Research On TDC Activity And Alkaloid Purification In Catharanthus Roseus

Catharanthus Roseus has attracted many people's attention in biopharmacy, it was make certain the mode plant in study the pathway of secondary metabolites, for the cause of it is applied efficiency in anti-tumour at present. It has been reported that TDC is the key enzyme in biosynthesis of alkaloid from Catharanthus Roseus; we can inprove the indole alkaloid accumulation by the way change the environment factor. The paper based from inducement of the Catharanthus Roseus callus, through filter the high produce alkaloid callus cells and determination the activity of TDC and POD in the pathway in cultures, and study on the relation between environment factor, enzyme and alkaloid accumulation, discuss fine determination and purification of the alkaloid. The result is followed:1 .From the study on the condition to induce callus, we find it can induce callus only use 2.4-D, the rate is low only use NAA, the effect is best when simultaneity use concentration of 2.4-D at 2.0 mg/L and 6-BA at 0.5 mg/L, 6-BA can improve the induce rate. Through the UV irradiation on callus, we attained a high producted callus and rename it R40.2.The differentiation of apparatus make little impact on TDC activity, typtamine content is positive correlation with TDC; POD activity has some relationship with the content of chlorophyll; th content of alkaloid is asymmetry and positive correlation with POD.3.It can improve the activity of TDC and POD(acidity) in culture by the way illumination, added some filter adjust reagent Na⁺ and K⁺, anti-oxidation reagent PVP. Some environment factors as Na⁺ ,K⁺ and PVP can improve the accumulation of alkaloid in culture.4. HPLC method for simultaneous determinate alkaloid in Catharanthus Roseus was established, with the Mobile phase of HPLC consisted of super purified water(containing 0.005 mol · L^-1 ammonium hydrogen phosphate), methanol(containing 0.67% triethylamine) and acetonitrile. The elution was designed as follows: 0-10 min, gradient, the amount of methanol was increased linearly from 34% to 52% and acetonitrile from 20% to 28% ; 10-34 min, gradient, the amount of methanol was decreased linearly from 52% to 45% and acetonitrile...