Study On Chemical Constituents And Bioactivities Of Safflower Injection

Safflower injection made from safflower by hydro-decoction and alcohol precipitation.It has the functions of blood circulation,anti-oxidation,lipid-lowering,and anti-inflammatory,and is mainly used for the prevention and treatment of coronary heart disease,vasculitis,occlusive cardiovascular,cerebrovascular diseases,myocardial infarction and other diseases.At present,there are many reports on clinical research and pharmacological studies about safflower injection.However,there are few studies on its active ingredients and they mainly focus on hydroxysafflor yellow A?HSYA?.To further elucidate the pharmacological substance basis of safflower injection,this study evaluated the prolonged activated partial thromboplastin time?APTT?,prothrombin time?PT?,platelet inhibition activity,and antioxidant activity of safflower injection,and based on these biological activity indicators,activity-guided isolation method was used for separation.The anticoagulant active site and chemical composition of safflower injection were studied.1.The anticoagulant activity of safflower injection produced by seven companies was evaluated by prolonging the normal plasma activated partial thromboplastin time?APTT?,prothrombin time?PT?and in-vitro inhibition of platelet aggregation.The in-vitro anti-oxidation activity of safflower injection was studied using the screening model of scavenging 1,1-Diphenyl-2-picrylhydrazyl free radical?DPPH??.The content of total polyphenols and total flavonoids in safflower injection was determined by Folin-Phenol method and AlCl₃ colorimetric method.The results showed that safflower injections from different manufacturers had a prolonged effect on APTT?P>0.05?but PT was not significant?P<0.05?.That is,safflower injection had a significant effect on endogenous coagulation pathways.While no significant effect on the coagulation pathway.Besides,it can significantly prolong ADP-induced platelet aggregation in rabbits.And in a certain range,its DPPH?scavenging effect was dose-dependent.2.The macroporous resin D101 column was used to crudely separate the safflower injection and obtain four components.Then the biological activity of each component was evaluated by in-vitro biological activity screening method.The experimental results showed that the four chemical constituents all had the effect of prolonging APTT,PT and inhibiting platelet aggregation in vitro,and the order of prolonging APTT and PT was Fr₄>Fr₃>Fr₂>Fr₁.Furthermore,Fr₃ had strong inhibition platelet aggregation,antioxidant activity,and high total polyphenol content.3.The separation and purification by Sephadex LH-20 and preparation phase were repeated,and three compounds were isolated fromFr₄.Thethreecompoundswereidentifiedas p-hydroxybenzaldehyde,p-hydroxycinnamic acid and?8Z?-decaene-4,6-diyne-3-O-?-D-glucoside by UPLC-MS and ¹H NMR,¹³C NMR and standard comparisons.The results of prolonged APTT activity showed that the activity of p-hydroxycinnamic acid was the strongest,followed by p-hydroxybenzaldehyde.4.The contents of hydroxysafflor yellow A,p-hydroxycinnamic acid and p-hydroxybenzaldehyde which were the main chemical constituents of safflower injection were determined by HPLC.Pearson's method was used to compare the correlation between the anticoagulant activity of safflower injection,inhibition activity of platelet aggregation and antioxidant activity and its content of total flavonoids and total polyphenol.And the content of hydroxysafflor yellow A and in vitro anticoagulation were also compared and analyzed.Overall,the dose-effect relationship analysis of blood activity showed that there was a certain correlation between the total flavonoid content and the APTT prolongation rate,and the total polyphenol content had a good correlation with the DPPH?scavenging rate.However,there was no significant dose-response relationship between the content of hydroxysafflor yellow A and APTT prolongation rate.The conclusion was that the chemical basis of APTT prolonging activity may be its flavonols and flavonol glycosides,and the chemical basis of in vitro antioxidant activity may be polyphenols.