CN106309508B - Antrodia camphorata mycelium active substance, preparation method thereof, medical composition containing the same and application thereof - Google Patents
Antrodia camphorata mycelium active substance, preparation method thereof, medical composition containing the same and application thereof Download PDFInfo
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- CN106309508B CN106309508B CN201510506134.5A CN201510506134A CN106309508B CN 106309508 B CN106309508 B CN 106309508B CN 201510506134 A CN201510506134 A CN 201510506134A CN 106309508 B CN106309508 B CN 106309508B
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Abstract
The invention relates to an antrodia camphorata mycelium active substance, a preparation method thereof, a medical composition containing the antrodia camphorata mycelium active substance and application of the antrodia camphorata mycelium active substance. The active substance is prepared by the following steps: (a) culturing Antrodia Camphorata mycelium at predetermined temperature and pH for 5-7 days; (b) inoculating the cultured antrodia cinnamomea mycelia in the step (a) into a fermentation tank, and culturing for 7-20 days at the preset temperature and the preset pH value under stirring; (c) freeze-drying the cultured antrodia cinnamomea mycelia in the step (b), and grinding into powder to form antrodia cinnamomea mycelia freeze-dried powder; mixing a predetermined amount of solvent with the antrodia camphorata mycelium freeze-dried powder, shaking for extraction, and filtering to remove the antrodia camphorata mycelium freeze-dried powder to form antrodia camphorata mycelium extract; and concentrating and drying the antrodia camphorata mycelium extract liquid under reduced pressure to obtain the antrodia camphorata mycelium active substance.
Description
Technical Field
The invention relates to an active substance of antrodia camphorata mycelium, in particular to an active substance of antrodia camphorata mycelium for a medicine for preventing and/or treating xerophthalmia, a preparation method thereof, a medical composition containing the same and application thereof.
Background
Antrodia camphorata pattern
Antrodia camphorata also known as Antrodia camphorata, Balanophora canescens, and Balanophora canescens in caves, and Taiwan is also called Yinyang Paokougu. The fruiting body of Antrodia camphorata is perennial, has strong fragrance of camphor tree, which is different from the common Ganoderma lucidum, and the shape of the fruiting body is plate-shaped or bell-shaped. The surface of the platy antrodia camphorata is orange (yellow), the whole surface of the platy antrodia camphorata is provided with bacteria holes, and the bottom layer of the platy antrodia camphorata is provided with pale yellow white cork which is attached to the inner wall of the hollow core material of the antrodia camphorata to grow. The bell-shaped antrodia camphorata has the fruiting body (clock surface) in orange color, is full of fungus pores (4-5 fungus pores/mm), has extremely bitter taste of spores, is orange red when being fresh, and then becomes orange brown or brown, and the clock body is dark green brown peel. The basidiospores are observed by a microscope and are in a smooth colorless transparent micro-bent column shape.
Production of Antrodia camphorata
Natural antrodia camphorata only grows in the specific hollow trunk of the antrodia camphorata tree in taiwan, the antrodia camphorata tree is listed as a conservation tree species based on the concept of ecological protection, and the antrodia camphorata growing on the antrodia camphorata tree is also limited to be not collected, so that most of the antrodia camphorata sold on the market at present is produced by artificial cultivation. The artificial culture of antrodia camphorata mainly comprises basswood cultivation, space bag cultivation, dish cultivation, liquid state cultivation and the like, wherein the state of antrodia camphorata fruiting bodies cultivated by basswood is closest to that of natural antrodia camphorata, and the method for producing mycelium by separating antrodia camphorata strains from the natural fruiting bodies and performing liquid state fermentation is gradually the mainstream of antrodia camphorata volume production in recent years due to shorter time consumption and easy large-scale cultivation.
Efficacy of Antrodia camphorata
Traditionally, the natural antrodia camphorata handed over from the mouth to the ear has the effects of detoxifying, diminishing inflammation, relieving pain, resisting cancer, protecting the liver and the like, but all the effects are not verified by scientific methods, the antrodia camphorata market is developed vigorously in recent years, the antrodia camphorata becomes a popular research target, published papers reach hundreds of papers, the comprehensive results show that the antrodia camphorata has the effects of resisting inflammation, resisting oxidation, protecting the liver, resisting tumors, enhancing immunity, reducing blood pressure and the like, and the main physiological active ingredients of the antrodia camphorata comprise polysaccharide, triterpenes, superoxide dismutase, adenosine and the like. The artificially cultured antrodia camphorata mycelium can induce and generate the content and the composition of metabolites which are partially different from those of natural antrodia camphorata by adjusting the components of a liquid medium and the culture conditions, so the effect of the antrodia camphorata mycelium is not completely the same as that of a sporocarp, and the part needs to be systematically discussed by scientists. The applicant has already carried out research on the efficacy of the antrodia camphorata for a long time, and the produced antrodia camphorata mycelium capsule product passes two taiwan health food certifications of liver protection and blood pressure regulation (taiwan health food character a 00182), and continuously faces to the development effort of new efficacy ingredients.
Brief introduction to Dry eye
Xerophthalmia (keratocionjunctitis sicca) is a quite common disease (accounting for 15-20% of the adult population), and is mainly caused by lack of tears on the surface of an eyeball or poor quality of the tears, so that patients have uncomfortable symptoms such as eye acerbity, foreign body sensation, burning sensation or increased secretions, temporary blurred vision and the like. There are many possible causes of abnormal tear secretion, including excessive eye use, prolonged contact lens wear, inflammation of the ocular surface, aging, hormone imbalance, autoimmune diseases, and damage to the ocular surface due to some external factors, and Dry eye syndrome (Dry eye syndrome) is also called Dry eye syndrome because many of the factors cause Dry eye symptoms.
Brief introduction to tear film
The tear film is a layer of liquid film covering the cornea and conjunctiva, and has moistening and protecting functions. The structure of the tear film can be simply divided into three layers:
lipid layer (Lipid layer): the outermost layer of the tear film is mainly composed of wax, cholesterol, phospholipid, triglyceride, etc., is secreted by meibomian glands (tarsal glands), Zeiss glands, and Moll glands, and is liquid at normal body temperature to prevent tear evaporation.
(ii) tear water layer (Aqueous layer): the middle layer of the tear film is the thickest layer in the three-layer structure, mainly comprises water, inorganic salt, organic matter, tear protein (tear lipocalin), lactoferrin, lysozyme, lacritin protein and the like, and is secreted by main and auxiliary lacrimal gland cells (lacrimal gland).
(iii) mucus layer (Mucous layer): is the inner layer of the tear film. Covering the corneal epithelial surface, it is composed of mucopolysaccharide (mucin) and the like, and is secreted mainly by conjunctival epithelial goblet cells (junctional goblet cells) and partially by the main lacrimal gland. Mucus is in semisolid state, has high hydrophilicity, and forms a hydrophilic interface between the corneal epithelium layer and the lacrimal water layer of the lacrimal film, thereby playing roles in nourishing, protecting cornea and smoothing corneal surface.
The decrease of tear secretion, mucus secretion, lipid secretion, corneal epithelial roughness, and tear component change caused by any cause may result in the destruction of the physiological properties of the tear film, causing the corneal and conjunctiva to be dry, and further causing the occurrence of dry eye.
Dry eye treatment
The treatment of dry eye mainly aims at searching the cause of the disease and taking medicines according to the symptoms, so as to eliminate the disease for the first time, and if the cause of the disease cannot be eliminated, the symptom is relieved. Simply dry eye symptoms caused by poor daily habits such as excessive eye use and long-time wearing of contact lenses, doctors usually recommend improving the dry eye symptoms by methods of sufficient sleep, hot compress massage of eyes, multi-blink lubrication of eye surfaces, diet conditioning of physique and the like; if symptoms cannot be alleviated, further treatment methods can be divided into medical and surgical treatments: the internal medicine therapy can be given to artificial tears, mucolytic agents, anti-inflammatory drugs or antibiotics and the like according to the condition of a patient, and has the functions of moistening the ocular surface, reducing abnormal secretion or reducing inflammation so as to relieve the discomfort of the ocular surface; the surgical treatment is usually performed when the symptoms are serious, and comprises a lacrimal canaliculus cauterization operation, a lacrimal canaliculus plugging operation, an eyelid suturing operation and the like, wherein the permanent or temporary sealing of the lacrimal canaliculus can reduce the loss of tears and tear films on the ocular surface of a patient, and the suturing of the eyelid can avoid further damage and lesion of the cornea of the patient with serious dry eye syndrome.
Disclosure of Invention
The invention utilizes a specific method to artificially culture the antrodia cinnamomea mycelium and extract the active substances thereof. Animal experiments prove that the antrodia camphorata mycelium active substance prepared by the method has good protection effect on physical or chemical ocular surface injury, and can be used for new purposes of eye health care, prevention and/or treatment of xerophthalmia and the like.
The present invention provides a method for preparing an active substance of mycelia of Antrodia camphorata for preventing and/or treating dry eye, the method comprising the steps of:
(a) culturing Antrodia Camphorata mycelium at predetermined temperature and pH for 5-7 days;
(b) inoculating the cultured antrodia cinnamomea mycelia in the step (a) into a fermentation tank, and culturing for about 7-20 days at the preset temperature and the preset pH value under stirring;
(c) freeze-drying the cultured antrodia cinnamomea mycelia in the step (b), and grinding into powder to form antrodia cinnamomea mycelia freeze-dried powder;
(d) mixing a predetermined amount of solvent with the antrodia camphorata mycelium freeze-dried powder, shaking for extraction, and filtering to remove the antrodia camphorata mycelium freeze-dried powder to form antrodia camphorata mycelium extract liquid; and
(e) and (3) decompressing, concentrating and drying the antrodia camphorata mycelium extract liquid to obtain the antrodia camphorata mycelium active substance.
In one embodiment, the antrodia camphorata mycelium is a strain deposited in the China general microbiological culture Collection center, and the preservation number is as follows: mycelium of Antrodia camphorata of CGMCC NO. 0543.
In one embodiment, the predetermined temperature is 20-30 ℃ and the predetermined pH is 3-8.
In one embodiment, the culturing in step (a) of the preparation method is shaking culturing at a shaking rate of 50-250 rpm.
In one embodiment, the fermentation tank of step (b) of the above preparation method has a tank pressure of 0.5 to 1.0kg per square centimeter, an aeration rate of 0.05 to 2vvm, and a stirring rate of 5 to 250 rpm.
In one embodiment, the same medium is used in step (a) and step (b) of the preparation method.
In one embodiment, the medium used in the above preparation method comprises: cereals, sulfate compounds, phosphate compounds, saccharides, yeast extract, beans, or combinations thereof.
In one embodiment, the predetermined amount in step (d) of the above preparation method is 10-50 times of the weight of the lyophilized powder of antrodia camphorata mycelia.
In one embodiment, the solution in step (d) of the preparation method is an alcohol. Preferably, the alcohol is methanol or ethanol.
In another aspect, the present invention provides an active material of mycelia of antrodia camphorata for preventing and/or treating dry eye, which is prepared by the aforementioned preparation method.
In still another aspect, the present invention provides the use of the active substance of antrodia camphorata mycelium for the preparation of a medicament for the prevention and/or treatment of dry eye. Namely, the invention provides the application of the antrodia camphorata mycelium active substance in preparing the medicine for preventing and/or treating xerophthalmia.
In yet another aspect, the present invention provides a pharmaceutical composition for preventing and/or treating dry eye, the pharmaceutical composition comprising the antrodia camphorata mycelium active substance, and a pharmaceutically acceptable carrier, excipient, diluent or adjuvant.
In a further aspect, the present invention provides the use of the pharmaceutical composition for the preparation of a medicament for the prevention and/or treatment of dry eye. Namely, the present invention provides the use of the pharmaceutical composition for the preparation of a medicament for the prevention and/or treatment of dry eye.
Drawings
FIG. 1 is a graded decision criterion for a Corneal clarity (Corneal opacity) test, wherein a rating of 0 represents normal Corneal transparency; grade 1 represents mild opaqueness; grade 2 represents moderate opacity; grade 3 represents moderate opacity and the iris is unclear; grade 4 represents severe opaque degeneration with a marked white turbidity observed with corneal ulceration.
Fig. 2A to 2D are graphs showing results plotted according to results of a UVB mouse model Corneal appearance analysis test, in which fig. 2A is a Corneal smoothness (Corneal smoothness) test; FIG. 2B is a Corneal clarity (Corneal opacity) test; FIG. 2C is a Corneal map (Corneal topograph) test; figure 2D is a Corneal lesion staining (Corneal staining) test.
Fig. 3 is a photograph of a Central cornea (Central cornea) region stained by HE of mouse eyeball tissue in UVB mouse mode.
Fig. 4A to 4D are graphs of results plotted according to the results of corneal appearance analysis test in BAC mouse mode, in which fig. 4A is a corneal smoothness test; FIG. 4B is a corneal clarity test; FIG. 4C is a corneal map test; figure 4D is a corneal lesion staining test.
Fig. 5 is a photograph of the central corneal region of HE staining of mouse eyeball tissue in the BAC mouse model.
Detailed Description
In order to make the features and advantages of the present invention clearer, the following description of the embodiments of the present invention is provided with the accompanying drawings.
Principle of experiment
Ultraviolet (UV) is ubiquitous on the earth, and can be divided into UVA (315 plus 380nm), UVB (280 plus 315nm) and UVC (100 plus 280nm) according to different wavelength sections, the UVC has the strongest energy, but most of the UVC is absorbed by an ozone layer and has small influence on a human body, the UVA has stronger penetrating power, the eye water crystal pathological change and aging are caused, the reason for inducing cataract to penetrate is one, the UVB has weaker force than the UVA, the Ocular surface (Ocular surface) and the light injury of the cornea are mainly caused, and the long-term irradiation can cause the eye to have the symptoms of foreign body sensation and stabbing pain, corneal epithelium shedding and the like. The general public has less eye protection than wearing sunglasses and therefore unconsciously ignores the damage of UVB to the ocular surface resulting in chronic inflammation and even the development of dry eye.
Benzalkonium chloride (BAC, BAK) is a cationic surfactant, belongs to non-oxidative broad-spectrum bactericide, and can be used for sterilizing, disinfecting, antisepsis, emulsifying, descaling, solubilizing, etc. Early additions to ophthalmic water were often used as a preservative, but recent studies have shown that this compound causes tear film instability, loss of goblet cells, conjunctival squamous Metaplasia (Metaplasia) and Apoptosis (Apoptosis), disruption of the corneal epithelial barrier and damage to deeper ocular tissues, mild symptoms can lead to ocular inflammation and dry eye, and severe individuals can even cause permanent damage to the ocular surface and affect vision. The mechanism by which BAC contributes to these effects is not known, but current studies have demonstrated that it causes the release of pro-inflammatory cytokines (pro-inflammatory cytokines), apoptosis, and Oxidative stress (Oxidative stress) leading to an immune inflammatory response, and in addition, it interacts directly with lipid components on the tear film and cell membrane (Interaction).
Therefore, the present experiment was conducted based on the above principle, and two mouse animal models (UVB and BAC models) were designed to observe the prevention and/or treatment effects of the antrodia camphorata mycelium active substance on dry eye caused by physically or chemically induced ocular surface damage.
Experimental procedure
Antrodia camphorata mycelium culture and active substance preparation
The active substance of antrodia camphorata mycelium used in the following examples was prepared in the following manner:
(a) fresh antrodia camphorata mycelia (the strain is from a strain CGMCC NO.0543 which is deposited in the China general microbiological culture Collection center at 3/5/2001 and is publicly sold in Taiwan, and particularly referred to CN101803528B) are scraped from a plate and inoculated into a conical flask, the components in the table 1 are taken as a culture medium, and the culture medium is shake-cultured to the initial logarithmic propagation phase (log phase) (about 5-7 days) at the speed of 50-250rpm under the environment of 30 ℃ and pH 4.5. The conditions of temperature and pH value can be adjusted according to actual conditions, preferably 20-30 deg.C and pH value of 3-8. It is specifically noted that the present invention is not limited to the use of the above-mentioned deposited strain, and those skilled in the art can extract active substances using other mycelia of Antrodia camphorata according to the present invention and apply the same to the prevention and/or treatment of dry eye.
(b) Then, the culture in the Erlenmeyer flask was inoculated into a culture medium of a fermentation tank, the composition of which is also shown in Table 1, at 30 ℃ under a tank pressure of 0.5-1.0kg/cm2Then, air was introduced at a ventilation rate of 1.0vvm at a pH of 4.5, and the mixture was cultured at a stirring rate of 200rpm for about 10 days to yield the final product. The actual culture time, aeration rate and stirring rate can be adjusted as required, preferably 7 to 20 days of culture, the aeration rate is between 0.05 to 2.0vvm, and the stirring rate is between 5 to 250 rpm.
(c) Freeze drying collected antrodia camphorata mycelium, and grinding to obtain antrodia camphorata mycelium freeze-dried powder. Can be freeze-dried together with culture solution (liquid culture medium) in the fermentation tank to form Antrodia Camphorata mycelium whole solution lyophilized powder.
(d) Weighing antrodia camphorata mycelium freeze-dried powder, placing the antrodia camphorata mycelium freeze-dried powder into ethanol or methanol with the weight of 10-50 times of the antrodia camphorata mycelium freeze-dried powder for shaking extraction, wherein the extraction time is about 24 hours, and removing the freeze-dried powder by air suction filtration to obtain antrodia camphorata mycelium extract.
(e) The extract of the mycelium of Antrodia camphorata was concentrated to dryness under reduced pressure to obtain the active substance of the mycelium of Antrodia camphorata used in the following examples.
TABLE 1 Antrodia camphorata mycelium culture medium formula
| Composition (I) | Content (percentage by weight) |
| Cereals (e.g. wheat flour) | 1 |
| Peptone | 0.1 |
| Magnesium sulfate | 0.05 |
| Dipotassium hydrogen phosphate | 0.05 |
| Ferric sulfate | 0.05 |
| Sucrose | 2 |
| Yeast extract, powder and paste | 0.5 |
| Beans (such as soybean powder, mung bean powder, etc.) | 0.2 |
In particular, the formulation of the culture medium used in the present invention can be adjusted as required, or can be used in combination with commercially available culture medium formulations, and is not particularly limited.
Example 1 UVB mouse model
Female mice of ICR strain were used in this experiment, purchased from Lescow biotechnology, Inc. (Taiwan, China), 7-10 weeks old, and 25-33g in body weight, and were raised under conditions of 12 hours light-dark cycle, 20. + -. 2 ℃ and 50. + -. 5% relative humidity, and provided with clean feed and drinking water.
The test was carried out by randomly dividing the mice into 4 groups of 6 mice each, namely a control group (fed with an equivalent amount of physiological saline solution without UVB treatment), a UVB-treated group (fed with UVB treatment), a low-dose test group (fed with 10 mg/kg. bw of the above-prepared antrodia camphorata mycelium active substance, fed with UVB treatment) and a high-dose test group (fed with 100 mg/kg. bw of the above-prepared antrodia camphorata mycelium active substance, fed with UVB treatment). The unit of mg/kg · bw here indicates how many mg of the drug is administered per kg body weight, and bw represents body weight.
The experiment was carried out for 10 days, the experimental groups were each administered with the active substance of the mycelium of Antrodia camphorata at the experimental design dose every day, the UVB treatment group and the experimental groups started from day 4, the mice were anesthetized with 2.5% Avertin every day and placed in a dark box with 0.72J/cm irradiation of the eyeball upward2UVB at intensity for 90 seconds, the ocular surface of the mice was damaged. In all groups, corneal appearance evaluation and post-sacrifice eye tissue staining analysis were performed on day 10 of the experiment to evaluate whether the antrodia camphorata active substance could ameliorate ocular surface damage caused by UVB, and the experimental evaluation method is detailed in the next paragraph, "methods of implementation".
Example 2 BAC mouse model
Female mice of the ICR strain were used in this experiment, purchased from Lescow Biotech, Inc. (Taiwan), 7-10 weeks old, and fed under conditions of 12 hours light-dark cycle, 20 + -2 deg.C and 50 + -5% relative humidity, and provided clean feed and drinking water.
The test was conducted by randomly dividing the mice into 4 groups of 6 mice each, which were a control group (fed with an equivalent amount of physiological saline solution without BAC treatment), a BAC-treated group (fed with BAC treatment), a low-dose test group (fed with 10 mg/kg. bw of the above prepared antrodia camphorata mycelium active substance, fed with BAC treatment) and a high-dose test group (fed with 100 mg/kg. bw of the above prepared antrodia camphorata mycelium active substance, fed with BAC treatment).
The test is carried out for 14 days, the test group is fed with the active substance of the antrodia camphorata mycelium with the test design dose every day from day 1 to day 13, and from day 4 to day 13, the BAC treatment group and the test tissue mice take 5 mu L of 0.2% BAC to be spotted on the eyeballs every day, so that the eye surfaces of the mice are damaged. All groups were subjected to corneal appearance evaluation and histological staining analysis on the 14 th day of the experiment to evaluate whether the active substance of antrodia camphorata mycelium could improve ocular surface damage caused by BAC, and the experimental evaluation method is detailed in the next paragraph, "methods of implementation".
Implementation method-evaluation of degree of ocular surface injury
In the test, two mouse animal models (example 1-UVB and example 2-BAC) are adopted to observe the protective effect of the antrodia camphorata mycelium active substance on the ocular surface, and according to the test design, samples are respectively taken during the test and after the test is finished to evaluate the ocular surface damage degree and the xerophthalmia.
The evaluation items comprise cornea appearance analysis and HE staining histology analysis; wherein the corneal appearance analysis was performed with 4 assessments of corneal smoothness, corneal clarity, corneal map, and corneal damage staining.
The Corneal appearance analysis was classified into 4 tests of Corneal smoothness (Corneal smoothness), Corneal clarity (Corneal opacity), Corneal map (Corneal topograph) and Corneal damage staining (Corneal staining), and a higher rating score indicates a higher degree of Corneal damage.
(1) Corneal smoothness analysis (Corneal smoothenness): irradiating the surface of an eyeball by using an annular light source, and grading according to the integrity of an annular image of a cornea reflection light source into 0 grade to 5 grades; the image at level 0 is a complete and undistorted ring, the ring at levels 1-3, 1/4, 1/2, and 3/4 in sequence, is distorted at level 4, and the most severe level 5 is a ring that is extremely distorted to be indistinguishable.
(2) Corneal clarity analysis (Corneal opacity): irradiating the eyeball with a light source, observing the clarity of the cornea, and classifying into 0 grade to 4 grades according to the opacity degree; grade 0 is normal corneal transparency, grades 1-3 are mild, moderate, and moderate (unclear iris) opacity, respectively, while grade 4 is severe opacity degeneration, with marked white turbidity and accompanying corneal ulceration, and the detailed grading criteria are schematically shown in figure 1.
(3) Corneal mapping analysis (Corneal topograph): corneal map analysis projects the ocular surface in a quintuple annular pattern, allowing a wide range of corneal smoothness to be observed. The evaluation method comprises dividing the ocular surface into 4 zones equally by cross, wherein each zone has five circular arcs of 5, and each time 1 circular arc is distorted or cannot be judged, counting 1 point, and the total eyes are 20 points; higher score indicates higher degree of corneal unevenness, and is classified into 0-4 according to degree, score 0 is classified into 0, 1-4 is classified into 1, 5-9 is classified into 2, 10-14 is classified into 3, 15-19 is classified into 4, and the most serious 20 is classified into 5.
(4) Corneal lesion staining analysis (Corneal staining): since the damaged cornea is stained with a stain, the degree of corneal damage can be assessed by the stained area. The analysis results were rated as 0-5, those not stained were rated as 0, those below 25% were rated as 1, those 25-50% were rated as 2, those 50-75% were rated as 3, those 75% -99% were rated as 4, and all corneas were stained were rated as 5.
HE staining (hematoxin and Eosin stain; H & E stain) is one of the most commonly used staining methods in histology, the dye Hematoxylin (Hematoxylin) stains basophilic structures into bluish purple, such as intranuclear chromatin and intracellular ribosome, etc., and Eosin (Eosin) is an acid dye, which makes cytoplasm and extracellular matrix red, so as to facilitate histological identification. After the last day of the experiment, the mice were sacrificed, and the eyeball tissues were taken out and paraffin-embedded, sliced and HE-stained, and the number of cell layers, the shape and the thickness of the central cornea were observed to evaluate the effect of the active substance given to the antrodia camphorata mycelia.
The above experimental results were statistically analyzed by the mother-number-free assay Mann-Whitney U using SPSS software version 18, and are shown in FIGS. 2A to 5. In the experimental results shown in the figures, asterisks (×) indicate p <0.05, i.e. there is a significant difference between the two.
Results of the experiment
EXAMPLE 1 knotFruit UVB mouse model
Corneal appearance analysis
The test results are shown in fig. 2A to 2D, in which the X-axis from left to right is respectively the "bw" in the control group (Blank), the treatment group (UVB treatment), the low dose test group (10mg/kg · bw of the above prepared antrodia camphorata mycelium active substance, UVB treatment) and the high dose test group (100mg/kg · bw of the above prepared antrodia camphorata mycelium active substance, UVB treatment), the low dose test group and the high dose test group, so as to make the drawings clearer; the number of samples n in each group was 6, and the asterisks in the figure indicate p <0.05, i.e., there was a significant difference between the two. In four tests of corneal smoothness (corresponding to fig. 2A), corneal clarity (corresponding to fig. 2B), corneal map (corresponding to fig. 2C) and corneal damage staining (corresponding to fig. 2D), significant corneal damage was observed in the UVB-treated group (highest rating score), whereas the damage was improved in the active substance-fed group of the invention (lower rating score) and a dose effect was observed in the high dose group (100mg/kg · bw) compared to the low dose group; in both corneal smoothness (FIG. 2A) and corneal lesion staining (FIG. 2D) tests, the administration of high doses of active substance (100mg/kg bw of the above-prepared A. camphorata mycelium active substance) significantly reduced corneal lesions in mice.
HE staining
After the mice were sacrificed at day 10, the Central cornea (Central cornea) was taken and the corneal tissue condition was evaluated by HE staining, and as a result, as shown in fig. 3, several layers of structures were observed in the normal corneal epithelial layer, while the UVB-treated group, the low dose group and the high dose group were not much different from the control group in the number of cell layers and structure, indicating that UVB causes ocular surface damage and does not act on the corneal epithelial layer.
Example 2 results BAC mouse animal model
Corneal appearance analysis
The test results are shown in FIGS. 4A to 4D, in which the X-axis from left to right represents a control group (Blank), a treatment group (BAC treatment), a low dose test group (10 mg/kg. bw of the active substance of the prepared A. camphorata mycelium, BAC treatment) and a high dose test group (100 mg/kg. bw of the active substance of the prepared A. camphorata mycelium, BAC treatment); the number of samples n in each group was 6, and the asterisks in the figure indicate p <0.05, with significant differences between the two. Processing BAC results in an increase in the values of four tests, corneal smoothness (corresponding to fig. 4A), corneal clarity (corresponding to fig. 4B), corneal map (corresponding to fig. 4C), and corneal lesion staining (corresponding to fig. 4D), with significant differences between corneal clarity (fig. 4B) and corneal lesion staining (fig. 4D). The high dose of active substance administered to mice (the active substance fed with 100mg/kg · bw of the prepared antrodia camphorata mycelium) significantly reduced the degree of corneal damage due to BAC treatment in all four evaluations.
HE staining
After the mice were sacrificed at day 14, the Central cornea (Central cornea) was taken and evaluated for corneal tissue status by HE staining, and as a result, as shown in fig. 5, several layers of structures were observed in the normal corneal epithelial layer, the corneal epithelial cell layer was significantly damaged in the BAC-treated group, and the damage was improved in both the low dose group and the high dose group to which the active substance of antrodia camphorata mycelium was administered.
The above tests prove that the active substance of the mycelium of Antrodia camphorata prepared by the preparation method of the embodiment has the effect of physically and chemically damaging the cornea, and can improve the damage condition.
The above embodiments are specific illustrations of possible embodiments of the present invention, but the present invention is not limited to these embodiments. Those skilled in the art can make equivalent implementations or modifications without departing from the spirit of the present invention.
Claims (7)
1. The use of an active substance of antrodia camphorata mycelium for the preparation of a medicament for the prevention and/or treatment of dry eye; wherein, the antrodia camphorata mycelium is a strain deposited in China general microbiological culture collection center, and the preservation number is as follows: the antrodia camphorata mycelium of CGMCC NO.0543, and the preparation method of the antrodia camphorata mycelium active substance comprises the following steps:
(a) culturing Antrodia Camphorata mycelium at predetermined temperature and pH for 5-7 days;
(b) inoculating the cultured antrodia cinnamomea mycelia in the step (a) into a fermentation tank, and culturing for 7-20 days at the preset temperature and the preset pH value under stirring;
(c) freeze-drying the cultured antrodia cinnamomea mycelia in the step (b), and grinding into powder to form antrodia cinnamomea mycelia freeze-dried powder;
(d) mixing a predetermined amount of methanol or ethanol with the antrodia camphorata mycelium freeze-dried powder, shaking for extraction, and filtering to remove the antrodia camphorata mycelium freeze-dried powder to form antrodia camphorata mycelium extract; and
(e) and (3) decompressing, concentrating and drying the antrodia camphorata mycelium extract liquid to obtain the antrodia camphorata mycelium active substance.
2. Use of a pharmaceutical composition comprising an active substance of antrodia camphorata mycelium together with a pharmaceutically acceptable carrier, excipient, diluent or adjuvant for the preparation of a medicament for the prevention and/or treatment of dry eye; wherein, the antrodia camphorata mycelium is a strain deposited in China general microbiological culture collection center, and the preservation number is as follows: the antrodia camphorata mycelium of CGMCC NO.0543, and the preparation method of the antrodia camphorata mycelium active substance comprises the following steps:
(a) culturing Antrodia Camphorata mycelium at predetermined temperature and pH for 5-7 days;
(b) inoculating the cultured antrodia cinnamomea mycelia in the step (a) into a fermentation tank, and culturing for 7-20 days at the preset temperature and the preset pH value under stirring;
(c) freeze-drying the cultured antrodia cinnamomea mycelia in the step (b), and grinding into powder to form antrodia cinnamomea mycelia freeze-dried powder;
(d) mixing a predetermined amount of methanol or ethanol with the antrodia camphorata mycelium freeze-dried powder, shaking for extraction, and filtering to remove the antrodia camphorata mycelium freeze-dried powder to form antrodia camphorata mycelium extract; and
(e) and (3) decompressing, concentrating and drying the antrodia camphorata mycelium extract liquid to obtain the antrodia camphorata mycelium active substance.
3. Use according to claim 1 or 2, wherein the predetermined temperature is 20-30 ℃ and the predetermined pH value is 3-8.
4. The use according to claim 1 or 2, wherein the culturing in step (a) is shaking culture and the shaking rate is 50-250 rpm.
5. Use according to claim 1 or 2, wherein the fermenter in step (b) has a tank pressure of 0.5-1.0kg per square centimeter, an aeration rate of 0.05-2vvm and a stirring rate of 5-250 rpm.
6. Use according to claim 1 or 2, wherein the same medium is used in step (a) and step (b).
7. The use according to claim 1 or 2, wherein the predetermined amount in step (d) is 10-50 times the weight of the antrodia camphorata mycelium lyophilized powder.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1799562A (en) * | 2005-09-28 | 2006-07-12 | 莱阳农学院 | Antrodia camphorata mycelium fermented extract and application thereof |
| CN1865450A (en) * | 2005-09-28 | 2006-11-22 | 莱阳农学院 | Fatty acid extract of Antrodiacamphorata mycelium and its uses |
| CN101803528A (en) * | 2009-02-16 | 2010-08-18 | 葡萄王生技股份有限公司 | Novel cultural method of antrodia camphorata mycelium |
| CN104223044A (en) * | 2014-09-17 | 2014-12-24 | 中山安荞生物科技有限公司 | Extraction method for adenosine from antrodia camphorata mycelia |
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| CN1865450A (en) * | 2005-09-28 | 2006-11-22 | 莱阳农学院 | Fatty acid extract of Antrodiacamphorata mycelium and its uses |
| CN101803528A (en) * | 2009-02-16 | 2010-08-18 | 葡萄王生技股份有限公司 | Novel cultural method of antrodia camphorata mycelium |
| CN104223044A (en) * | 2014-09-17 | 2014-12-24 | 中山安荞生物科技有限公司 | Extraction method for adenosine from antrodia camphorata mycelia |
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| CN106309508A (en) | 2017-01-11 |
| TW201701892A (en) | 2017-01-16 |
| TWI552755B (en) | 2016-10-11 |
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