CN110846259B - High-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide and application thereof - Google Patents
High-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide and application thereof Download PDFInfo
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Abstract
The invention discloses a high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide and application thereof, wherein the taxonomic name of the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide is Microbacterium testaceum EBPDAK006, and the preservation number is CCTCC NO: and M2018785. The high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide provided by the invention can be used for obtaining the Hubei ophiopogon japonicus polysaccharide through liquid fermentation, can effectively solve the problem of difficult production of the Hubei ophiopogon japonicus polysaccharide, and can be used for rapidly, industrially and massively producing the Hubei ophiopogon japonicus polysaccharide through a microbial fermentation mode.
Description
Technical Field
The invention relates to the technical field of microorganisms, and particularly relates to a high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide and application thereof.
Background
The Hubei radix ophiopogonis polysaccharide is one of main active ingredients of a national geographical sign product and a land Chinese medicinal plant-Hubei radix ophiopogonis Lir iope spicata var. prolifera Y.T.Ma tuberous root, and researches show that: the Hubei ophiopogon root polysaccharide has pharmacological effects of resisting myocardial ischemia, resisting thrombosis, resisting anoxia, resisting aging, reducing blood sugar and the like, has positive effects on the aspects of improving the immune system of an organism, enhancing the gastrointestinal motility and the like, and has an anti-tumor effect according to latest research.
The traditional Hubei radix ophiopogonis polysaccharide production is mainly obtained by directly separating the root tuber extract of Hubei radix ophiopogonis. However, in the actual planting production of ophiopogon japonicus in Hubei, factors such as relatively small planting area caused by continuous cropping obstacle, large influence of climatic conditions, long root tuber production period, high polysaccharide extraction cost and the like exist, so that the market release of ophiopogon japonicus polysaccharide in Hubei is limited. At present, no endophytic bacterium capable of producing Hubei ophiopogon japonicus polysaccharide through microbial fermentation and application thereof are reported.
Disclosure of Invention
The invention mainly aims to provide a high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide, aiming at improving the yield of the Hubei ophiopogon japonicus polysaccharide through microbial fermentation.
In order to achieve the purpose, the invention provides a high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide, which is classified and named as Microbacterium terreus (EBPDAK 006) with the preservation number of CCTCC NO: and M2018785.
Alternatively, the 16S rRNA gene sequence of the high-producing bacterial strain of the Hubei ophiopogon japonicus polysaccharide is shown in SE Q ID NO. 1.
The method for separating the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide comprises the following steps:
cleaning fresh roots of the Hubei dwarf lilyturf tuber, soaking the roots in ethanol for 1-3 min, rinsing the roots with water for 2-4 times, soaking the roots in a sodium hypochlorite solution for 1-3 min, rinsing the roots with water for 3-5 times, and dipping sterile filter paper in the roots to obtain a sterilized sample;
cutting off two ends of browning parts of the sterilized sample, taking root tuber tissue blocks, adding water, grinding to obtain homogenate, diluting the homogenate, taking the suspension, coating a PDA culture medium flat plate, sealing by using a sealing film, and then culturing at a constant temperature of 37 ℃;
and (3) selecting white and round colonies with good growth and smooth surfaces, transferring the colonies into a new PDA culture medium plate, and respectively carrying out streaking and purification culture for multiple times to finally obtain the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide.
The invention also provides an application of the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide, and the Hubei ophiopogon japonicus polysaccharide is prepared by adopting the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide through liquid fermentation.
The preparation of the Hubei ophiopogon japonicus polysaccharide by fermentation specifically comprises the following steps:
inoculating the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide into a PDA culture medium test tube, and performing activated culture to obtain an activated strain;
inoculating the activated strain into a seed culture medium, and performing shake culture to obtain seeds;
inoculating the seeds into a liquid fermentation culture medium, and performing shaking culture to obtain a fermentation mixture;
quickly freezing the fermentation mixture into a solid state, and performing low-temperature freeze drying treatment to obtain a fermentation product;
grinding the fermentation product into powder, washing the crushed material for multiple times by using ethanol, and drying to obtain a dried sample;
adding the dried sample into water, heating in water bath, filtering, repeating the above steps again on filter residues, combining filtrates, centrifuging, and dialyzing supernatant to obtain dialysate;
concentrating the dialysate under reduced pressure to obtain extract, and precipitating the extract with ethanol to obtain ethanol precipitation product;
washing the alcohol precipitation product with absolute ethyl alcohol, acetone and ethyl ether in sequence, drying to obtain Hubei ophiopogon japonicus polysaccharide, and storing at low temperature.
The high-yield bacterial strain of Hubei radix ophiopogonis polysaccharide provided by the invention can be used for producing Hubei radix ophiopogonis polysaccharide after liquid fermentation, has the advantages of rapidness, factory production and scale production, can avoid the pollution to the environment caused by chemical synthesis, can avoid various adverse factors in the planting production of Hubei radix ophiopogonis medicinal materials, and has great application value.
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In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other related drawings can be obtained according to the drawings without creative efforts.
FIG. 1 is a morphological diagram of Microbacterium rubrum on PDA medium in example 1 of the present invention;
FIG. 2(a) is an HPLC detection chart of acid hydrolyzed monosaccharide of polysaccharide extracted from Hubei radix Ophiopogonis tuber;
FIG. 2(b) is a HPLC detection chart of acid hydrolysis monosaccharides in polysaccharide extracts from fermentation broths of strains prepared in example 2 of the present invention;
FIG. 3 shows OH and O pairs of Hubei Ophiopogon japonicus polysaccharides in polysaccharide extract from fermentation broth of the strain prepared in example 2 of the present invention2-DPPH, ABTS radical scavenging activity.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.
The invention provides a high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide, which is classified and named as Microbacterium tetakeum EBPD AK006, and the preservation number is CCTCC NO: m2018785, wherein the preservation date is 11/12 in 2018, the preservation unit is China Center for Type Culture Collection (CCTCC), and the preservation address is China, Wuhan university.
In the practical application process, for the reasons of convenient transportation and the like, it is necessary to enlarge the application range of the Hubei radix ophiopogonis polysaccharide high-yield bacterial strain in the form of a composition (particularly a microbial agent) by culture enlargement. The invention also provides a composition containing the culture of the high-producing bacterial strain of Hubei ophiopogon japonicus polysaccharide.
Further, the composition is in the form of a liquid, frozen or dried powder.
The preparation is expressed in the form of preparation commonly used in the industry, such as granules, suspending agents, wettable powder, emulsion or liquid.
The invention also provides a separation method of the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide, and in an embodiment of the separation method of the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide provided by the invention, the separation method of the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide comprises the following steps:
step S10, cleaning fresh roots of the Hubei dwarf lilyturf tuber, soaking the roots in ethanol for 1-3 min, rinsing the roots with water for 2-4 times, soaking the roots in a sodium hypochlorite solution for 1-3 min, rinsing the roots with water for 3-5 times, and dipping the roots in sterile filter paper to obtain a sterile sample.
In specific implementation, a large amount of tap water is used for fully cleaning soil on the surfaces of fresh roots of the liriope spicata plants, then the fresh roots of the liriope spicata plants are put into a clean beaker, deionized water is added, and the beaker is placed in an ultrasonic cleaner for repeated cleaning until the water after cleaning becomes extremely clear; after the water on the surface of the dry radix ophiopogonis root tuber is dipped by sterile filter paper, the treatment is carried out according to the following steps: firstly, soaking the root tuber in 75% (V/V) ethanol for 2min, and rinsing the root tuber with sterile water for 2-4 times; and then soaking the fabric in a sodium hypochlorite solution with the effective chlorine content of 4-6% for 2min, continuously rinsing the fabric for 3-5 times by using a large amount of sterile water, and dipping the fabric in dry water by using sterile filter paper.
And step S20, cutting off two ends of the browning part of the sterilized sample, taking root tuber tissue blocks, adding water, grinding to obtain homogenate, diluting the homogenate, taking the suspension, coating a PDA culture medium flat plate, sealing by a sealing film, and culturing at constant temperature of 37 ℃.
In specific implementation, the tuber root sample with the surface disinfected is taken, brown stain parts at two ends of the tuber root are cut off under the aseptic condition, about 1.0g of tuber root tissue block is weighed, 5mL of sterile water is added for full grinding, and 0.5m of tuber root tissue block is suckedL homogenate is diluted to 10-3And then coating the suspension on a PDA culture medium plate, sealing the PDA culture medium plate by using a parafilm sealing film, and then culturing at the constant temperature of 37 ℃ for 12-18 h.
And S30, selecting a colony which grows well, is white and round, has a flat and smooth surface, and is transferred to a new PDA culture medium flat plate, and performing streaking and purification culture for multiple times respectively to finally obtain the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide.
The invention also provides an application of the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide, and the Hubei ophiopogon japonicus polysaccharide is prepared by adopting the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide through liquid fermentation. Wherein the liquid fermentation culture medium is a PDB culture medium, and the formula of the liquid fermentation culture medium is as follows: 200g of potato, 20g of glucose and 1000mL of distilled water. In an embodiment of the application of the high-yield bacterial strain of ophiopogon japonicus polysaccharide in Hubei of the invention, the method comprises the following steps:
inoculating the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide into a PDA culture medium test tube, and performing activated culture to obtain an activated strain;
inoculating the activated strain into a seed culture medium, and performing shake culture to obtain seeds;
inoculating the seeds into a liquid fermentation culture medium, and performing shaking culture to obtain a fermentation mixture;
quickly freezing the fermentation mixture into a solid state, and performing low-temperature freeze drying treatment to obtain a fermentation product;
grinding the fermentation product into powder, washing the crushed material for multiple times by using ethanol, and drying to obtain a dried sample;
adding the dried sample into water, heating in water bath, filtering, repeating the above steps again on the filter residue, combining the filtrates, centrifuging, and dialyzing the supernatant of the centrifugate to obtain a retention solution;
concentrating the reserved solution under reduced pressure to obtain an extract, and then carrying out alcohol precipitation on the extract by using ethanol to obtain an alcohol precipitation product;
washing the alcohol precipitation product with absolute ethyl alcohol, acetone and ether in sequence, drying to obtain a strain fermentation liquid polysaccharide extract, namely Hubei ophiopogon japonicus polysaccharide, and storing at low temperature.
In specific implementation, the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide is taken, a small number of bacterial colonies are picked by an inoculating needle under the aseptic condition, and the bacterial colonies are inoculated into a sterilized PDA culture medium test tube and subjected to activation culture at 37 ℃ for 16 hours; taking the activated and cultured strain, transferring the strain into a sterilized liquid potato seed culture medium under an aseptic condition, and carrying out shaking culture at 37 ℃ and 180rpm for 16 hours to obtain seeds; respectively filling the prepared liquid fermentation culture medium into 250mL triangular flasks, wherein each flask is about 100mL, sterilizing, and cooling for later use; inoculating seeds according to the inoculation amount of 10% under the aseptic condition, performing shaking culture at 37 ℃ and 180rpm for 2 days, and fermenting; placing the fermented solid-liquid mixture in an ultra-low temperature refrigerator for quick freezing into a solid state, and performing low-temperature freeze drying treatment by using a small freeze dryer to obtain a dried fermented product; grinding the strain fermentation product freeze-dried at low temperature into powder, respectively adding 100mL of 95% ethanol repeatedly for three times, washing the ground material, and drying; adding distilled water into the dried sample according to a ratio of 1: 10(g/mL), carrying out water bath at 80 ℃ for 2h, filtering, repeating the operation on the filter residue once again, combining the filtrate, centrifuging, taking the supernatant, and dialyzing for 48 h; concentrating the dialyzed retention solution under reduced pressure to obtain extract, and then carrying out alcohol precipitation on the extract by using 100mL of 95% ethanol; washing the alcohol precipitation product with absolute ethanol, acetone and diethyl ether in sequence, drying to obtain the polysaccharide extract of the strain fermentation liquor, and storing at 4 ℃ for later use.
Example 1 isolation procedure of high-producing bacterial strains of Hubei Ophiopogon japonicus polysaccharides
(1) Sampling: the GAP base of the Hubei radix ophiopogonis, which is the original producing area of the Hubei radix ophiopogonis in the town of Ozily Temple of Xiangyang, Hubei province, is respectively sampled at multiple points to be used as a plant raw material for separating the high-yield bacterial strains of the Hubei radix ophiopogonis polysaccharide.
(2) Preparing a culture medium:
the formula of the PDA culture medium is as follows: 200g of potato, 20g of glucose, 15-20 g of agar and 1000ml of distilled water
(3) Strain separation: cleaning soil on root tuber surface of fresh Hubei radix Ophiopogonis plant with large amount of tap water, placing into clean beaker, adding deionized water, and placing into ultrasonic cleanerRepeatedly cleaning until the water after cleaning becomes extremely clear; after the water on the surface of the tuber root of ophiopogon japonicus in north of dry lake is dipped by sterile filter paper, the treatment is carried out according to the following steps: soaking root tuber in 75% (V/V) ethanol for 2min, and rinsing with sterile water for 3 times; soaking for 2min by using a sodium hypochlorite solution with the effective chlorine content of 4-6%, continuously rinsing for 4 times by using a large amount of sterile water, and dipping the water in sterile filter paper; taking the radix Ophiopogonis Hubei with surface disinfected, cutting off brown stain parts at two ends of the root tuber under aseptic condition, weighing about 1.0g of root tuber tissue block, adding 5mL of sterile water, grinding thoroughly, sucking 0.5mL of homogenate, diluting to 10%-3Then coating the suspension on a PDA culture medium plate, sealing with a parafilm sealing film, and culturing at 37 deg.C for 12-18 h; and (3) selecting a colony which is white and round and has a flat and smooth surface, transferring the colony into a new PDA culture medium plate, and respectively carrying out streaking and purification culture for multiple times to finally obtain the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide.
Example 2 preparation of Hubei Ophiopogon japonicus polysaccharides
The high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide obtained in example 1 is taken, a small number of colonies are picked up by an inoculating needle under the aseptic condition, and the colonies are inoculated into a sterilized PDA culture medium test tube and activated and cultured for 16 hours at 37 ℃.
Taking the activated and cultured strain, transferring the strain into a sterilized liquid potato seed culture medium under the aseptic condition, and carrying out shaking culture at 37 ℃ and 180rpm for 16 hours to obtain the seed.
Respectively filling the prepared liquid fermentation culture medium into 250mL triangular flasks, wherein each flask is about 100mL, sterilizing, and cooling for later use; under aseptic conditions, the seeds were inoculated at 10% inoculum size and cultured with shaking at 37 ℃ and 180rpm for 2 days.
After the fermentation is finished, the fermentation solid-liquid mixture is placed in an ultra-low temperature refrigerator to be frozen into a solid state quickly, and then a small freeze dryer is used for low-temperature freeze drying treatment, so that a dried fermentation product is obtained.
Grinding the strain fermentation product freeze-dried at low temperature into powder, respectively adding 100mL of 95% ethanol repeatedly three times, washing the ground product, and oven-drying.
Adding distilled water into the oven-dried sample at a ratio of 1: 10(g/mL), water bathing at 80 deg.C for 2 hr, filtering, repeating the above operation for 1 time, mixing filtrates, centrifuging, collecting supernatant, and dialyzing for 48 hr.
Collecting the dialyzed retention solution, concentrating under reduced pressure to obtain extract, and precipitating with 100mL 95% ethanol.
Washing the alcohol precipitation product with absolute ethyl alcohol, acetone and ether in sequence, drying to obtain the polysaccharide extract of the strain fermentation liquor, and storing at 4 ℃ for later use.
Example 3 identification of Strain and detection of Hubei Ophiopogon japonicus polysaccharide produced by Strain
(1) Morphological characteristic identification of strain culture medium
The high-producing bacterial strain of Hubei ophiopogon japonicus polysaccharide obtained in example 1 was streaked on a PDA medium plate, cultured at 28 ℃ for 24 hours, and observed for the characteristics of colony shape, size, color, and the like. As shown in FIG. 1, the colonies of the high-producing bacterial strain of ophiopogonpolysaccharide in Hubei are white, round and smooth.
(2) 16S rRNA gene sequencing of high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide
The 16Sr RNA gene sequence of the high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide is shown in an attachment, and the sequence comparison is carried out on the sequence comparison result at NCBI website (http:// blast. NCBI. nlm. nih. gov/blast. cgi), wherein the homology with Microbacterium testaceum. MH590622.1 is 99%.
(3) Color reaction of fermentation product
The following two color reaction methods are adopted for detection:
(a) the naphthol-concentrated sulfuric acid reaction method comprises the following steps: 200 mul of the polysaccharide extract sample of the strain fermentation liquor is dissolved in 400 mul of 15 percent 1-naphthol, and 100 mul of concentrated sulfuric acid is added, and the sample is positive with blue-purple.
(b) The anthrone-concentrated sulfuric acid reaction method comprises the following steps: 400 mul of polysaccharide extract sample of the strain fermentation liquor is taken and added with 300 mul of anthrone sulfuric acid solution, and the sample shows blue green as positive.
The results of both reactions of the fermentation product were positive. Namely, the bacterial strain with high yield of the Hubei ophiopogon polysaccharide can produce the polysaccharide through fermentation.
(4) Thin layer chromatography detection (TLC)
Taking 20mg of the strain fermentation broth polysaccharide extract prepared in the example 2, adding 5mL of pure water for dissolving, then adding 0.25mL of HCL with the concentration of 12mol/L, placing the solution in an environment with the temperature of 120-126 ℃ for reacting for 4h, and diluting to 10mL after neutralization and dialysis to obtain the acidolysis solution of the strain fermentation broth polysaccharide extract.
Performing thin layer chromatography on acidolysis solution of polysaccharide extract of strain fermentation broth by taking acidolysis product of polysaccharide extracted from Hubei radix Ophiopogonis tuber as positive control. The developing solvent is n-butyl alcohol: anhydrous ethanol: glacial acetic acid: pure water is 2:3:1:1(V/V/V/V), the developer is prepared by weighing 4g of diphenylamine, 4mL of aniline and 20mL of 85% phosphoric acid, dissolving in 200mL of acetone, and heating in an oven at 105 ℃ for 3min until clear red spots appear.
The results show that the acidolysis solution of the polysaccharide extract of the strain fermentation broth contains an Rf value similar to that of the positive control. Namely, the high-yield bacterial strain fermentation of the Hubei ophiopogon japonicus polysaccharide can produce the Hubei ophiopogon japonicus polysaccharide.
(5) High performance liquid chromatography detection (HPLC)
Taking 20mg of the strain fermentation broth polysaccharide extract prepared in the example 2, adding 5mL of pure water for dissolving, then adding 0.25mL of HCL with the concentration of 12mol/L, placing the solution in an environment with the temperature of 120-126 ℃ for reacting for 4h, and diluting to 10mL after neutralization and dialysis to obtain the acidolysis solution of the strain fermentation broth polysaccharide extract.
Placing 1mL of the acidolysis solution of the prepared strain fermentation broth polysaccharide extract in a centrifuge tube, adding 1mL of 0.5mol/mL PMP-methanol solution and 0.3 mol/mL LNaOH solution, vortexing for 1min, cooling to room temperature in 70 ℃ water bath for 60min, neutralizing 1mL of 0.3mol/L HCL solution, extracting with 2mL of chloroform for 3 times, combining water layers, filtering with a water system filter membrane, and performing ultrasonic treatment for 0.5h before use. Meanwhile, the acidolysis product of the polysaccharide extracted from the Hubei radix ophiopogonis tuber is taken, and the operation is repeated to complete the preparation of the positive control sample.
HPLC chromatographic conditions
Shimadzu LC-20 high performance liquid chromatograph (detector: SPD-M20A, column oven: CTO-20A, vacuum degasser DGU-20A3, chromatographic column Inert Sustain C18); utilizing acetonitrile: isocratic elution with phosphate buffer (pH 5.0) 20:80 at a flow rate of 1 mL/min; spectral data acquisition channel: ch1(250 nm); the sample volume is 10 mu L; recording data for 30 min; the signal strength is in units of m AU.
FIG. 2(a) is an HPLC detection chart of acid-hydrolyzed monosaccharide of polysaccharide extracted from radix Ophiopogonis Hubei tuber as shown in FIG. 2(a) and FIG. 2 (b); FIG. 2(b) is a HPLC check chart of acid hydrolysis monosaccharide in polysaccharide extract from fermentation broth of the strain prepared in example 2 of the present invention. The comparison of the two figures shows that the peaks are relatively consistent, which indicates that the high-yield bacterial strain fermentation of the Hubei ophiopogon japonicus polysaccharide can produce the Hubei ophiopogon japonicus polysaccharide.
(6) Antioxidant activity detection method
Respectively testing OH and O pairs of polysaccharide extracts in strain fermentation liquor by adopting an alpha-deoxyribose method, a pyrogallol autoxidation method, a DPPH method and an ABTS method2-DPPH, ABTS radical scavenging activity.
FIG. 3 shows the polysaccharide extract pair OH and O in the fermentation broth of the strain prepared in example 22-DPPH, ABTS radical scavenging activity. As can be seen from FIG. 3, the polysaccharide extract from the fermentation broth of the high-yielding bacterial strain of ophiopogon japonicus polysaccharide in Hubei has a certain in vitro scavenging activity on the four free radicals, and the antioxidant ability is positively correlated with the concentration.
In conclusion, the bacterial colony of the high-yield bacterial strain of the Hubei ophiopogon japonicus polysaccharide provided by the invention is white and round, and the surface is flat and smooth; homology to Microbacterium testaceum. mh590622.1 is 99%; the two color reaction results are positive, the acidolysis monosaccharide of the strain fermentation broth polysaccharide extract has similar Rf value with the positive control acidolysis monosaccharide, and HPLC comparison maps show that the high-yield bacterial strain fermentation of the Hubei ophiopogon japonicus polysaccharide can produce the Hubei ophiopogon japonicus polysaccharide; the Hubei radix ophiopogonis polysaccharide produced by fermentation has certain in-vitro removal activity, which shows that the high-yield bacterial strain of the Hubei radix ophiopogonis polysaccharide has good performance and can be widely applied to the production of the Hubei radix ophiopogonis polysaccharide.
The above is only a preferred embodiment of the present invention, and it is not intended to limit the scope of the invention, and various modifications and changes will occur to those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present invention shall be included in the scope of the present invention.
SEQUENCE LISTING
<110> Hubei academy of culture and management
<120> high-yield bacterial strain of Hubei ophiopogon japonicus polysaccharide and application thereof
<130> 2019.12.13
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1486
<212> DNA
<213> Microbacterium testaceum
<400> 1
tacggttacc ttgttacgac ttagtcctaa ttaccgatcc caccttcgac ggctccctcc 60
acaagggttg ggccaccggc ttcaggtgtt accgactttc atgacttgac gggcggtgtg 120
tacaagaccc gggaacgtat tcaccgcagc gttgctgatc tgcgattact agcgactccg 180
acttcatgag gtcgagttgc agacctcaat ccgaactggg accggctttt tgggattcgc 240
tccacctcac ggtattgcag ccctttgtac cggccattgt agcatgcgtg aagcccaaga 300
cataaggggc atgatgattt gacgtcatcc ccaccttcct ccgagttgac cccggcagta 360
tcccatgagt tcccaccatt acgtgctggc aacatagaac gagggttgcg ctcgttgcgg 420
gacttaaccc aacatctcac gacacgagct gacgacaacc atgcaccacc tgttcaccag 480
tgtccaaaga gttgaccatt tctggcccgt tctggtgtat gtcaagcctt ggtaaggttc 540
ttcgcgttgc atcgaattaa tccgcatgct ccgccgcttg tgcgggtccc cgtcaattcc 600
tttgagtttt agccttgcgg ccgtactccc caggcgggga acttaatgcg ttagctgcgt 660
cacggaaacc gtggaatggt ccccacaact agttcccaac gtttacgggg tggactacca 720
gggtatctaa gcctgtttgc tccccaccct ttcgctcctc agcgtcagtt acggcccaga 780
gatctgcctt cgccatcggt gttcctcctg atatctgcgc attccaccgc tacaccagga 840
attccaatct cccctaccgc actctagtct gcccgtaccc actgcaggcc cgaggttgag 900
cctcgggatt tcacagcaga cgcgacaaac cgcctacgag ctctttacgc ccaataattc 960
cggataacgc ttgcgcccta cgtattaccg cggctgctgg cacgtagtta gccggcgctt 1020
tttctgcagg taccgtcact ttcgcttctt ccctgctaaa agaggtttac aacccgaagg 1080
ccgtcatccc tcacgcggcg ttgctgcatc aggctttcgc ccattgtgca atattcccca 1140
ctgctgcctc ccgtaggagt ctgggccgtg tctcagtccc agtgtggccg gtcaccctct 1200
caggccggct acccgtcgac gccttggtga gccattacct caccaacaag ctgataggcc 1260
gcgagcccat cccagaccaa aaaatctttc caacccccac catgcgatga gagctcatat 1320
ccagtattag acgccgtttc cagcgcttat cccagagtcc agggcaggtt gctcacgtgt 1380
tactcacccg ttcgccactg atccaccaag caagcttggc ttcaccgttc gacttgcatg 1440
tgttaagcac gccgccagcg ttcatcctga gccaggatca aactct 1486
Claims (2)
1. The bacterial strain for producing the Hubei dwarf lilyturf tuber polysaccharide is characterized in that the taxonomy of the bacterial strain for producing the Hubei dwarf lilyturf tuber polysaccharide is named as Microbacterium terreus (EBPD AK 006), and the preservation number of the bacterial strain is CCTCC NO: and M2018785.
2. The use of the bacterial strain producing ophiopogonpolysaccharide from Hubei province as claimed in claim 1, wherein the Hubei ophiopogonpolysaccharide is obtained by liquid fermentation.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2012145946A1 (en) * | 2011-04-25 | 2012-11-01 | 上海张江中药现代制剂技术工程研究中心 | Application of dwarf lilyturf tuber polysaccharide extract in preparation of dietary supplement, health food or medicine with the function of weight loss |
| CN107739718A (en) * | 2017-10-18 | 2018-02-27 | 湖北文理学院 | It is a kind of to assist raw Eurotium fungi and the application in steroid saponin is prepared in the tuber of dwarf lilyturf |
| CN108553480A (en) * | 2018-06-11 | 2018-09-21 | 上海中医药大学 | Application of the ophiopogonpolysaccharide extract in preparing the drug with anti-inflammatory effect |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2012145946A1 (en) * | 2011-04-25 | 2012-11-01 | 上海张江中药现代制剂技术工程研究中心 | Application of dwarf lilyturf tuber polysaccharide extract in preparation of dietary supplement, health food or medicine with the function of weight loss |
| CN107739718A (en) * | 2017-10-18 | 2018-02-27 | 湖北文理学院 | It is a kind of to assist raw Eurotium fungi and the application in steroid saponin is prepared in the tuber of dwarf lilyturf |
| CN108553480A (en) * | 2018-06-11 | 2018-09-21 | 上海中医药大学 | Application of the ophiopogonpolysaccharide extract in preparing the drug with anti-inflammatory effect |
Non-Patent Citations (2)
| Title |
|---|
| 湖北麦冬多糖含量的测定;王晓华等;《湖北中医杂志》;20050325;第27卷(第3期);第50-51页 * |
| 襄麦冬多糖及皂苷体外活性研究;陈哲等;《食品工业科技》;20180731;第39卷(第13期);第7-12页 * |
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