A method for separating and purifying l-citrulline
A technology for separation and purification, citrulline, applied in chemical instruments and methods, microorganism-based methods, biochemical equipment and methods, etc., can solve the problem of low arginine deiminase activity, and the enzymatic method is difficult to meet industrial production. , affecting the production of citrulline by enzymatic method, etc., to achieve the effect of high product purity, overcoming the bottleneck of citrulline production and saving dosage
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Embodiment 1
[0035] Example 1 A bacterial strain MQO-153 producing arginine deiminase, the present invention through repeated chemical mutagenesis and ultraviolet mutagenesis to Streptococcus faecalis (Streptococcus faecalis purchased from Beijing Microorganism Culture Collection Center), And after screening a large number of strains, a strain MQO-153 with high arginine deiminase activity was obtained, and its preservation number is CGMCC No.10726; Center; the preservation date is April 21, 2015; the preservation address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing.
Embodiment 2
[0036] Example 2 The method for preparing arginine deiminase by using bacterial strain MQO-153, the specific steps are as follows:
[0037] (1) Slant culture: inoculate the bacterial strain MQO-153 on the slant medium under aseptic conditions for cultivation, the cultivation temperature is 30° C., and the cultivation time is 24 hours. At this time, the bacterial colony is large and smooth without pigment accumulation;
[0038] Slant medium: Slant medium (g / L): beef extract 5, peptone 10, sodium chloride 5, agar 20; pH is 7.0-7.3; preferably 7.2; the sterilization temperature of the slant medium is 115 ° C, sterilized The time is 15 minutes.
[0039] (2) Expansion culture: pick bacterium colony from the slant culture medium of step (1), add water and dilute to obtain thalline solution, the concentration after dilution is 3 * 10 5 -5×10 5 cells / mL, inoculate the bacterial cell solution onto the seed medium and place it in a shaker for expanded culture, the inoculum size is 10%...
Embodiment 3
[0044] Example 3 The method for preparing L-citrulline from L-arginine through the conversion of arginine deiminase, the specific steps are as follows:
[0045] (1) Obtaining the substrate: the arginine ceramic membrane filtrate was adsorbed by 732 ammonia resin, and eluted with 2.0N ammonia water to obtain arginine positive column purified solution;
[0046] (2) Transformation culture: the bacteria containing arginine deiminase were washed once with normal saline, and the bacteria were collected after centrifugation. Transfer the bacteria to the transformation solution, which includes 0.45mol / L acetate buffer and 0.5g / L CTAB, add 10% L-arginine purification solution, and incubate at 37°C for 24h to obtain L-citrulline Amino acid conversion solution.
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