A kind of method for synthesizing caffeic acid with levodopa as substrate
A caffeic acid and biosynthesis technology, applied in microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problems of long production cycle, poor solubility, and no significant increase in yield, and achieve high production efficiency and substrate. The effect of high solubility and high yield
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0026] Construction method of recombinant Escherichia coli: TcXAL was optimized and synthesized by GenScript Biotechnology Co., Ltd., and cloned into pUC57-Simple, and the recombinant plasmid was named pUC57-TcXAL. Recombinant vector pUC57-TcXAL and expression vector pET-28a(+) were digested with restriction endonuclease Bam HI / HindIII, the cut products were separated by agarose gel electrophoresis, and the target gene TcXAL (2070bp) and expression vector ( 5368bp). Mix the cleaved target gene and expression vector at a molar ratio of 4:1, and use T4 ligase to ligate overnight at 16°C. The ligation product was transformed into Escherichia coli JM109 competent cells, and LB plates containing 50 μg / mL kanamycin were spread. Positive transformants were verified by colony PCR, and the primer sequences used were SQ4 / SQ5. Transfer the positive transformants to liquid LB medium containing 50 μg / mL kanamycin, culture overnight at 37°C and 220 rpm, extract the plasmid, and transform ...
Embodiment 2
[0030] Temperature condition optimization in the conversion process of caffeic acid of embodiment 2
[0031]The temperature conditions during the conversion of caffeic acid were optimized. The recombinant E. coli DCA-1 was used as the catalyst, PBS (50mM, pH 7.0) was used as the reaction medium, and the concentration of levodopa was 1g / L. The transformation was carried out under the temperature conditions of 20°C, 25°C, 30°C, 37°C and 42°C respectively, and the rotation speed was set at 220 rpm. Since caffeic acid reached its maximum production at 6 hours, samples were taken at 6 hours to determine the production and conversion of caffeic acid.
[0032] Such as Figure 4 As shown, the results show that the production of caffeic acid increases with the increase of temperature, and when the temperature exceeds 37°C, the production of caffeic acid does not increase but decreases. This may be related to the degradation of caffeic acid under higher temperature conditions. After ...
Embodiment 3
[0033] Example 3 The Effect of pH on the Production Results of Caffeic Acid
[0034] The pH conditions in the conversion process of caffeic acid were optimized, the recombinant E. coli DCA-1 was used as the catalyst, and the concentration of levodopa was 1g / L. Transformation conditions were set at 37°C, 220 rpm. PBS (50 mM) with different pH (6.0, 6.5, 7.0, 7.5, 8.0, 8.5) was used as the transformation medium. Samples were taken at 6 hours to measure the yield and conversion rate of caffeic acid.
[0035] Such as Figure 5 As shown, the results showed that the production of caffeic acid increased with the increase of pH, and when the pH exceeded 7.5, the production of caffeic acid began to decrease. This may be related to the easy conversion of caffeic acid into benzoquinones under alkaline conditions. After conversion at 37°C for 6 hours, the caffeic acid reached the maximum yield, namely 910.90 mg / L, with a conversion rate of 99.70%, which is the highest conversion rate ...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More