Application of helicase in directionally hydrolyzing rutin for biosynthesizing Quereetin
A biosynthesis and helicase technology, applied in the direction of fermentation, etc., can solve the problems of unrealized directional transformation of quercetin, no rutinoside bonds, etc., and achieve good industrial application prospects, high catalytic efficiency and specificity, and enzyme catalysis High conversion effect
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Embodiment 1
[0029] A. The free enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0030] Rutin was dissolved in a pH 7 buffer to obtain a rutin substrate solution with a concentration of 0.005 g / L. The substrate solution was placed in a 50 mL Erlenmeyer flask with a stopper, the helicase crude enzyme preparation was added in a ratio of 50 g / L, the cork was tightly ground, and the enzymatic hydrolysis reaction was carried out in a rotary shaker at 120 rpm. The reaction temperature is 40°C, and the reaction time is 0.5h. After the reaction was completed, the molar conversion rate of the hydrolyzed quercetin detected by HPLC was 92%.
[0031] B. The immobilized enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0032] Prepare 0.5%wt colloidal solution for immobilized enzyme with carrageenan, add 50g / L helicase crude enzyme powder, mix well, drop into cross-linking agent solution and solidify for 3 hours...
Embodiment 2
[0035] A. The free enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0036] Rutin was dissolved in a pH3 buffer to obtain a rutin substrate solution with a concentration of 0.100 g / L. Place the substrate solution in a 50mL conical flask with a stopper, add the crude helicase preparation at a ratio of 0.1g / L, plug the cork tightly, and carry out the enzymatic hydrolysis reaction in a rotary shaker at 120rpm , the reaction temperature is 10°C, and the reaction time is 36h. After the reaction was completed, the molar conversion rate of the hydrolyzed quercetin detected by HPLC was 22%.
[0037]B. The immobilized enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0038] Use gelatin to make 15%wt colloidal solution for immobilized enzyme, add 0.100g / L helicase crude enzyme powder, mix well, drop into crosslinking agent solution and solidify for 12h, and make square immobilized particles with ...
Embodiment 3
[0041] A. The free enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0042] Rutin was dissolved in a pH 9 buffer to obtain a rutin substrate solution with a concentration of 0.025 g / L. Place the substrate solution in a 50mL conical flask with a stopper, add the crude helicase preparation at a rate of 0.5g / L, plug the cork tightly, and carry out the enzymatic hydrolysis reaction in a rotary shaker at 120rpm , the reaction temperature is 70°C, and the reaction time is 0.1h. After the reaction was completed, the molar conversion rate of the hydrolyzed product quercetin detected by HPLC was 23%.
[0043] B. The immobilized enzyme of helicase catalyzes the directional biosynthesis of quercetin by hydrolyzing rutin:
[0044] Prepare a 5%wt colloidal solution for immobilized enzymes with sodium alginate, add 0.5g / L helicase crude enzyme powder, mix well, drop into the crosslinking agent solution for 0.5h, and make spherical shapes with u...
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