CN113801798B - Acidocella adephagia strain A50, extracellular polysaccharide produced by same and application thereof - Google Patents
Acidocella adephagia strain A50, extracellular polysaccharide produced by same and application thereof Download PDFInfo
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- CN113801798B CN113801798B CN202110881679.XA CN202110881679A CN113801798B CN 113801798 B CN113801798 B CN 113801798B CN 202110881679 A CN202110881679 A CN 202110881679A CN 113801798 B CN113801798 B CN 113801798B
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- Prior art keywords
- strain
- extracellular polysaccharide
- polysaccharide
- blastobotrys adeninivorans
- bile acid
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- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
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- A—HUMAN NECESSITIES
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
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- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
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- A—HUMAN NECESSITIES
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/06—Antihyperlipidemics
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- C—CHEMISTRY; METALLURGY
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- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
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- C—CHEMISTRY; METALLURGY
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- C12P19/00—Preparation of compounds containing saccharide radicals
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Abstract
The invention belongs to the technical field of microorganisms, and particularly relates to a Blastobotrys adeninivorans A strain and extracellular polysaccharide produced by the strain, and discloses the effect of extracellular polysaccharide in combination with bile acid. The strain of the invention was isolated from a Liupu tea sample of the pile fermentation process and identified as Blastobotrys adeninivorans by molecular and morphological analysis. The extracellular polysaccharide of the strain contains 78.08% of total sugar and 17.81% of uronic acid, and the ultraviolet spectrum shows that the extracellular polysaccharide is free of nucleic acid protein, the infrared spectrum shows that the extracellular polysaccharide is acidic polysaccharide and is sulfated polysaccharide, and the alpha-pyran ring and the furan ring exist at the same time. The extracellular polysaccharide has better bile acid binding effect in vitro, has half-inhibition concentration IC50 value of 64.325mg/ml, has potential blood lipid reducing capability, and has good application prospect in the aspects of preventing and treating hyperlipidemia and the like.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to a Blastobotrys adeninivorans A strain capable of producing extracellular polysaccharide and having a bile acid binding effect and application thereof.
Background
Hyperlipidemia refers to a condition characterized by an abnormally elevated level of one or more lipid components in the plasma (or serum). Atherosclerosis caused by this disorder is an important factor in the occurrence of cardiovascular diseases. Conversion to bile acids in the liver is the main way cholesterol is metabolized in the body. More than 95% of bile acid generated by the liver can be reabsorbed by human body, and the small part which is not absorbed can be discharged with feces. Bile acid binders can bind with bile acids in the small intestine to form insoluble complexes and be discharged outside the body, resulting in accelerated conversion of cholesterol in the liver or blood to bile acids, thereby exerting an effect of lowering cholesterol in the body.
Bile acid binders fall into two broad categories: anion exchange resins and dietary fiber polysaccharides. The administration of some dietary fiber polysaccharides such as oat and barley beta-glucan, pectin in fruit, guar gum, results in increased bile acid excretion in feces to 35% -65%. The microbial polysaccharide is widely used in various fields such as food and pharmacy because of low cost and easy production, and is not affected by seasons. However, compared to plant polysaccharides, there is less research on bile acid binding activity of microbial exopolysaccharides, which is detrimental to the development of microbial resources.
Liupu tea is considered as a healthy and health-preserving leisure drink, and Liupu tea polysaccharide has been studied to bind bile acid in blood. The special tea making process makes the formation of unique quality of Liupu tea not leave the action of microbe in the pile fermentation process. Studies have reported that exopolysaccharide of Eurotium cristatum is one of the active ingredients of Fuzhuan tea polysaccharide, and has lipid-lowering activity. However, no functional information has been reported about Blastobotrys adeninivorans exopolysaccharides in Liupu tea.
Disclosure of Invention
In order to overcome the defects in the prior art, the invention provides the following technical scheme:
the invention provides a Blastobotrys adeninivorans A strain capable of producing extracellular polysaccharide and having a bile acid binding effect, wherein Blastobotrys adeninivorans A is preserved in China Center for Type Culture Collection (CCTCC) at the year 2021 and the month 04 and has the preservation number of CCTCC NO: M2021485.
The Blastobotrys adeninivorans strain belongs to the genus Blastobotrys of the family Trichomonasceae of the order Saccharomyces, of the order Saccharomyces.
(1) The A50 strain is obtained by fermenting tea leaves in the pile fermentation process of Liupu tea enterprises in Guangxi Zhuang nationality, chinese urban areas, and the identification result shows that the colony of the Alternaria alternata A50 on a PDA culture medium is white and dry, and is circular and opaque; cells were observed under the microscope to be round or quasi-round.
Further, the a50 strain produces extracellular polysaccharide in PDB medium. The fermentation liquor of the strain is subjected to centrifugation to collect supernatant, then 3 times of absolute ethyl alcohol with volume is added for precipitation twice, and the fermentation liquor is dried. Adding distilled water for dissolution, removing protein by Sevag method, adding three times of 95% ethanol, and centrifuging for precipitation. Dissolving the precipitate with distilled water, loading into dialysis bag with molecular weight cut-off of 8000Da-12000 and Da, and lyophilizing sugar solution to obtain EPS-A50.
The EPS-A50 had a composition of 78.08% total sugars, 17.81% uronic acid. The ultraviolet spectrum has no absorption peak at 260nm-280nm, indicating no nucleic acid or protein. Infrared spectrum indicates that the extracellular polysaccharide is an acidic polysaccharide and is a sulfated polysaccharide; the monosaccharide composition has mannose, with both an alpha-pyran ring and a furan ring. The molecular weight of EPS-A50 was 320.799kDa (26.28%), 26.618kDa (72.30%), 1.786kDa (1.42%), the monosaccharide composition consisting of Man, galA and Gal, the molar ratios being 0.99, respectively: 1.34:39.02.
furthermore, the extracellular polysaccharide has 23.81% -79.24% of in-vitro binding bile acid effect at the concentration of 10-110mg/mL, and the half-inhibitory concentration IC50 value is 64.325mg/mL. Therefore, the microbial agent can be applied to preparation of microbial agents, for example, can be used as a bile acid binding agent for reducing blood fat, and can be especially used for preparing drugs for preventing and/or treating hyperlipidemia.
The microbial agent can be prepared into preparations acceptable in foods, health-care products or medicines. The strain is from Liupu tea, so that the components are safe and reliable, and the applicability is strong.
Compared with the prior art, the invention has the following advantages: the strain has reliable source and safe components. And the microbial fermentation technology is adopted, so that the enlarged production is easy to realize. The extracellular polysaccharide produced by the strain has good effect of combining bile acid and strong applicability.
Drawings
FIG. 1 is a microscopic image of strain A50.
FIG. 2 is a diagram of the A50 strain neighbor-training algorithm system evolution tree.
FIG. 3 is an ultraviolet spectrum of EPS-A50.
FIG. 4 is an infrared spectrum of EPS-A50.
FIG. 5 is a graph showing the molecular weight of EPS-A50.
FIG. 6 is a diagram showing the monosaccharide composition of EPS-A50.
FIG. 7 is a graph showing the effect of EPS-A50 binding bile acids in vitro.
Detailed Description
The technical solutions of the present invention will be clearly and completely described in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1 Blastobotrys adeninivorans A50 isolation, cultivation and identification of Strain
1. Isolation and culture of strains
(1) Collecting bacterial colonies: 2g of fresh Liupu tea sample (collected place: fermented tea sample in pile fermentation process of Liupu tea enterprise in Guangxi Zhuang nationality, phoenix state) in pile fermentation process is taken, sheared by scissors, poured into a blue mouth bottle filled with 20mL of sterile water, and oscillated for 30min under the condition of 80r/min, so that microorganisms attached to the tea fall off.
(2) Colony purification: 1mL stock solutions of bacterial colonies are respectively diluted to 10 -1 、10 -2 、10 -3 、10 -4 、10 -5 、10 -6 Is absorbed by the bacteria suspension with gradient concentration200. Mu.L of the bacterial suspension was taken and poured into PDA and uniformly coated with a disposable coating rod. The culture was carried out in an incubator at 28℃for 2 to 3 days. After the bacterial colonies grow out of the coated culture dish, sorting by naked eyes, picking the colony edges of the strains with different forms, inoculating the colony edges to a PDA, and repeating the purification for more than 4 times until the colony becomes a single colony.
2. Morphological identification of strains
(1) The bacterial colony of the A50 strain on the PDA is white, dry, circular and opaque.
(2) And (3) observing by a microscope: as shown in FIG. 1, the cells are round or quasi-round.
3. Molecular characterization
(1) First 16S rRNA sequencing was performed: the single colony after purification was inoculated to PDA and cultured in an incubator at 28℃for 12 to 24 hours. Taking 50 mu L of sterilized resin solution in eight connecting tubes, picking up a small part of thalli by using a toothpick, placing the thalli in the eight connecting tubes and grinding for 1min. The eight-connecting tube is kept at the temperature of 100 ℃ in a metal bath for 10min, and the supernatant is obtained by centrifugation to obtain the template DNA. The template DNA fragments were amplified and sent to biosystems for sequencing. The sequence of 16S rRNA of the A50 strain is obtained and is shown as SEQ ID NO. 1.
(2) All the sequences of 16S rRNA from the species Acinetobacter adenivorum were collected in the EzTaxon database (http:// www.ezbiocloud.net/ezTaxon). All sequences were aligned and cut using software CLUSTAL_X, and the evolutionary tree was constructed using software MEGA X, with algorithms of neighbor-joining and Maximum-likelihood algorithms (Bootstrap: 1000 alternatives).
Phylogenetic analysis was performed using sequence alignment, and the results of the evolutionary tree are shown in FIG. 2. The results show that the A50 strain belongs to Blastobotrys, the closest strain is Blastobotrys adeninivorans, and is preserved, the A50 strain is preserved in China Center for Type Culture Collection (CCTCC) at the year 04 and the month 29 of 2021, and the preservation number is CCTCC NO: M2021485, and the preservation address is: university of martial arts in chinese.
Example 2 Blastobotrys adeninivoransA50 fermentation culture of Strain
1. Seed liquid culture
The strain is taken out of the freezing tube box according to the serial number, placed at room temperature for dissolution, inoculated to PDA, cultured for 24 hours at 28 ℃, then inoculated to select single bacterial colony to be inoculated to a YEPD liquid culture medium (20.0 g of peptone, 10.0g of yeast powder, 20.0g of glucose and 1000.0mL of distilled water), fermented for 18-24 hours at 28 ℃ and 120r/min, and the absorbance value of bacterial liquid at 600nm is regulated to be about 1.0, thus obtaining seed liquid.
2. Expansion culture
500mL Erlenmeyer flasks containing 250mL PDB were inoculated at a 5% inoculation rate and fermented at 28℃for 168 hours at 120 r/min.
Example 3 Blastobotrys adeninivoransA50 extraction, purification and component determination of extracellular polysaccharide EPS-A50 produced by Strain
1. Extracting and purifying
After fermentation, the fermentation broth is centrifuged at 6000r/min for 20min, three times of absolute ethyl alcohol is added, the sediment is deposited overnight in a refrigerator at 4 ℃, a certain volume of distilled water is added for remelting after the sediment is dried in the air, and then 3 times of 95% ethanol is added at 4 ℃ overnight. Collecting the precipitate, and air drying. Distilled water is added to dissolve and prepare 1% sugar solution, and protein is removed by Sevag method. Mixing n-butanol with distilled water in equal volume, and performing ultrasonic treatment for 10min to obtain water saturated n-butanol solution as the upper layer solution. 4 volumes of chloroform were added to the n-butanol solution to obtain Sevag reagent. One third of the volume of Sevag reagent was added to the sugar solution and stirred with a magnetic stirrer for 20min, poured into a separatory funnel and allowed to stand until a clear delamination was seen, and the lower organic reagent and the middle deformable proteins were removed. The upper layer was collected as a deproteinized sugar solution. The deproteinization operation was repeated until no milky precipitate was seen. The organic reagent was then removed by rotary evaporation at 60 ℃. Three volumes of 95% ethanol were added and the precipitate was collected by centrifugation at 4 ℃ overnight in a refrigerator. Dissolving the precipitate with distilled water, loading into dialysis bag with molecular weight cutoff of 8000Da-12000 Da, dialyzing in distilled water of 100 times volume, changing water every 4 hours, and dialyzing for 48 hr. After dialysis, the sugar solution was freeze-dried to give EPS-A50.
2. Component measurement
Methods for determining total sugar, uronic acid and protein content were slightly modified with reference to the literature. The results showed that EPS-A50 contained 78.08.+ -. 1.59% total sugar, 17.81.+ -. 1.21% uronic acid, and no protein.
Example 4 Blastobotrys adeninivorans A50 ultraviolet and Infrared Spectrometry analysis of extracellular polysaccharide EPS-A50 produced by the Strain
1. Ultraviolet spectrum
The extracellular polysaccharide sample was dissolved in distilled water and adjusted to an appropriate concentration, followed by scanning analysis with an ultraviolet spectrometer (Shimadzu corporation, UV 2501 PC) at a scanning wavelength in the range of 200nm to 400 nm. As shown in FIG. 3, EPS-A50 had no absorption peak at 260nm-280nm, indicating that it did not contain nucleic acid and protein.
2. Infrared spectrum
The extracellular polysaccharide sample 3mg and potassium bromide solid powder 270mg were weighed, mixed and ground until the sample particles were free of lens, and then pressed into uniform flakes by a tabletting machine. Scanning with Fourier transform infrared spectrometer (Fourier transform-infra-red, FT-IR) with a scanning range of 4000cm -1 -400cm -1 。
EPS-A50 as shown in FIG. 4 at 3407.47cm -1 The strong absorption peak at this point can be attributed to the stretching vibration peak of the O-H bond, indicating the presence of hydroxyl groups; 2933.45cm -1 The peak at this point is-CH on the sugar chain 2 Or the stretching vibration peak of the C-H bond of the-CH. The above is the characteristic absorption peak of polysaccharide. 1639.50cm -1 The peaks near where this is due to the presence of bound water. 1418.50cm -1 The peak may be the characteristic of carboxyl or carboxylate, indicating that the extracellular polysaccharide is an acidic polysaccharide, which is consistent with the determination of physicochemical index. Polysaccharide at 1239.15cm -1 There is an absorption peak, which is the stretching vibration peak of s=o, indicating a sulfated polysaccharide. 1100cm -1 -1010cm -1 There are 3 peaks representing pyran rings and 2 peaks representing furan rings. The results showed only 2 peaks for the polysaccharide, indicating a furan ring. EPS-A50 at 1079.18cm -1 The absorption peak of the ether bond C-O-C and the hydroxyl group of the pyran ring. Polysaccharide at 1021.13cm -1 The C-O stretching vibration peak is shown. 931.67cm -1 The peak indicates that mannose may be contained. At 890cm -1 And 812cm -1 No absorption peak (specific signal of beta configuration) at 850cm -1 The peak was left and right, indicating that the alpha-glycosidic bond was contained. EPS-A50 at 765.13cm -1 The absorption peak is caused by the symmetrical stretching vibration of the alpha-pyran ring.
The above results illustrate: the extracellular polysaccharide is acidic polysaccharide and is a sulfated polysaccharide, and the monosaccharide composition is mannose. EPS-A50 has both an alpha-pyran ring and a furan ring.
Example 5 Blastobotrys adeninivorans A50 detection of molecular weight and monosaccharide composition of extracellular polysaccharide EPS-A50 produced
1. Determination of molecular weight
The molecular weight of the extracellular polysaccharide was determined by gel permeation chromatography (Gel Permeation Chromatography, GPC). Extracellular polysaccharide and dextran standards (580, 29330, 10730, 31420, 63350, 143400, 280500, 841700, 2560000 and 7500000 Da) were dissolved in ultrapure water to prepare 1mg/mL solutions. Chromatographic conditions: waters GPC model 1515 equipped with a differential refractive detector, column: WATERS Ultrahydrogel 500column (7.8X100 mm), WATERS Ultrahydrogel 250column (7.8X100 mm) and WATERS Ultrahydrogel 120column (7.8X100 mm) are used in series. Mobile phase: ultrapure water; flow rate: 1.0mL/min; column temperature: 30 ℃; sample injection amount: 40. Mu.L.
As a result, FIG. 5 shows that FIG. 5 (a) is a standard curve of dextran standard, R 2 =0.9985 indicates a good linear range and a reliable method. FIG. 5 (b) is a GPC chart of EPS-A50, showing that the molecular weight of EPS-A50 is 320.799kDa (26.28%), 26.618kDa (72.30%), 1.786kDa (1.42%) by substituting the retention time into the standard curve.
2. Monosaccharide composition determination
Hydrolysis of polysaccharide: placing 0.0500g of extracellular polysaccharide powder into a hydrolysis tube, adding 1mL of HCl (3.0 mol/L), sealing, mixing, hydrolyzing at 110deg.C for 4-6 hr, cooling, adjusting pH to neutrality with 3.0mol/L NaOH, and absorbing 400 μL for derivatization.
Shan Biao and gradient mixing. (1) Single label. The monosaccharide controls (arabinose, mannose, glucose, galactose, galacturonic acid, glucuronic acid, fucose and rhamnose) were dissolved in ultrapure water at a concentration of 2 mmol/L. And (2) mixing. The monosaccharide controls were co-dissolved in ultrapure water so that the concentration of each standard was 1.6mg/mL, and post-dilution was a gradient mix of 6 concentrations of 1.6, 1.2, 1.0, 0.8, 0.4, 0.2 mg/mL.
Monosaccharide derivatization: 400 mu L of standard solution (Shan Biao and gradient mixed standard) and hydrolyzed polysaccharide sample are taken, 0.5mol/L PMP-methanol solution and 0.3mol/L NaOH are added into 400 mu L of the standard solution, and the mixture is uniformly mixed and reacted for 100min in a water bath at 70 ℃. Then, 500. Mu.L of 0.3mol/L HCl was added, the PMP reagent was extracted 3 times with 5mL of chloroform each time, the chloroform layer was discarded, and after centrifugation of the water layer, 1mL of the supernatant was filtered through a 0.45 μm organic filter membrane and analyzed by high performance liquid chromatography (High Performance Liquid Chromatography, HPLC).
Analysis was performed using a Waters 2984LC high performance liquid chromatograph. Chromatographic conditions: waters C18 column (4.6X105 mm,5 μm); taking acetonitrile-0.02 mol/L ammonium acetate solution (17:83) as a mobile phase chromatographic condition; column temperature is 30 ℃; the flow rate is 1.0mL/min; the detection wavelength is 250nm, and the sample injection amount is 10 mu L.
From FIGS. 6 (a) and 6 (b), the PMP pre-column derivatization method effectively separated 8 monosaccharide standards, and the retention time and peak area were brought into the standard curve to calculate the monosaccharide composition of EPS-A50, which was found to consist of Man, galA and Gal, at a molar ratio of 0.99:1.34:39.02.
example 6 extracellular polysaccharide EPS-A50 produced by Arthrobacter adenylatus in vitro binding to bile acid.
First, a bile acid stock solution is prepared: glycocholic acid (9 mmol/L), chenodeoxycholic acid (9 mmol/L), glycodeoxycholic acid (9 mmol/L), taurochenodeoxycholic acid (4.5 mmol/L), taurodeoxycholic acid (4.5 mmol/L) was dissolved in phosphate buffer solution with pH=6.8, i.e. prepared as a 36mmol/L bile acid stock solution. Stock solutions were stored in a-20 ℃ freezer and diluted to 0.72 μmol/mL bile acid working solution prior to use.
Reference is made to the scheme of Wu et al and modifications are made. 10mg/mL-110mg/mL of exopolysaccharide was placed in a 15X 150mm glass tube, 1mL of 0.01mol/L HCl was added, and acid digestion of the stomach was simulated at 37℃for 2 hours at 120 r/min. The pH of the sample was adjusted to 7-7.5 with 0.2mol/L NaOH. To each sample to be tested were added 4mL of bile acid working solution (0.72. Mu. Mol/mL) and 5mL of pancreatin (10 mg/mL in phosphate buffer at pH=6.8) and simulated in vitro intestinal digestion at 37℃at 120r/min for 2 hours. Centrifuging the mixture at 6000r/min for 20min, collecting supernatant, and storing at-20deg.C. Distilled water was used as a negative control.
And detecting the content of bile acid in the supernatant according to the determination method of the total bile acid kit. The formula of bile acid content is:
wherein, the enzyme labeling instrument (Infinite M200 PRO) reads the absorbance A0 at 405nm, and reads the absorbance A1 after 3min, wherein DeltaA=A1-A0. The ΔA measurement is the ΔA value of the sample, the ΔA calibration is the ΔA value of the calibrator, and the concentration of bile acid calibrator is 50. Mu. Mol/L.
As a result, the IC50 value of the half inhibitory concentration of EPS-A50 was 64.325mg/ml as shown in FIG. 7. Shows that EPS-A50 has potential hypolipidemic capacity and is expected to be a medicine for preventing and treating hyperlipidemia, and the strain is expected to be developed into a commercial strain.
The embodiments of the present invention have been described in detail above, but the present invention is not limited to the described embodiments. It will be apparent to those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, and yet fall within the scope of the invention.
Sequence listing
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<120> Acinetobacter adeniveum A50 strain, extracellular polysaccharide produced and application thereof
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Claims (8)
1. Acidocella adephagiaBlastobotrys adeninivorans) Use of strain a50 for the preparation of a bile acid binder, characterized in thatBlastobotrys adeninivorans A50 strain is preserved in China Center for Type Culture Collection (CCTCC) at 2021, 04 and 29 days, and the preservation number is CCTCC NO: M2021485.
2. The use according to claim 1, wherein the bile acid binder is used in food or pharmaceutical products.
3. The use according to claim 1, wherein the yeast Arthrobacter adephagia is usedBlastobotrys adeninivorans A50 strain is fermented conventionally, and supernatant is taken.
4. An extracellular polysaccharide EPS-A50, which is characterized in that the extracellular polysaccharide is prepared from an adeps Zosterae MarinaeBlastobotrys adeninivorans A50 strain is fermented according to the conventional method, and supernatant fluid is taken to obtain; the saidBlastobotrys adeninivorans A50 strain is preserved in China Center for Type Culture Collection (CCTCC) at 2021, 04 and 29 days, and the preservation number is CCTCC NO: M2021485.
5. Use of the exopolysaccharide EPS-a50 according to claim 4 for the preparation of a medicament for the prophylaxis and/or treatment of hypolipidemic conditions.
6. The use according to claim 5, characterized by the use in the preparation of a medicament for the prevention and/or treatment of hyperlipidemia.
7. Acidocella adephagiaBlastobotrys adeninivorans) A50 strain, characterized in that it comprisesBlastobotrys adeninivorans A50 strain is preserved in China Center for Type Culture Collection (CCTCC) at 2021, 04 and 29 days, and the preservation number is CCTCC NO: M2021485.
8. A microbial agent comprising the yeast of Arthrobacter adephagia of claim 7Blastobotrys adeninivorans A50 strain.
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| RU205304U1 (en) * | 2021-04-12 | 2021-07-08 | Федеральное государственное бюджетное образовательное учреждение высшего образования "Тульский государственный университет" (ТулГУ) | DEVICE FOR EXPRESS ANALYSIS OF WATER POLLUTION BY BIODEGRADABLE ORGANIC COMPOUNDS |
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