Method for preparing tautomeric acid
The technology of allosteric acid and allosteric acid is applied in the preparation field of preparing allosteric acid by hydrolyzing allosteric acid with lipase, which can solve the problems of severe hydrolysis conditions, easy generation of by-products and the like, and achieves mild reaction conditions, response-specific effect
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Embodiment 1
[0020] Example 1 Preparation of allosteric acid by using Novozymes lipase to hydrolyze and ferment tamomycin
[0021] Dissolve the relative mutamycin sample in 0.2M phosphate buffer to prepare a 22500μg / mL mutamycin solution; take 1g of Novozym lipase and dissolve it in 50mL of 0.2M pH7.0 Phosphate buffer solution to make 200U / mL enzyme solution. Add 0.3 mL of enzyme solution (225000 μg: 60 U = 3750: 1) to 10 mL of tamomycin solution, react at 35° C. for 3 h, and hydrolyze tamomycin to obtain about 10 mL of tamomycin hydrolyzate.
[0022] Separation and purification of allosteric acid: adjust 10 mL of the above-mentioned tausteric acid hydrolyzate, adjust the pH to 4.0 with 1 mol / L HCl, then extract three times with the same volume of ethyl acetate, combine the extracts, and recover the ethyl acetate; Then adjust the pH of the raffinate phase (7 mL of the combined aqueous phase) to 2.0 with 1 mol / L HCl, extract 3 times with the same volume of ethyl acetate, combine the ethyl ...
Embodiment 2
[0023] Example 2 Utilizing Aspergillus niger lipase to hydrolyze and ferment tamomycin to prepare allosteric acid
[0024] Dissolve the relative mutamycin sample in 0.4M disodium hydrogen phosphate-citrate buffer to prepare a 40,000 μg / mL mutamycin solution; take 1.5 g of Aspergillus niger lipase and dissolve it in 50 mL 0.4M pH 7.0 disodium hydrogen phosphate-citric acid buffer solution to make 300U / mL enzyme solution. Add 0.5mL of enzyme solution (activity ratio: 400000U: 150U = 2667:1) to 10mL of tamomycin solution, react at 37°C for 4.0h, the hydrolysis of tamomycin is completed to obtain about 10mL of tamomycin hydrolyzate .
[0025] Separation and purification of allosteric acid: adjust the pH of 10 mL of the above-mentioned tausteric acid hydrolyzate to 4.0 with 1 mol / L HCl, then extract three times with the same volume of ethyl acetate, combine the extracts, and recover the ethyl acetate; Adjust the pH of the raffinate phase (combined aqueous phase 8mL) to 2.0 with 1...
Embodiment 3
[0026] Example 3 Utilizing Yeast Lipase to Hydrolyze and Ferment Tatamycin to Prepare Allosteric Acid
[0027] Dissolve the relative mutamycin sample in 0.3M phosphate buffer to prepare a 30000μg / mL mutamycin solution; take 2g of yeast (Candida rugosa) lipase and dissolve it in 50mL of 0.2M pH 7.0 phosphate buffer solution to make 400U / mL enzyme solution. Add 0.4 mL of enzyme solution (activity ratio: 300000 U: 120 U = 2500: 1) to 10 mL of tamomycin solution, react at 35° C. for 3 h, and hydrolyze tamomycin to obtain about 10 mL of tamomycin hydrolyzate.
[0028] Separation and purification of allosteric acid: adjust 10 mL of the above-mentioned tausteric acid hydrolyzate, adjust the pH to 4.0 with 1 mol / L HCl, then extract three times with the same volume of ethyl acetate, combine the extracts, and recover the ethyl acetate; Then adjust the pH of the raffinate phase (7 mL of the combined aqueous phase) to 2.0 with 1 mol / L HCl, extract 3 times with the same volume of ethyl ac...
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