total sugar content determination comprises the following specific steps:
taking glucose as a standard substance, numbering 6 clean test tubes by adopting a phenol-concentrated sulfuric acid method, respectively sucking 0.1mg/ml standard glucose solution 0, 0.2, 0.4, 0.6, 0.8 and 1.0ml into the test tubes, then adding distilled water to make up to 1ml, respectively adding 1ml of 6% phenol and 5ml of concentrated sulfuric acid into each test tube, fully mixing uniformly and standing for 10 min. Reacting in a water bath at 30 deg.C for 20min, taking appropriate amount of reaction solution, measuring absorbance at 490nm, and drawing standard curve with glucose concentration (ug/ml) as abscissa and absorbance as ordinate. Taking 2ml of fermentation liquor, adding 5ml of water and 1.5ml of concentrated hydrochloric acid, carrying out water bath at 100 ℃ for 3 hours, and metering the volume of distilled water to 25 ml. Sucking 0.1ml of the above liquid into test tubes, adding 0.9ml of distilled water, adding 1.0ml of phenol reagent and 5.0ml of concentrated sulfuric acid into each test tube, mixing the reaction solution, standing for 10min, and reacting in a 30 ℃ water bath for 20 min. Taking a proper amount of reaction liquid to detect the absorbance at 490nm, and calculating the total sugar concentration (mg/ml) in the fermentation liquid according to a formula. Thereby knowing the decomposition rule of the saccharides in the fermentation liquid and detecting whether the components change before and after the fermentation.
As shown in FIGS. 1-3, it can be seen that Lactobacillus acidophilus and Bifidobacterium breve can grow from fruiting body of Ganoderma lucidum (shown in FIG. 1), and the concentration of Lactobacillus acidophilus and Bifidobacterium breve is 33.2mg/ml, the conditions are suitable, and the total sugar content is high (shown in FIG. 2). Through mass spectrometry detection, the component changes of lactobacillus acidophilus and bifidobacterium breve before and after fermentation are obvious (shown in figure 3), and the response degree of ganoderic acid C2 in fermentation liquor is larger than that in a ganoderma lucidum fruiting body culture medium (shown in table 1), so that a preparation obtained by combined fermentation of lactobacillus acidophilus and bifidobacterium breve is adopted for subsequent animal experiments.
TABLE 1 comparison of Pre-fermentation (A) and post-fermentation (B) Components and amounts
Example 3
Application of Ganoderma encarpium-Lactobacillus acidophilus and Bifidobacterium breve compound fermentation broth in animal model with low immunity is provided.
Mainly observing the change condition of animal body weight and the condition of feces, detecting colon tissue HE staining pathology, detecting IL-17 and TNF- α by an ELISA double-antibody sandwich method, and detecting the structure of mouse intestinal flora by a PCR-denaturing gradient gel electrophoresis method.
The animal model is constructed by using 6-8 week-old male Balb/c mice, and is divided into 4 groups (8 mice in each group), namely a Control (CON) group, a dexamethasone model group (DEX) group, a fermentation liquor (FT) group and a lucid Ganoderma (GL) group, the rest 3 groups of mice except the CON group are subjected to intraperitoneal injection of dexamethasone 20mg/(kg.d) every day for 7 days continuously, a low immunity model is established, the FT group lucid ganoderma fermentation liquor is given by 0.2ml from the 8 th day, the GL group lucid ganoderma liquor is given by 0.2ml, the GL group lucid ganoderma liquor is fed continuously for 14 days, eyeballs are picked on the 21 st day to kill the groups of mice, blood, colon, excrement, spleen and other tissues are taken, a weight change curve of the mice is drawn, colon tissue ELISA pathological staining is performed, the IL-17 and TNF- α levels are detected by a double antibody sandwich method, and the intestinal flora structure of the mice.
The experimental results show that:
1. the Ganoderma encarpium-Lactobacillus acidophilus and Bifidobacterium breve compound fermentation liquid can recover body weight of hypoimmunity mice (shown in figure 4).
2. Colon HE stained pathological section shows: the colon tissue of the CON group mouse is intact, the mucous membrane structure is arranged orderly, and inflammatory cell infiltration is not seen. The DEX group tissues have serious epithelial damage, and mucosa and submucosa are infiltrated by a large amount of inflammatory cells mainly comprising neutrophils. The infiltration of cells into the mucosa and submucosa of the FT and GL groups was reduced and the epithelial lesions were mild (fig. 5).
3. ELISA (competitive assay) detection of small intestine tissue homogenate showed that the DEX group TNF- α was higher than other groups, and inflammatory factors were reduced by treatment with Ganoderma encarpium fermentation broth (FIG. 6).
4. The results of PCR-denaturing gradient gel electrophoresis show that: after the treatment of the ganoderma lucidum fruiting body compound fermentation liquor, FT group bands are increased, and intestinal microbial diversity is increased (shown in figure 7 a). The DGGE map is subjected to digital processing by using Quantity One, then, non-weighted pairwise arithmetic mean algorithm (UPGMA) is adopted to perform clustering analysis (shown in figure 7 b) on the types of the lanes, the CON group is clustered into a cluster, and the similarity of the rest groups is higher, which shows that the structure of the intestinal flora of the mouse subjected to the dexamethasone trunk prognosis is obviously different compared with that of the normal group. The bands with larger changes in PCR-DGGE spectra of 4 groups of mice are subjected to gel cutting sequencing, NCBI (https:// www.ncbi.nlm.nih.gov /) is used for retrieval and homology comparison, sequencing comparison results after PCR-DGGE band gel cutting recovery are shown in Table 2, and the number and diversity of Geobacillus bacteria in the FT group are obviously increased after ganoderma lucidum fruiting body composite probiotic fermentation liquor is dried.
TABLE 2 sequencing comparison results after PCR-DGGE band gel cutting recovery
In summary, the following steps: lactobacillus acidophilus and Bifidobacterium breve can grow well in the culture medium of Ganoderma fruiting body, and the conditions are suitable when the Ganoderma fruiting body concentration is 33.2mg/ml, and the total sugar content is high. The Lactobacillus acidophilus and Bifidobacterium breve can be converted into ganoderic acid C2 with increased fermentation content by using ganoderic acid N, ganoderic acid AM1, ganoderic acid B, ganoderic acid A and ganoderic acid H in Ganoderma fruiting body. The ganoderic acid has effects of enhancing immunity, resisting tumor, protecting liver, removing toxic substance, and resisting HIV virus.
The Ganoderma encarpium-Lactobacillus acidophilus and Bifidobacterium breve compound fermentation broth has functions of recovering animal model weight with low immunity, improving colon tissue morphology, and reducing reaction; the diversity of intestinal flora is increased, and the backwall mycoderm is the dominant bacterium. Researches find that the ratio of bacteroidetes/firmicutes in intestinal tracts of middle-aged men is positively correlated with the levels of serum IgG and IgM, and the serum IgG and IgM are main components of a human humoral immune system, so that the human body can be protected from being attacked by various pathogenic bacteria, and the immunity of the human body is enhanced.
The foregoing is illustrative and explanatory only of the inventive concept, and it is intended that those skilled in the art, upon attaining an understanding of the present invention, will readily produce alterations to, variations of, and equivalents to such embodiments without departing from the spirit and scope of the invention as defined by the appended claims.
Claims (9)
1. A ganoderma lucidum fruiting body compound probiotic preparation for enhancing organism immunity is characterized in that a ganoderma lucidum fruiting body extracting solution and a compound probiotic strain are fermented together to obtain a fermentation product, namely the ganoderma lucidum fruiting body compound probiotic preparation.
2. The ganoderma lucidum fruiting body complex probiotic preparation according to claim 1, wherein the complex probiotic is lactobacillus acidophilus and bifidobacterium breve.
3. The ganoderma lucidum fruiting body composite probiotic preparation according to claim 2, wherein the ratio of the number of viable bacteria of lactobacillus acidophilus and bifidobacterium breve in the composite probiotic is 1: 1.
4. the method for preparing the ganoderma lucidum fruiting body composite probiotic preparation according to claim 2, wherein the steps are as follows:
1) preparing an MRS culture medium containing a ganoderma lucidum extracting solution:
preparing Ganoderma lucidum fruiting body powder into Ganoderma lucidum extractive solution, adding into MRS culture medium, and performing high pressure treatment to obtain MRS culture medium containing Ganoderma lucidum extractive solution;
2) inoculating and fermenting
Respectively inoculating Lactobacillus acidophilus and Bifidobacterium breve into the MRS culture medium containing Ganoderma extractive solution, and fermenting to obtain Ganoderma fruiting body compound probiotic preparation.
5. The method for preparing the ganoderma lucidum fruiting body composite probiotic preparation according to claim 4, wherein firstly, the ganoderma lucidum fruiting body powder is prepared into ganoderma lucidum extract with the concentration of 8.8-33.2mg/ml, and then added into MRS culture medium containing 0.5% sugar.
6. The method for preparing the ganoderma lucidum fruiting body composite probiotic preparation according to claim 5, wherein firstly, the ganoderma lucidum fruiting body powder is prepared into the ganoderma lucidum extracting solution with the concentration of 33.2mg/ml, and then is added into the MRS culture medium containing 0.5% of sugar.
7. The method for preparing the ganoderma lucidum fruiting body composite probiotic preparation according to claim 4, wherein in the step 2), lactobacillus acidophilus and bifidobacterium breve strains are respectively inoculated into the MRS culture medium containing the ganoderma lucidum extracting solution at a concentration of 1%.
8. The method for preparing the ganoderma lucidum fruiting body composite probiotic preparation according to claim 4, wherein the inoculation in step 2) is followed by culturing at 37 ℃ for 48h for fermentation.
9. The use of the ganoderma lucidum fruiting body complex probiotic preparation according to claim 1, 2 or 3 for enhancing immunity.
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Cited By (3)
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| CN112155199A (en) * | 2020-09-29 | 2021-01-01 | 上海怡宝化妆品集团有限公司 | Capsule containing Ganoderma lucidum sporophore spore powder fermentation liquid and preparation method thereof |
| CN113750129A (en) * | 2021-09-17 | 2021-12-07 | 无锡弘焕微生态科技有限公司 | Traditional Chinese medicine probiotic fermentation liquor preparation for relieving side effects of chemotherapy |
| CN113875987A (en) * | 2021-09-17 | 2022-01-04 | 无锡弘焕微生态科技有限公司 | Formula food with anti-aging function |
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| CN110882287A (en) * | 2019-12-31 | 2020-03-17 | 合肥康春堂药业有限责任公司 | Ganoderma lucidum fruiting body microbial fermentation preparation for treating intestinal flora disorder and intestinal barrier function impairment |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112155199A (en) * | 2020-09-29 | 2021-01-01 | 上海怡宝化妆品集团有限公司 | Capsule containing Ganoderma lucidum sporophore spore powder fermentation liquid and preparation method thereof |
| CN112155199B (en) * | 2020-09-29 | 2022-09-23 | 上海洁士宝日化集团有限公司 | Capsule containing Ganoderma lucidum sporophore spore powder fermentation liquid and preparation method thereof |
| CN113750129A (en) * | 2021-09-17 | 2021-12-07 | 无锡弘焕微生态科技有限公司 | Traditional Chinese medicine probiotic fermentation liquor preparation for relieving side effects of chemotherapy |
| CN113875987A (en) * | 2021-09-17 | 2022-01-04 | 无锡弘焕微生态科技有限公司 | Formula food with anti-aging function |
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