Locust Flower Total Extraction Process And Quality Standards

Purpose:1. To optimize the extraction technology for total flavonoids from Robinia pseudoacacia L.(TFR), and give reference for development and utilization of Robinia pseudoacacia L.; To study the purification process of total flavonoids in Robinia pseudoacacia L. with macroporous resin, and give science referenc for industrial production.2. In accordance With Chinese Pharmacopoeia 2005 (Volume l), the quality standard of TFR was researched. According to the experimental results,the quality standard of TFR has been drafted. The results of the stability experiments showed that TFR was stable under the conditions.3. Through vitro Anticancer activity screening of total flavonoids of Robinia pseudoacacia L.enlarge application rang, and give reference for further study of its anticancer mechanism.Material and method:1. Material:Harvested from April to June 2009 were collected from Dalian, Liaoning Province, Shenyang, Jinzhou, Chaoyang, Jilin, Siping City, Siping Tiancheng County ten batches of samples from different areas (see table 1-1), and bud and the flowering of the different harvest periods2. Method:1) Optimize the extraction technology for total flavonoids from Robinia pseudoacacia L.by single factor test and orthogonal test; Using static absorption to optimum maeroporous resin which was used for purifying total flavonoids in Robinia pseudoacacia L. Using dynamic absorption to the condition of purifying total flavonoids of Robinia pseudoacacia L.by macroporous resin.2) Researches on the quality standard of TFRIn accordance With Chinese Pharmacopoeia 2005 (Volume l), the quality standard of TFR was researched. TFR was identified by chemical reaction and thin layer chromatography (TLC).General testing methods had been carried out, such as limit test for heavy metals and arsenic,determination of loss on drying and water, etc.The contents of total flavonoids and two xanthones were determined. Total flavonoids,calculated as Robinin,was determined by colourmetry. Two xanthones, were determined by HPLC.According to the experimental results,the quality standard of TFR has been drafted. 3) MTT assay was employed to investigate the effects done by total flavonoids on the growth and proliferation ability of SMMC-7721,MGC-8,HT-29 cells.Results:1. By single factor test and orthogonal test, the optimum process of total flavonoids was as follows: the crude drug was extracted three times, the 75% alcohol density, 40 times alcohol was added and 1 h every time; Optimum maeroporous resin was D101,and the optimum condition was as follow:the speed that sample flows through macroporous resin was 2.0 mL·min^-1,and the total flavonoids absorbed on maeroporous resin can eluted by 75%ethanol of 12 BV.2. Researches on the quality standard of TFRIn accordance With Chinese Pharmacopoeia 2005 (Volume l), the quality standard of TFR was researched. TFR was identified by chemical reaction and thin layer chromatography (TLC).General testing methods had been carried out, such as limit test for heavy metals and arsenic,determination of loss on drying and water, etc.The contents of total flavonoids and two xanthones were determined. Total flavonoids,calculated as Robinin,was determined by colourmetry. Two xanthones, were determined by HPLC.According to the experimental results,the quality standard of TFR has been drafted. The results of the stability experiments showed that TFR was stable under the conditions of 60℃,RH92.5% and 4500Lx士500Lx for at least 10 days,respectively. And TFR was stable after being placed at 40℃,RH75% and at room temperature for 3 months.3. Total flavonoids have more inhibitory effects on viability of SMMC-7721 cells,MGC-8 cells,HT-29 cell than negative control group;and inhibitory effects depend on drug's concentration.Conclusion:1) By single factor test and orthogonal test, the optimum process of total flavonoids was as follows: the crude drug was extracted three times, the 75% alcohol density, 40 times alcohol was added and 1 h every time; Optimum maeroporous resin was D101,and the optimum condition was as follow:the speed that sample flows through macroporous resin was 2.0 mL·min^-1,and the total flavonoids absorbed on maeroporous resin can eluted by 75%ethanol of 12 BV.2) Researches on the quality standard of TFRIn accordance With Chinese Pharmacopoeia 2005 (Volume l), the quality standard of TFR was researched. TFR was identified by chemical reaction and thin layer chromatography (TLC).General testing methods had been carried out, such as limit test for heavy metals and arsenic,determination of loss on drying and water, etc.The contents of total flavonoids and two xanthones were determined. Total flavonoids,calculated as Robinin,was determined by colourmetry. Two Prenylated xanthones, were determined by HPLC.According to the experimental results,the quality standard of TFR has been drafted.The results of the stability experiments showed that TFR was stable under the conditions of 60℃,RH92.5% and 4500Lx士500Lx for at least 10 days,respectively. And TFR was stable after being placed at 40℃,RH75% and at room temperature for 3 months. 3) Total flavonoids have more inhibitory effects on viability of SMMC-7721 cells,MGC-8 cells HT-29 cell than negative control group, and inhibitory effects depend on drug's concentration.