The Study On Quality,Material Spectrum And Activities Of Alcohol Extract From Coreopis Tinctoria

Objective:1.The extraction process of Coreopsis tinctoria was further amplified to verify the yield stability and quality reliability of the extracts.2.To establish quality standards of Coreopsis tinctoria alcohol extracts?ETE?according to Chinese pharmacopoeia guidelines.3.To purify and isolate the effective parts of Coreopsis tinctoria alcohol extracts.4.To explore the?-glucosidase inhibitory component of Coreopsis tinctoria.5.To study on anticoagulant activity and its material spectrum of Coreopsis tinctoria extracts.Methods:1.Based on the declaration of Coreopsis tinctoria extraction technology of flavonoids in our research group,to gradually enlarge production10 batch extracts with multi-function extraction unit and multi-stage flash equipment.2.The quality standard of Coreopsis tinctoria alcohol extracts were established by multiple fields,such as TLC,moisture inspection,residue inspection,determination of total flavonoids and total phenolic acid,QAMs aspects and fingerprint method.3.The activity components were separated by extraction,macroporous resin and polyamide column chromatography.The purity products were determined by HPLC area normalization method,and the molecular weight was determined by mass spectrometry.4.The?-glucosidase inhibitor of Coreopsis tinctoria were screened by the ultrafiltration mass spectrometry.5.Bleeding time was tested by the cutting tail in mice,and clotting time was measured by slide method.The ratio of platelet aggregation of ADP and COL induced in healthy volunteers was determined by Born method.The extract of the fingerprint were determined by High performance liquid chromatography?HPLC?,and the change of each component were identified and compared.Furthermore,the possible anticoagulant components were speculated according to dose-response relationship Results:1.The yield was basically stable on amplification process,and the 10 batches of Coreopsis tinctoria extracts were obtained.2.Three main components?marein,flavanokanin and okanin?from Coreopsis tinctoria were identified by TLC.The moisture of Coreopsis tinctoria alcohol extracts should not exceed 15%,and the residue on ignition should not exceed16%.The chlorogenic acid,flavanomarein,marein,3,5-Dicaffeoylquinic acid,total flavonoids and total phenolic acid from Coreopsis tinctoria alcohol extracts should not exceed 0.8%,4%,8.5%,0.5%,9%,28.0%,respectively.The similarity calculation of HPLC map of 10 batches of Coreopsis tinctoria alcohol extracts were calculated,and the similarity of each batch of samples were all exceed 0.9%.3.The content of effective components from Coreopsis tinctoria after gathering by macroporous resin was reached50%.Furthermore,a component?flavanomarein?of flavonoids was isolated from ETEP by polyamide chromatography column.4.The half maximal inhibitory concentration(IC₅₀)of the purified extracts from Coreopsis tinctoria?ETEP?,flavanomarein and marein to?-glucosidase were 0.052,0.892,0.455mg/mL,respectively.The ultrafiltration mass spectrometry results showed that there were six components from Coreopsis tinctoria bonded with?-glucosidase.5.The results of experiments in vivo showed that the bleeding time and blood coagulation time in mice were significantly prolonged by Coreopsis tinctoria alcohol extracts?p<0.01?.Furthermore,the results showed that the aggregation of platelet in healthy volunteer induced by ADP were inhibited by ETE and ETEP?P<0.05?,but the aggregation of platelet induced by COL were inhibited non-significantly.Conclusion:1.The amplified extraction process of Coreopsis tinctoria had the advantages of simple operation,low cost,and the extract rate were 33%.This process can be used for further trials in production and research.2.The quality standards of ETE were established,which provide protection for its industrialization.3.The content of effective components from Coreopsis tinctoria was reached 50%after optimizing the process of isolation and purification.Furthermore,the isolated monomer?flavanomarein?could be used in the study on material basis and mechanism of Coreopsis tinctoria.4.At least 6components from Coreopsis tinctoria had the inhibitory action to?-glucosidase.5.Coreopsis tinctoria could play an anti-clotting effect by inhibiting ADP induced platelet aggregation.In addition,combined with the change of the content of each component in the fingerprint to speculate that the anticoagulant components from Coreopsis tinctoria were purificated chlorogenic acid,flavanomarein,marein and 3,5-Dicaffeoylquinic acid.