Traditional Chinese medicine polysaccharide extract freeze-drying protective agent, direct-to-vat starter and preparation methods thereof
A traditional Chinese medicine polysaccharide extract freeze-drying protective agent, a direct-to-vat starter and preparation methods thereof are disclosed, which belong to the technical field of feed starter. The compound traditional Chinese medicine polysaccharide extract freeze-drying protective agent includes Poria cocos polysaccharide extract, Atractylodes macrocephala polysaccharide extract, Radix codonopsis polysaccharide extract, L-tyrosine, α-cyclodextrin and polyethyleneimine. Its application in the freeze-drying protection of direct-to-vat starter of aquatic fermentation feed can reduce the damage to the cells caused by freeze-drying, improve the survival rate of the cells in the starter after freeze-drying, effectively prolong the storage period of the starter, and ensure that the fermentation activity remains at a high level during storage.
1 . A preparation method of a direct-to-vat starter, comprising
mixing a mixed strain solution of Enterococcus faecalis, Candida utilis, Aspergillus niger, Bacillus coagulans and Brevibacillus laterosporus with a freeze-drying protective agent according to a volume ratio of 1:1 to 1:4, and carrying out freeze-drying to obtain the direct-to-vat starter; wherein the freeze-drying protective agent consists of the following components:
Poria cocos polysaccharide extract 10.0-30.0 g/100 mL, Atractylodes macrocephala polysaccharide extract 10.0-20.0 g/100 mL, Radix codonopsis polysaccharide extract 5.0-20.0 g/100 mL, L-tyrosine 2.0-10.0 g/100 mL, α-cyclodextrin 2.0-7.0 g/100 mL and polyethyleneimine 2.0-5.0 g/100 mL.
2 . The preparation method of claim 1 , wherein in the mixed strain solution,
the viable concentration of Enterococcus faecalis is 1.0×10 10 -6.0×10 10 CFU/mL;
the viable concentration of Candida utilis is 3.0×10 8 -6.0×10 8 CFU/mL;
the viable concentration of Aspergillus niger is 8.0×10 7 -2.0×10 8 CFU/mL;
the viable concentration of Bacillus coagulans is 5.0×10 7 -3.0×10 8 CFU/mL; and
the viable concentration of Brevibacillus laterosporus is 1.0×10 8 -4.0×10 8 CFU/mL.
3 . The preparation method of claim 2 , wherein a preparation method of the mixed strain solution of Enterococcus faecalis, Candida utilis, Aspergillus niger, Bacillus coagulans and Brevibacillus laterosporus comprises:
(1) activating Enterococcus faecalis, Candida utilis, Aspergillus niger, Bacillus coagulans and Brevibacillus laterosporus respectively;
(2) after activation, mixing all activated strains and inoculating with a multiplication medium for multiplication culture, culturing the strains at 34° C. for 12 h, after culture is carried out, centrifuging the culture and discarding the supernatant, and performing resuspension with physiological saline to obtain the mixed strain solution;
the multiplication medium used in the process of multiplication culture is a composite medium of Jerusalem artichoke and Poria cocos;
a preparation method of the composite medium of Jerusalem artichoke and Poria cocos comprises: mixing Jerusalem artichoke powder, Poria cocos water extract and water at a mass ratio of 2:3:5, hydrolyzing at 65° C. for 60 min, heating to 95° C. and hydrolyzing for 60 min, cooling, filtering with 8 layers of gauze, and adjusting pH value of the filtrate to 7.0~7.3 to obtain the composite medium of Jerusalem artichoke and Poria cocos;
a preparation method of the Jerusalem artichoke powder comprises: slicing fresh Jerusalem artichoke, drying the fresh Jerusalem artichoke slices at 70~80° C. until the water content is lower than 8%, and then crushing the dried Jerusalem artichoke slices and sieving the crushed Jerusalem artichoke slices through 100 meshes to obtain the Jerusalem artichoke powder; and
a preparation method of the Poria cocos water extract comprises: slicing fresh Poria cocos , drying the fresh Poria cocos slices at 70~80° C., and then crushing the dried Poria cocos slices; adding 10~15 times the mass of absolute ethanol, extracting for 2~4 h and subjecting the extract to filtering to obtain a residue; adding 8~10 times the mass of distilled water to the residue, heating and refluxing to extract for 2 h, filtering twice to obtain filtrates, combining the filtrates, subjecting the combined filtrate to concentrating and drying under reduced pressure to obtain the Poria cocos water extract.
4 . The preparation method of claim 2 , wherein in the freeze-drying step, a prefreezing process is carried out for 3 h at −80° C., and the mixed strain solution is quickly transferred to a vacuum freeze-drying machine for freeze-drying after being completely frozen; the freeze-drying conditions comprise a vacuum degree of 10 Pa and a cold trap temperature of −55° C.; and the frozen mixed strain solution is first heated at 10° C. for 2 hours, then heated up to 20° C. for 3 hours, and finally heated up to 30° C.; for a total duration of the vacuum freeze-drying process is 20 hours.