AUH-JLC159 and transforming method thereof for preparation of (-)-5-OH-equol
A technology of AUH-JLC159, -5-OH-, applied in the field of Snaekia and its preparation of -5-OH-equol in genistein conversion
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Embodiment 1
[0045] 1. Isolation and cultivation of strain JLC159
[0046] (1) Collection and cultivation of rooster manure samples
[0047] Use a sterilized cotton swab to pick a fresh fecal sample, put it into 1 ml of fresh BHI liquid medium, and place it in an anaerobic workstation at 37°C, as a microbial flora for screening specific functional microbial strains;
[0048] (2) Isolation and cultivation of transformed strains
[0049] ①Single colony isolation culture
[0050] Use the microbial flora in the fresh BHI liquid medium for gradient dilution, and dilute to a concentration of 10 –1 , 10 –2 、10 –3 、10 –4 、10 –5 、10 –6 、10 –7 、10 –8 , and then 100 microliters of concentration were respectively 10 –5 、10 –6 、10 –7 、10 –8 The dilution of microbial flora was uniformly coated on the pre-prepared BHI solid medium, and the BHI solid medium coated with the dilution of microbial flora was placed in an anaerobic workstation for 48 hours. Pick dozens to hundreds of single colon...
Embodiment 2
[0084] The isolation method of bacterial strain JLC159 is the same as that in Example 1.
[0085] In the anaerobic workstation, the seed liquid of the above-mentioned pre-cultivated strain JLC159 was transferred to a 250-ml Erlenmeyer flask containing 100 ml of liquid medium at a 15% inoculum size, and 40 mmol / L of genistein was added at the same time. 1.25 ml of mother liquor was cultured in an anaerobic workstation for 3 days. Use an equal volume of ethyl acetate to extract the culture in the Erlenmeyer flask twice, filter the extract and evaporate to dryness on a rotary evaporator, then add 100% methanol solution, pass through an organic membrane with a pore size of 0.45 microns, and use a preparative column to perform HPLC analysis. Separation and preparation were carried out above to obtain a conversion rate of (-)-5-OH-EQ of 83.3%.
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