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Bacillus natto and application thereof to fermenting ruditapes philippinarum to prepare active polysaccharides

A technology of Philippine clams and active polysaccharides, applied in the directions of application, fermentation, organic active ingredients, etc., to achieve the effect of obvious anti-cancer activity and increase of polysaccharide yield

Active Publication Date: 2018-08-17
QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although fermented products of Bacillus natto have anti-tumor, blood pressure-lowering, anti-oxidation, and thrombolytic health-care effects, researchers at home and abroad have mainly focused on nattokinase in the research on the active substances, but there are few anti-tumor active polysaccharides. to report

Method used

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  • Bacillus natto and application thereof to fermenting ruditapes philippinarum to prepare active polysaccharides
  • Bacillus natto and application thereof to fermenting ruditapes philippinarum to prepare active polysaccharides
  • Bacillus natto and application thereof to fermenting ruditapes philippinarum to prepare active polysaccharides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Screening and identification of embodiment 1 bacillus natto

[0039] (1) Preliminary screening of Bacillus natto: Weigh 0.2g natto in 20mL 0.9% normal saline, shake for 1h, heat the bacterial suspension in a constant temperature water bath at 80°C for 10min, then rapidly cool to room temperature, then The bacterial suspension was used as the original bacterial suspension; 0.5mL of the original bacterial suspension was added to 4.5mL sterile saline to make 10-1 Dilute the bacterial suspension, prepare 10 by gradient dilution as above -2 ~10 -8 Dilute the bacterial suspension. Take 100 μl each of the above 10 -5 ~10 -8 Add the diluted bacterial suspension to the beef extract peptone medium plate with a salt content of 2%, set 3 parallels for each treatment, and spread the diluted bacterial suspension evenly. The above plate was moved to a 37°C constant temperature incubator and incubated upside down for 24 hours. Observe the growth of colonies, select a plate with un...

Embodiment 2

[0052] Example 2 The preparation method of the anti-tumor polysaccharide of Philippine clam

[0053] (1) Pretreatment of Philippine clams: select fresh and live clams with a length of about 4 cm, steam them in a steamer after spitting out sand with clean water, and take out the meat after the shells are opened, and beat them into a homogenous paste.

[0054] (2) Preparation of seed bacteria suspension: After activating the 7 isolated Bacillus natto strains, pick 1 to 2 rings of bacteria from the inclined surface and transfer them to the triangular flask containing the seed liquid medium, Cultivate on a shaking table at 37°C for 24 hours at 200r / min to obtain a seed suspension with a concentration of 1x10 8 ~8x10 8 cfu / mL. Wherein, the seed liquid culture medium is: peptone 1%, beef extract 0.5%, sodium chloride 0.5%, glucose 0.5%, and the pH is adjusted to 7.2-7.4.

[0055] (3) Fermentation: add distilled water to the clam meat according to the ratio of material to liquid o...

Embodiment 3

[0060] Example 3 Polysaccharide (RPP) Yield Determination

[0061] Accurately weigh 10mL of glucose standard solution in a 100mL volumetric flask, add water to make up to the mark (i.e. 100μg / mL glucose standard solution), draw 0.2mL, 0.4mL, 0.6mL, 0.8mL, 1.0mL of glucose standard solution respectively In a dry and clean test tube, add water to 1mL, and the blank control is 1mL water. Add 1mL of 6% phenol and 5mL of concentrated sulfuric acid to all the test tubes, shake them well and let them stand for 5min. After heating in a boiling water bath at 100°C for 15min, take them out and cool them down to room temperature. Measure its absorbance and draw a standard curve, such as figure 1 As shown, the linear regression equation is: y=0.0087x+0.0061, R=0.0087, and the glucose concentration x has a good linear relationship with the absorbance value y.

[0062] Determination method of polysaccharide content in the sample: add 50 ml of water to dissolve the polysaccharide powder pr...

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Abstract

The invention discloses Bacillus subtilis subsp. natto PS-I-HepG2. The Bacillus subtilis subsp. natto PS-I-HepG2 is used for fermenting ruditapes philippinarum to prepare active polysaccharides. The preparation method of the active polysaccharides comprises pretreatment of ruditapes philippinarum, preparation of bacterium suspension, fermentation, extraction, alcohol precipitation, deproteinization, ultra-filtration and freeze-drying to obtain ruditapes philippinarum polysaccharide powder. According to the Bacillus subtilis subsp. natto PS-I-HepG2, hepatoma carcinoma cell in vitro activity inhibiting experiments prove that the polysaccharides prepared by fermenting the ruditapes philippinarum through the Bacillus subtilis subsp. natto PS-I-HepG2 have significant anti-cancer activity and achieve a three-fold yield compared those produced before the ruditapes philippinarum is fermented. The active polysaccharides obtained through the Bacillus subtilis subsp. natto PS-I-HepG2 via the preparation method can be applied to preventing and treating solid tumors such as lung cancer and liver cancer.

Description

technical field [0001] The invention relates to the field of polysaccharide preparation, in particular to the isolation and identification of a strain of natto bacteria and its use for fermenting clam philippines to prepare active polysaccharides. Background technique [0002] The Philippine clam (Ruditapes philippinarum) belongs to the mollusca (Mollusca), bivalve (Veneroida), curtain clam (Veneridae), clam (Ruditapes), commonly known as clams, flower clams, clams, clams, etc. Wide-temperature and wide-haline shellfish are distributed in my country from Liaoning in the north to Guangxi and Hainan in the south. Philippine clams are fresh and tender, with excellent taste and rich nutrition. They have high economic value and medicinal value. They are also an important species for tidal flat farming in my country. In 2016, the annual output of Philippine clams was about 3 million tons, accounting for 90% of the world's total output, and the output of a single species was the hi...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/02C12P19/04A23L17/50A23L29/00A23L33/125A61K31/715A61P35/00C12R1/07
CPCA23V2002/00A61K31/715A23L17/50A23L17/65A23L29/065A23L33/125C12N1/02C12P19/04C12N1/205C12R2001/07A23V2200/308A23V2250/51
Inventor 魏玉西张雪梅高翔
Owner QINGDAO UNIV
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