Preliminary Study Of The Main Component Of Xuefu Zhuyu Decoction-Contained In Rat Serum

Research BackgroundXuefu Zhuyu Decoction (XFZYD), a representative prescription of promoting blood circulation and removing blood stasis, has the effect of promoting blood circulation to remove blood stasis, and promoting qi to relieve pain. It is mainly used to cure syndrome of chest pain, headache, dysphasia, insomnia and dreaminess, palpitation, irritability and susceptibility which are caused by blood stasis. And it is also used to treat coronary heart disease, angina pectoris, and traumatic headache in modern medicine. The clinical study found that XFZYD can also treat hemorrhagic disease and amenorrhea which caused by the stagnation of the blood. However, recent researches showed that the action mechanism of Xuefu Zhuyu Decoction-Contained serum (XFZYD-CS) may be related to the angiogenesis promotion. But someone considered that the action mechanism of XFZYD may be related to the angiogenesis inhibition. The role of XFZYD has caused widespread attention, it is worthy of studying further whether XFZYD has a two-ways regulation or other causing factors.ObjectiveTo explain the function of XFZYD and further understand its action basis, HPLC, LC-MS/MS, etc were used to analyze the main ingredients of XFZYD-CS and blank serum. Through qualitative and quantitative analysis, paeoniflorin(PF) was the initial screening of the main component in XFZYD-CS. The other major pharmaceutical ingredients in XFZYD-CS will be identified gradually.MethodsSD rats were administrated with Xuefu Zhuyu preparations, the blood samples were collected and serum was separated. The serum samples were analyzed by LC-MS/MS after having pre-processing by acetonitrile protein precipitation method. Chromatographic columns:Atlantis(?) T3(2.1×100mm3μm), column temperature:40℃, flow rate:0.3mL/min, injection volume:2μL, mobile phase:A,0.05%aqueous formic acid; B, methanol, gradient elution. Ion source:electrospray ionization(ESI) ionization source, source temperature:325℃, test mode:detection of negative ions, drying gas flow (Gas Flow):11L/min, nebulizer pressure (Nebulizer):45psi, acquisition conditions:the PF479→120.9, fragmentation voltage:150V, collision voltage:25VResults1. The linear relationship was good with the concentration of PF ranged from9.14to230.0ng/mL. Regression equation was Y=1.1666X-7.2314(r=0.9983). The lowest quantification limit was9.14ng/mL.2. The concentration of PF in XFZYD-CS:the average value of PF in XFZYD-CS of high dose group:55.97ng/mL, SD:13.77%, RSD:24.6%; the average value of PF in XFZYD-CS of the low-dose group:8.38ng/mL, SD:1.84%, RSD:21.9%; the average value of PF in XFZYD-CS of middle dose group:23.77ng/mL, SD:5.58%, RSD:23.5.3. The concentration values of PF in XFZYD-CS of low, medium and high dose group were significantly different.ConclusionThe blank serum and ingredients of XFZYD-CS were determined by HPLC, LC-MS/MS. The determination method of PF in rats’ serum was established by using LC-MS/MS, the PF concentration in XFZYD-CS were also analyzed.The results were statistically analyzed; the concentrations of PF in XFZYD-CS of low, medium and high dose group were significantly different. However, whether the differences of PF concentration in XFZYD-CS were the basis of action mechanism of XFZYD should be further studied.