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CN-121975639-A - Bremia total shape, application thereof and preparation method of fungal chitosan

CN121975639ACN 121975639 ACN121975639 ACN 121975639ACN-121975639-A

Abstract

The invention discloses a Bremia total shape and application thereof, and a preparation method of fungus chitosan, belonging to the technical field of biology and application thereof. The strain is preserved in China general microbiological culture Collection center (CGMCC) No.42357, and the preservation date is 2025, 12 and 10. The strain is the Bremia total shape (LICHTHEIMIA RAMOSA) BT-0-A-001-JZ-001 separated from the brewing environment, and fungus chitosan with high purity and high deacetylation degree can be obtained by culturing. As an effective source of chitosan, the problem that the traditional chitosan extraction is excessively dependent on fishery resources is solved, and the problem that the raw materials for producing chitosan are limited is solved.

Inventors

  • CHEN LANG
  • MAO XIANG
  • He Hanyun
  • Ren Fanwen

Assignees

  • 成都柏腾医药科技有限公司

Dates

Publication Date
20260505
Application Date
20260407

Claims (9)

  1. 1. The Aureobasidium total is characterized in that Aureobasidium total BT-0-A-001-JZ-001 is preserved in China general microbiological culture Collection center (CGMCC) No.42357, and the preservation date is 2025, 12 and 10 days.
  2. 2. A process according to claim 1, wherein the Bremia totalis is in the form of living cells.
  3. 3. A process according to claim 2, wherein the Bremia totalis is in the form of a fungal medium, a lyophilized powder or a fermentation broth.
  4. 4. A process according to claim 3, wherein the Aureobasidium total is Aureobasidium total inclined plane and the culture medium is PDA culture medium.
  5. 5. The use of Bremia total body shape according to claim 1 to 4 for preparing fungal chitosan.
  6. 6. A method for preparing fungal chitosan, characterized in that the method is carried out by using the Brevibacterium racemosum according to any one of claims 1-4, and comprises the following steps: S1, preparing seed liquid, namely taking the Bremia total shape, taking 1-2 loops, inoculating the Bremia total shape to the inclined surface of a PDA test tube, culturing for 3-5 days at 28-35 ℃ to obtain an activated strain, preserving the activated strain in a refrigerator at-4~0 ℃ for standby, taking 2-3 loops of activated strain under the aseptic condition, inoculating the strain in a first liquid culture medium, and culturing for 2-3 days at 28-35 ℃ and 80-280 rpm to obtain the seed liquid; S2, fermenting and preparing mycelium, namely inoculating the seed liquid into a second liquid culture medium in an inoculum size of 3-10wt% under the aseptic condition, culturing for 4-9 days at 28-35 ℃ and 80-280 rpm, sieving the fermented liquid after the culturing is finished by a sieve with 50-100 meshes to obtain mycelium, adding deionized water with the same volume as the fermented liquid into the mycelium, washing the mycelium, filtering the mycelium by a sand core funnel, repeatedly washing the mycelium twice, and collecting and drying the mycelium at 40-50 ℃ to obtain dry mycelium of fungi; S3, deproteinizing the dry mycelium of the fungus, namely crushing the dry mycelium of the fungus, sieving the crushed dry mycelium of the fungus by a sieve with 50-60 meshes, collecting powder, adding a NaOH solution with the solid-to-liquid ratio of 1:20-1:40 (w/v), treating the mixture for 15-30 min at the temperature of 120-125 ℃, taking out the mixture, cooling the mixture for standby, putting the cooled liquid into a centrifuge tube, centrifuging the cooled liquid at 4500-6000 rpm for 3-15 min, taking out the supernatant, removing the supernatant, collecting the precipitate, adding deionized water for washing the precipitate, centrifuging the precipitate, repeatedly washing the precipitate until the liquid is neutral, and collecting the precipitate to obtain deproteinized mycelium; S4, extracting and purifying chitosan, namely adding glacial acetic acid solution with the concentration of 10% into deproteinized mycelium according to the dry weight of initial total-shaped aureobasidium in S2, wherein the dry weight is 1:20-1:40 (w/v), stirring for 6-12 hours at room temperature, centrifuging for 5-10 minutes at 4000-6000 rpm, collecting supernatant, regulating the pH to 10.0 by using 2mol/L NaOH solution, centrifuging for 3-10 minutes at 4000-6000 rpm, collecting precipitate, adding deionized water for washing for 1-2 times, centrifuging for 3-10 minutes at 4000-6000 rpm, collecting precipitate, adding acetone for washing for 1 time, centrifuging for 3-10 minutes at 4000-6000 rpm, collecting precipitate, and freeze-drying for 5-10 hours at-50-30 ℃ to obtain a fungal chitosan product.
  7. 7. The method of claim 6, wherein the first liquid medium in S1 is PDA liquid medium, 1000mL of PDA liquid medium comprises potato powder 5g and glucose 20g, and the sterilization is performed at 121 ℃ for 20min.
  8. 8. The method of claim 7, wherein the second liquid medium in S2 is identical to the first liquid medium in S1, and is the PDA liquid medium.
  9. 9. The method of claim 7, wherein the second liquid medium in S2 is a modified SDB liquid medium, 1000mL of the modified SDB liquid medium comprises 20g glucose, 5g yeast extract or yeast extract powder, 5g peptone, K 2 HPO 4 0.2~1g,(NH 4 ) 2 SO 4 0.5~2g,NaCl 0~0.1g and 0.1-0.5 g MgSO 4 •7H 2 O, and the modified SDB liquid medium is sterilized at 121 ℃ for 20min.

Description

Bremia total shape, application thereof and preparation method of fungal chitosan Technical Field The invention belongs to the technical field of biology and application thereof, and particularly relates to a Bremia total shape, application thereof and a preparation method of fungal chitosan. Background Chitosan (CS), also known as deacetylated chitin, is the deacetylated product of chitin, which is a white or pale yellow solid. The chitosan is taken as a natural cationic polysaccharide in the nature, has excellent biocompatibility, degradability, broad-spectrum antibacterial activity and other performances, and has great application potential in the fields of medicine, food, agriculture, environmental protection and the like. At present, chitosan is mainly extracted from shellfish processing wastes such as shrimps, crabs and the like, the production raw materials of the chitosan are seriously dependent on fishery resources and limited by seasons and regions, a large amount of strong acid and strong alkali are used in the extraction process, and the problems of high equipment requirements, serious environmental pollution and the like exist. The use of microbial fermentation to produce chitosan is considered as a very promising alternative. The chitosan produced by fungus fermentation has the remarkable advantages of wide raw material sources, controllable fermentation process, mild extraction conditions and the like, and is more and more paid attention to. In the prior art, chinese patent CN 120290331A discloses an aspergillus niger, application thereof and a preparation method of chitosan, wherein the method is used for extracting chitin after fermentation of the aspergillus niger and deacetylating the obtained chitin to obtain the chitosan. The method still uses NaOH aqueous solution with the concentration of 40-60wt% for deacetylation, has high alkali concentration, has high equipment requirement and is easy to cause environmental pollution. Chinese patent CN106243245B provides a preparation method of chitosan, which is characterized in that the chitosan is obtained from insect crustaceans through calcium removal, deproteinization, purification and deacetylation, and the operation is simple and convenient, and the equipment is conventional. However, the method still extracts chitin from the crust and obtains chitosan after deacetylation, and the problems of limited raw materials and the like are not eliminated. Disclosure of Invention The invention aims to overcome the defects of the prior art and provides a Bremia total shape, application thereof and a preparation method of chitosan. The aim of the invention is realized by the following technical scheme: The first aspect of the invention is to provide a Bremia total shape (LICHTHEIMIA RAMOSA) BT-0-A-001-JZ-001 which is preserved in China general microbiological culture Collection center with the preservation number of CGMCC No.42357 and the preservation date of 2025, 12 months and 10 days. The invention provides a Bremia total shape which is in a living cell form, for example, a strain is in a fungus culture medium form, a freeze-dried powder form or a fermentation broth form. The lyophilized powder form is generally a long-term preserved form of the strain. Preferably, when the strain is in the form of a fungal medium, the Bremia total is a Bremia total slant and the medium is PDA medium. The second aspect of the invention is to provide an application of the Bremia total shape for preparing fungal chitosan. The third aspect of the present invention is to provide a method for preparing fungal chitosan, which is a method for preparing the fungus chitosan by using the Bremia total shape (LICHTHEIMIA RAMOSA) BT-0-A-001-JZ-001, and comprises the following steps: S1, preparing seed liquid, namely taking the Bremia total shape, taking 1-2 loops, inoculating the Bremia total shape to the inclined surface of a PDA test tube, culturing for 3-5 days at 28-35 ℃ to obtain an activated strain, preserving the activated strain in a refrigerator at-4~0 ℃ for standby, taking 2-3 loops of activated strain under the aseptic condition, inoculating the strain in a first liquid culture medium, and culturing for 2-3 days at 28-35 ℃ and 80-280 rpm to obtain the seed liquid. S2, fermenting to prepare mycelium, namely inoculating the seed liquid into a second liquid culture medium in an inoculum size of 3-10wt% under the aseptic condition, culturing for 4-9 days at 28-35 ℃ and 80-280 rpm, sieving the fermented liquid after the culturing is finished by a sieve with 50-100 meshes to obtain mycelium, adding deionized water with the same volume as the fermented liquid into the mycelium, washing the mycelium, filtering the mycelium by a sand core funnel, repeatedly washing the mycelium twice, and collecting and drying the mycelium at 40-50 ℃ to obtain dry fungus mycelium. S3, deproteinizing the dry mycelium of the fungus, namely crushing the dry mycelium of the fungus, sieving the cr