Studies On Quality Control Method Of Flos Chrysanthemi Indici And The Preparation Process Of Standardized Extract For Injection

Flos chrysanthemi indici is the dried flowers of Chrysanthemum indicum L.of compositae. In this thesis, quality control method of flos chrysanthemi indici and the preparation process of the standardized extract are presented.Firstly, a study on quality control of flos chrysanthemi indici was carried out. The total flavonoids in flos Chrysanthemi indici samples were determined by UV-Vis spectrophotometry. Two bioactive constituents namely chlorogenic acid and caffeic acid in flos chrysanthemi indici samples were determined simultaneously by high performance liquid chromatography. The samples were separated on a Thermo Hypersil BDS C₁₈ column (4.6 mm×250 mm, 5μm) with the mobile phase of methanol-0.5% acetic acid (20:80, v/v). The flow rate was 0.8 rnL·min^-1. The detection was set at 325 nm and the column temperature at room temperature. The contents of chlorogenic acid and caffeic acid were 0.26-0.74% and 0.0050-0.056% respectively. The other two bioactive constituents namely linarin and luteolin in flos Chrysanthemi indici samples were determined simultaneously by high performance liquid chromatography, too. The samples were separated on the same Thermo Hypersil BDS C₁₈ column (4.6 mm×250 mm, 5μm) with the mobile phase of methanol- tetrahydrofuran-water (50:4:46, v/v). The flow rate was 0.6 mL·min^-1. The detection was set at 350 nm and the column temperature at room temperature. The contents of linarin and luteolin were 0.50-2.7% and 0.037-0.09% respectively. With chlorogenic acid as reference substance, HPLC-Fingerprint that can indicate substances existing in flos chrysanthemi indici was established with ultraviolet detection. All constituents were separated completely in 60 min. Peaks existing in chromatograms of all samples were assigned as "common peaks" for flos Chrysanthemi indici. There are 15 "common peaks" in the fingerprint. The fingerprint data were evaluated by Similarity Evaluation System for Chromatographic Fingerprint of TCM of the Chinese Pharmacopoeia Committee. The common pattern was established. Good similarities with cosine coefficient above 0.90 were found in fingerprints of 10 flos chrysanthemi indici samples from different sources. Under the same condition, 6 samples were analyzed. The similarities between their HPLC-FPS and the common pattern of flos chrysanthemi indici were all lower than 0.90.Secondly, to get the standardized extract for injection of flos chrysanthemi indici, the process of extraction and purification was studied. The best process was determined as follows: The herbal material was refluxed in 90℃water bath with 70% ethanol 3 times, 1 h per time, 16 times of 70% ethanol for the first time, 8 times for the second and third time. The extract was combined and was condensed under vaculm. The concentrated solution was diluted to 20 times of the quantity of herbal materials using water. The diluted solution was placed in 90℃water bath for 15 min and then centrifuged for 15 min at 3000 rpm. The supernatant was removed and adjusted to pH 3.0 with 10% hydrochloric acid solution and then purified by macroreticular resin of HPD 300 column (d:h=1:10). The maximum adsorbing capacity was 0.4 g herbal materials per ml resin. The flow rate was 2 BV·h^-1. After washed with 5 BV of 5% ethanol, column was eluted with 5 BV of 50% ethanol. The eluate of 50% ethanol was collected and then condensed under vacumn. The condensed solution was diluted to 20 times of the quantity of herbal materials using water and purified by polyamide column (d:h=1:10). The maximum adsorbing capacity was 0.2 g herbal materials per ml resin. The flow rate was 2BV.h^-1. After washed with 5 BV of water, 70% ethanol was used. The eluate of 70% ethanol were collected and the content of chlorogenic acid and linarin in the purified extract was above 25%.