Preparation Of Anti-urolithiasis Effective Fractions Of Alisma Orientalis (Sam.) Juzep. And Study Its Anti-urolithiasis Effect

Objective: Based on the early pharmacology experiments. Total triterpenes was identified to be effective fractions of Alisma orientalis preliminary.Macroporous resin was used in separation and purification of the total triterpenes of Alisma orientalis, the content of total triterpenes in the effective fractions reached More than 50 percent. And Alisol B 23-acetate was chose as evaluation index and its amount reached 1.29 percent and above. And the Anti-urinary tract stones effect of effective fractions was studied in vitro.Methods and results: A new method was established for the determination of total triterpenes from the effective fractions of Alisma orientalis. Total triterpenes was measured by spectrophotometer with 5% vanillin-glacial acetic acid solution and perchloric acid as color-developing agent and detected at 554 nm wavelength. The calibration curve was linear (r=0.9996) in the range of 23.8～142.8ug and the curve was Y=0.0055x-0.0512.The experiment showed that the spectrophotometer was suitable for the determination of total triterpenes from Alisma orientalis.A new method was established for the determination of Alisol B 23-acetate from the effective fractions of Alisma orientalis. The content of Alisol B 23-acetate was measured by HPLC-ELSD. The experiment showed that the calibration curve was linear in the range of 0.1384ug～0.6920ug and the curve was Y=494467X-4426.9,r=0.9999. The results of the experiments of accuracy,reproducibility and stability were good.The observation of the extracted term of the effective fractions of Alisma orientalis with the amount of Alisol B 23-acetate as evaluation index gave the result that the best extracted term was that Alisma orientalis was extracted by 70% alcohol for three times, each time last for 1.5h. The volume of alcohol was ten times of the medicinal substances each time. In several kinds of macro absorption resins, conclusion was reached that the A resin was the best and the best technology term was that First, the extractum of Alisma orientalis was dissolved by a certain amount of 50% ethanol solution and then the sample was fully adsorpted by the HPD-100 macro absorption resin; Second, a certain amount of distilled water, 30% ethanol, 90% ethanol was used to elute the sample in order. Then, the eluting of 90% ethanol solution was collected. Solvent was evaporated. Dried and then the residues was collected. Alisol B 23-acetate was used as an evaluation index and gave the result that the concentration of total triterpenes reached up to 50 percent, and the concentration of Alisol B 23-acetate also reached up to 1.29 percent. Experiments showed that: HPD-100 macro resin was suitable for isolation and purification of triterpenoid compounds which polarity was weak in Alisma orientalis.To study the effect of total triterpenes of Alisma Orientalis on calcium oxalate crystal formation in vitro and to identify the effective constituents. We detected the growth index (I.I) of calcium oxalate crystal by a seeded crystallization technique. Both the high and low dose of total triterpenes can reduce the growth index of crystallization reach 87.75%. The total triterpenes of Alisma Orientalis seems to inhibit significantly calcium oxalate crystal formation in vitro and to be the effective constituent of Alisma Orientalis.