Acid and choline-resistant separated strain of lactobacillus with ability of reducing and assimilating cholesterol
A technology of Lactobacillus and isolates, which is applied in the field of novel Lactobacillus isolates, can solve the problems such as difficulty in maintaining the number of bacteria, insufficiency, complex nutritional requirements, etc.
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Embodiment 1
[0064] Example 1 Isolation and Screening of Lactobacillus Isolates
[0065] Materials and Methods:
[0066] 1. Medium and diluent:
[0067] 1. Screening medium:
[0068] (A) Tomato juice agar medium
[0069] Casein enzymatic hydrolyzate (Sigma, Louis, Mo, USA) 10g
[0070] Skimmed milk 10g
[0071] Tomato Juice 400ml
[0072] Agar 12g
[0073] Distilled water up to 1L
[0074] (B) Rogosa agar medium
[0075] Rogosa agar (Merck, Darmstadt, Germany) 74.5 g
[0076] Distilled water 1L
[0077] After dissolving by microwave heating, wait to cool to 55°C, adjust the pH value to pH 5.5 with acetic acid
[0078] (C) Lactobacillus selection medium (LBS)
[0079] Rogosa agar 8.4g
[0080] Tomato Juice 40ml
[0081] Distilled water 60ml
[0082] After dissolving by microwave heating, wait to cool to 55°C, adjust the pH value to pH 5.5 with acetic acid
[0083] (D) Rogosa+X-glu agar medium
[0084]80 mg of 5-bromo-4-chloro-3-indolyl 1-β-D-glucopyranoside (X-glu) (Sigma,...
Embodiment 2
[0131] Example 2 Acid resistance test
[0132] 1. Experimental operation procedure:
[0133] With reference to the method disclosed in J.E.Holcombe et al., (1991), Cult.Dairy Prod.J., 26(3):4-5 and US 5,711,977 granted to Y.S.Yang et al., neutral (pH 7) and imitative gastric acid Environment (pH 2) to test the number of surviving bacteria.
[0134] After each test strain is activated twice with MRS broth, get its bacterial solution and centrifuge (3000rpm, 10 minutes), after removing the supernatant, add 1ml of normal saline solution (0.85%NaCl, pH 7), first with a bamboo stick Stir the bacteria, shake them evenly, and then take 0.5ml of the dispersed and floated bacteria solution and put them in the physiological saline solution of pH 7 and pH 2 respectively. number of surviving bacteria. The smaller the value of Δlog bacterial count [= log (bacterial count at pH 7 - bacterial count at pH 2)], the strain with better acid resistance is considered.
[0135] 2. Results:
...
Embodiment 3
[0139] Example 3. bile salt tolerance test
[0140] 1. Experimental operation procedure:
[0141] After each test strain was activated twice with MRS broth, 1% of the inoculum was inoculated into 10ml MRS broth and 10mL MRS broth (MRSO) containing 0.3% ox bile, and cultured at 37°C for 24 hours. Spectrophotometer was used to measure the bacterial concentration (OD 660 ). In addition, the number of remaining bacteria was also counted, where Δlog bacteria value=log(number of bacteria in MRS broth-number of bacteria in MRSO broth), and the smaller the number of Δlog bacteria, the better the bile salt tolerance of the test strain.
[0142] 2. Results:
[0143] After the lactic acid bacteria are treated with bile salts, the survival rate of different strains is very different. Therefore, it is important to screen the lactic acid bacteria strains with tolerance to bile salts in the selection of probiotics (S.E.Gilliland et al., (1984), J . Dairy Sci., 67: 3045-3051; P. Marteau...
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