A kind of Lactobacillus plantarum capable of improving sleep and its application
A technology of Lactobacillus plantarum and Lactobacillus, applied in the field of microorganisms, can solve the problems such as acid resistance, bile salt resistance and colonization ability of lactic acid bacteria not involved, and achieve the effects of excellent simulated gastrointestinal tract survival ability, prolonged sleep time, and wide application prospects.
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Embodiment 1
[0039] The screening of embodiment 1 lactobacillus plantarum
[0040] 1 Experimental method
[0041] Strains were screened using fresh human feces and pickled vegetable water as samples.
[0042] (1) Pipette 0.5 mL of the sample into 5 mL of MRS medium, culture at 37° C. for 18 to 24 hours for enrichment.
[0043] (2) Gradient dilution: absorb 0.5mL of the enriched sample into 4.5mL sterile saline to obtain 10 -1 Diluent, then draw 0.5mL 10 -1 Diluted solution in 4.5mL normal saline to obtain 10 -2 Diluent, according to this operation, successively get 10 -3 , 10 -4 , 10 -5 , 10 -6 Diluent.
[0044] (3) Plating culture: draw 100 μL of gradient dilution and spread it on the MRS plate, 10 -4 , 10 -5 , 10 -6 One plate per gradient; cultured at 37°C for 48h.
[0045] (4) Separation by streaking: Select colonies with typical characteristics on the coated plate according to the shape, size, edge, and transparency of the colony, pick the colonies with an inoculation loop,...
Embodiment 2
[0056] Example 2 Exploration of the Survival Ability of Different Lactobacillus Plantarum to Simulate the Gastrointestinal Tract
[0057] 1 Test method
[0058] (1) Preparation of simulated gastric juice: Dissolve pepsin in sterilized physiological saline (0.9% w / v, adjust pH to 3.0 with hydrochloric acid) so that the final concentration is 3 g / L. Filter through a 0.22μm sterile filter membrane and prepare for immediate use.
[0059] (2) Configuration of simulated intestinal juice: dissolve trypsin in sterilized physiological saline (0.9% w / v, adjust pH to 8.0 with NaOH) to make the final concentration 1g / L, and add bile salt to make the final concentration 0.3 %. Filter through a 0.22μm sterile filter membrane and prepare for immediate use.
[0060] (3) Experiments were carried out after successive activation of three generations of different Lactobacillus plantarum (18 hours each time). Determination of the continuous activation of three generations of strains in A 600 ...
Embodiment 3
[0069] Example 3 Exploration of the In Vitro Adhesion Ability of Different Lactobacillus plantarum to HT-29 Cells
[0070] 1 Test method
[0071] Normal HT-29 cells were cultured and subcultured, and different Lactobacillus plantarum were continuously activated for 3 generations. The cultured cells were collected, counted with a hemocytometer, and resuspended to 2×10 in 1640 medium (adding double antibodies and fetal bovine serum). 5 individual / mL. Add acid-washed and sterilized coverslips to 6-well plates. Inoculate 2 mL of cell suspension, and wash 3 times with sterile PBS after the cells adhere to the coverslip (about 12 hours). Take a certain amount of bacterial liquid to collect the bacterial cells by centrifugation, wash the bacterial cells once with sterile PBS, and centrifuge again to collect the bacterial cells, and resuspend the bacterial cells in 1640 medium (without adding double antibodies and fetal bovine serum) to a bacterial concentration of 10 8 CFU / mL. A...
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