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CN-121975888-A - Ulva pertusa polysaccharide and uniform polysaccharide fermented by bacillus bailii, and preparation methods and applications thereof

CN121975888ACN 121975888 ACN121975888 ACN 121975888ACN-121975888-A

Abstract

The invention discloses a method for preparing ulva pertusa polysaccharide by co-fermenting bacillus bailii SCAU-104 and ulva pertusa algae powder and application thereof. Compared with the traditional hot water extraction of polysaccharide (UPP), the polysaccharide (UBV) obtained by fermentation extraction has lower molecular weight, higher sulfate group retention and more optimized structure. Removing protein from UBV by macroporous resin, ultrafiltering to desalt, purifying by chromatography, and separating to obtain uniform polysaccharide UBV-1. Research shows that UBV is mainly composed of rhamnose, galactose and glucuronic acid, and is acidic sulfated polysaccharide with single molecular weight. In vitro experiments show that UBV significantly promotes RAW264.7 macrophage proliferation and NO and pro-inflammatory factor release, and the immunocompetence is superior to UPP. The method is green and controllable, and the obtained polysaccharide has high purity and strong activity, and can be widely applied to development of functional foods, cosmetics, skin care products and immunoregulation products.

Inventors

  • HUANG RIMING
  • ZHANG RONGXIN
  • ZHENG QIANWANG
  • Yu Jieting
  • ZHANG LIN
  • ZHANG XIAOYONG
  • HOU QILI
  • MO YONGHUI
  • HUANG YAONAN
  • LIU QIN
  • GAO JIEXIN

Assignees

  • 华南农业大学

Dates

Publication Date
20260505
Application Date
20251223

Claims (10)

  1. 1. A preparation method of ulva pertusa polysaccharide fermented by bacillus bailii is characterized by comprising the following steps: (1) Performing strain activation on bacillus belicus Bacillus velezensisSCAU-104 to obtain seed liquid, wherein bacillus belicus SCAU-104 strain is from the Guangdong province microorganism strain collection center, and the collection number is GDMCCNo.64045; (2) Inoculating seed liquid of bacillus belay SCAU-104 into a culture medium containing ulva pertusa algae powder, and fermenting at 25-37 ℃ for 24-96 hours to obtain a fermentation liquid product; (3) Filtering, concentrating, centrifuging to remove precipitate, removing protein from supernatant with macroporous resin, collecting filtrate, dialyzing to obtain UBV, and lyophilizing to obtain UBV powder.
  2. 2. The method for preparing ulva pertusa polysaccharide fermented by bacillus clarias according to claim 1, wherein the method comprises any one or more of the following characteristics of i) -iii): i) The inoculation amount of the bacillus bailii SCAU-104 seed solution is 1-5% v/v of the volume of the culture medium; ii) the weight percentage content of the ulva pertusa algae powder in the culture medium is 5-10 g/L; iii) In the dialysis step, a dialysis bag with the molecular weight cut-off of 8000-15000 Da is used.
  3. 3. The ulva pertusa polysaccharide UBV prepared by the method for preparing ulva pertusa polysaccharide by fermentation of bacillus bailii according to claim 1 or 2.
  4. 4. The UBV of Ulva pertusa polysaccharide according to claim 3, wherein the Ulva pertusa polysaccharide is mainly composed of rhamnose, glucuronic acid, xylose, galactose and glucosamine, and the molar ratio is in the range of (27% -30%) (22% -25%) (1.6% -1.8%) (38% -43%) (0.7% -0.8%).
  5. 5. The UBV of Ulva pertusa polysaccharide according to claim 3, wherein the Ulva pertusa polysaccharide has a total sugar content of 61% -72%, a sulfate group content of 24% -26%, The UBV consists of polysaccharide with molecular weight of 269239 and 86.67 percent and polysaccharide with molecular weight of 5046 and 13.33 percent; The molar ratio of the rhamnose to the glucuronic acid to the xylose to the galactose to the glucosamine is 40.59 percent, 28.63 percent, 23.47 percent, 4.87 percent and 0.74 percent.
  6. 6. Use of ulva pertusa polysaccharide UBV according to claim 3 for the preparation of a modulating immune formulation.
  7. 7. The ulva pertusa uniform polysaccharide fermented by bacillus bailii is characterized by having a structure shown in the following general formula: the ulva pertusa uniform polysaccharide mainly comprises rhamnose, glucuronic acid and xylose, and the molar ratio is 47.6%, 32.1% and 20.3% respectively.
  8. 8. The Ulva pertusa uniform polysaccharide fermented by Bacillus clathratus according to claim 7, wherein the maximum peak molecular weight of the Ulva pertusa uniform polysaccharide is 208564 daltons.
  9. 9. The method for preparing ulva pertusa uniform polysaccharide fermented by bacillus clarkii according to claim 7, which is characterized by comprising the following steps: (1) Performing strain activation on bacillus belicus Bacillus velezensis SCAU-104 to obtain seed liquid, wherein bacillus belicus SCAU-104 strain is from the Guangdong province microorganism strain collection center, and the collection number is GDMCCNo.64045; (2) Inoculating seed liquid of bacillus belay SCAU-104 into a culture medium containing ulva pertusa algae powder, and fermenting at 25-37 ℃ for 24-96 hours to obtain a fermentation liquid product; (3) Filtering, concentrating, centrifuging to remove precipitate, removing protein from supernatant with macroporous resin, collecting filtrate, dialyzing to obtain UBV, and lyophilizing to obtain UBV powder; (4) UBV is ultrafiltered, and UBV-1 is obtained by retaining the upper part of the membrane.
  10. 10. The method for preparing ulva pertusa uniform polysaccharide by bacillus clarias fermentation according to claim 9, wherein the ultrafiltration step is carried out through an ultrafiltration membrane with the molecular weight cutoff of 8000-15000 Da.

Description

Ulva pertusa polysaccharide and uniform polysaccharide fermented by bacillus bailii, and preparation methods and applications thereof Technical Field The invention relates to the technical field of polysaccharides, in particular to ulva lactuca polysaccharide prepared by co-fermenting bacillus bailii and ulva lactuca algae powder, and specifically relates to a fermented polysaccharide obtained by fermenting bacillus bailii SCAU-104 serving as a strain in a culture medium containing the ulva lactuca algae powder. The homogeneous polysaccharide UBV-1 is obtained through separation and purification, and the main structural characteristics are clarified through methylation and nuclear magnetic analysis. The invention also relates to the application of the fermented polysaccharide in functional foods and immunomodulators. Background Ulva pertusa (Ulva lactuca) belongs to the phylum Chlorophyta, commonly known as sea lettuce or sea lettuce, and is a common marine alga in the coastal intertidal zone. The algae is rich in nutrients and active ingredients such as polysaccharide, protein, vitamins, etc. The ulva pertusa polysaccharide is taken as sulfated polysaccharide, has remarkable physiological activities of immunoregulation, antioxidation, anticoagulation and the like, and has wide application prospect in the fields of health-care food, biological medicine and daily chemical products. At present, the ulva pertusa polysaccharide is mainly prepared by a traditional hot water leaching method. The method has the advantages of simple operation and low equipment investment, but has obvious technical limitations that the obtained polysaccharide has generally higher molecular weight and poorer solubility, is difficult to be effectively absorbed by host enzyme in human small intestine, and has limited bioavailability, thereby affecting the full play of the active function. Therefore, the exploration of a preparation way of the low molecular weight ulva pertusa polysaccharide to improve the solubility and bioavailability thereof has become an important research direction in the field. For example, the separation and structural study of Ulva pertusa polysaccharide [ D ] Qingdao university of science and technology, 2018. Sun. The extraction of Ulva pertusa polysaccharide and the study of its biological activity [ D ] Zhejiang university of sea, 2021. The polysaccharide is purified by the conventional hot water extraction method, and then the activity thereof is studied. In order to improve the traditional preparation process, the prior research provides a plurality of technical paths such as physical degradation, chemical degradation, biological degradation and the like. Among them, biodegradation has been receiving a great deal of attention because of its mild reaction conditions and high selectivity. The method utilizes specific glycoside hydrolase or polysaccharide lyase to directionally cut the polysaccharide main chain, thereby not only avoiding the problems of high energy consumption and difficult control of molecular weight distribution of physical degradation methods (such as ultrasonic and microwave treatment), but also reducing the damage to active groups such as glycosidic bonds, sulfate groups and the like in the chemical degradation process, preventing the polysaccharide structure from being heterogeneous, and further effectively retaining the natural biological activity of the polysaccharide. This is particularly critical for the development of immune-functional sulfated polysaccharides such as ulva pertusa polysaccharides which rely on specific structures. However, the existing biodegradation method mostly adopts a mode of constructing engineering bacteria expressing ulva pertusa polysaccharide lyase. The scheme needs to be subjected to complex molecular biological processes such as gene cloning, vector construction, heterologous expression optimization and the like, has high technical threshold and long development period, and meanwhile, the engineering bacteria are required to strictly control the induction conditions and the culture environment in culture, so that the production cost is high, and the method is not suitable for industrial application. Disclosure of Invention In order to solve the above-mentioned technical problems, the "microbial fermentation method" is receiving attention as a simple and efficient biodegradation strategy. According to the method, engineering bacteria are not required to be constructed manually, bacillus behenii and ulva pertusa algae powder are selected for co-fermentation, and effective breaking of ulva pertusa polysaccharide chains is realized by utilizing a compound enzyme system such as glycoside hydrolase and polysaccharide lyase secreted by microorganisms. The method has the advantages of simple and convenient process, mild condition and lower cost, and can improve the solubility and molecular weight distribution of polysaccharide while maintaining the structure of