Preparation method of unsaturated hyaluronic acid odd oligosaccharides
A technology of hyaluronic acid and hyaluronidase, applied in the field of biomedicine, can solve the problems of low reaction efficiency, complex process, non-single reaction product structure, etc., and achieve the effects of single structure, simplified preparation process and improved preparation efficiency.
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Embodiment 1
[0037] Step 1) Prepare the hyaluronic acid (HA) solution with a concentration of 10 mg / mL from the lyophilized powdered hyaluronic acid (HA) with pure water, and prepare the lyophilized powdered microbial hyaluronidase with an enzymatic hydrolysis working solution 200IU / mL hyaluronidase solution;
[0038] Further, the enzymolysis working solution is pure water;
[0039] Further, the microbial hyaluronidase is derived from Streptomyces hyalurolyticus;
[0040] Step 2) Take 5 parts of the hyaluronic acid (HA) solution, add 1 part of the hyaluronidase solution under ice bath conditions, shake and mix, and incubate in a 37°C water bath for 24 hours to prepare a sample; The sample was moved to a water bath at 100°C, heated for 5 minutes, centrifuged, and precipitated;
[0041] Step 3) using a low-pressure chromatography column (2.6 × 100cm) with a filler of Bio-gel P10 to separate and purify the sample after deprecipitation, wherein the mobile phase concentration is 0.2M sodium chl...
Embodiment 2
[0044] Step 1) Prepare lyophilized powdered hyaluronic acid (HA) with pure water to form a hyaluronic acid (HA) solution with a concentration of 8 mg / mL, and prepare lyophilized powdered microbial hyaluronidase with enzymatic hydrolysis working solution 150IU / mL hyaluronidase solution;
[0045] Further, the enzymolysis working solution is pure water;
[0046] Further, the microbial hyaluronidase is derived from Streptomyces hyalurolyticus;
[0047] Step 2) Take 8 parts of the hyaluronic acid (HA) solution, add 2 parts of the hyaluronidase solution under ice bath conditions, shake and mix, and incubate in a 37°C water bath for 30 hours to prepare a sample; The sample was moved to a water bath at 100°C, heated for 7 minutes, centrifuged, and precipitated;
[0048] Step 3) using a low-pressure chromatography column (2.6 × 100cm) filled with Bio-gel P6 to separate and purify the sample after deprecipitation, wherein the mobile phase concentration is 0.1M potassium chloride, and ...
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