Application of sanguinarine in preparation of helicobacter pylori urease inhibitor
A technology of Helicobacter pylori and urease inhibitor, which is applied in the application field of sanguinarine in the preparation of Helicobacter pylori urease inhibitor, can solve the problems of loss of effect of acetylhydroxamic acid, low toxicity and the like, and achieves inhibition of Helicobacter pylori urease Activity, effect on reducing resistance, promising development prospects
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Embodiment 1
[0039] Embodiment 1: Sanguinarine inhibits Helicobacter pylori urease activity test
[0040] Mix a certain concentration of Helicobacter pylori urease and a series of concentrations of sanguinarine solutions, incubate at 37°C for 20 minutes, then add urea solution at room temperature to react in the dark for 20 minutes, and then add Berthelot color developing solution in the dark for 10 minutes. Pipette 200 μL of the incubation solution onto a 96-well plate, and measure the OD at 595 nm with a microplate reader 绝对 value, get the corresponding OD相对 Value, the solvent added to the substance was used as a blank control, and the parallel determination was performed 3 times. Calculate the corresponding OD relative value according to formula 1. According to formula 2, the residual enzyme activity (Residual activity, RA) was obtained, and the corresponding half inhibitory concentration IC was obtained through the concentration-residual enzyme activity curve 50 , expressed as mean ±...
Embodiment 2
[0044] Example 2: Research on the type of inhibition of sanguinarine to Helicobacter pylori urease
[0045] A certain concentration of Helicobacter pylori urease was mixed with a series of concentrations of sanguinarine solutions (0, 0.25, 0.5, 1.0 mM), and incubated at 37° C. for 20 min. Then add a series of concentrations of urea solution (0.468-7.5mM) at room temperature to react in the dark for 20min, then develop color according to the method of Example 1 and measure the OD 绝对 value and obtain the corresponding OD 相对 value. The solvent of the added substance was used as a blank control, and the parallel determination was performed 3 times. The experiment passes the reciprocal of the reaction velocity (1 / v, that is, 1 / OD 相对 ) to the reciprocal (1 / [urea]) of the substrate concentration to make a Lineweaver-Burk diagram, and finally obtain the kinetic parameter K by combining the L-B curve with formula 3 M , v max , and through the L-B curve to obtain the inhibition con...
Embodiment 3
[0048] Embodiment 3: The influence of thiol-containing compound on Helicobacter pylori urease activity
[0049] incubation time test
[0050] Mix the Helicobacter pylori urease solution, the sulfhydryl compound solution (dithiothreitol, cysteine or glutathione solution, the concentration is 1.25mM) and the 1.0mM sanguinarine solution, and pre- After incubation for 5, 10, 20 or 40 minutes, add urea to react for 20 minutes at room temperature, then develop color and measure OD according to the method in Example 1 绝对 value and obtain the corresponding OD 相对 Value, and calculate the corresponding residual enzyme activity RA. The solvent of the added substance was used as a blank control, and the parallel determination was performed 3 times.
[0051] Interaction test of sanguinarine-mercapto-urease
[0052] Mix the Helicobacter pylori urease solution, the sulfhydryl compound solution (dithiothreitol, cysteine or glutathione solution, the concentration is 1.25mM) and the 1.0...
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