Preparation Of The Extracts Of Clinopodium Herb And Preliminary Pharmacokinetic Study Of Buddlejasaponin IVb

The Clinopodium Herb has the effect of astringent and hemostatic,and is often used for the treatment of functional uterine bleeding,postpartum hemorrhage and other symptoms,but there are few reports about the material basis of the hemostatic effect of the Clinopodium Herb.Based on the effect of Clinopodium Herb in the treatment of functional uterine bleeding,the research group obtained the compound buddlejasaponin IVb from Clinopodium Herb.Some researchers have previously studied the effects of buddlejasaponin IVb on platelet aggregation and thrombin time?TT?in rabbits in vitro.The results proved that buddlejasaponin IVb Promote the aggregation of rabbit platelets and significantly shorten the thrombin time?TT?,indicating that buddlejasaponin IVb can reduce blood flow by promoting platelet aggregation to accelerate thrombosis,and then play a role in hemostasis,and can affect the role of fibrinogen in plasma.Hemorrhagic buddlejasaponin IVb is the research object,and the pharmacokinetics,intestinal absorption,plasma protein binding rate,and tissue distribution of hemorrhagic buddlejasaponin IVb in the body are systematically studied to clarify the in vivo treatment process of buddlejasaponin IVb It provides experimental basis for the medicinal properties,new drug development,and drug delivery strategy of buddlejasaponin IVb in the bloodstream,and lays the foundation for the research on the Clinopodium Herb and new hemostatic drugs.1.Preparation of The extracts of Clinopodium Herb and determination of Buddlejasaponin IVb contentIn this experiment,an active site with high blood flow and blood flow saponin a content was first prepared,and then repeatedly purified using macroporous resin to prepare5.12%-5.60%Hemorrhagic buddlejasaponin IVb?Hepatic Extract??Dry Extract?.The blood stream extract prepared in this experiment will provide a sample basis for the pharmacokinetics of Hemorrhagic buddlejasaponin IVb.2.Pharmacokinetic study of Buddlejasaponin IVbFirstly,A UPLC-MS/MS method was established for the determination of Buddlejasaponin IVb in rat plasma.Isoquercetin was selected as the internal standard.Mass spectrometry was performed using an electrospray ion?ESI?source and a multi-reaction ion monitoring?MRM?was used in the positive ion mode.The results showed that Buddlejasaponin IVb had a good linear relationship within the range of37.5-7500 ng·m L^-1,with r²>0.99,and the LLOD is 0.1 ng·m L-1,the lower limit of quantification is 0.4 ng·m L-1,and the recovery was greater than 85%.The established method has good specificity,precision,accuracy and stability and there was no obvious matrix effect,which meet the requirements of biological sample analysis.SD rats intragastric administration of high,medium and low doses of Clinopodium Herb extract?interrupted Clinopodium Herb extract?dry extract?interrupted blood flow buddlejasaponin IVb content is 5.37%?(0.36,0.72,1.44 g·Kg^-1),to determine the blood concentration of buddlejasaponin IVb in the rat's blood after administration,and calculate the pharmacokinetic parameters and absolute bioavailability using DAS 2.0.The peak time?Tmax?of buddlejasaponin IVb after the interrupted Clinopodium Herb extract were?1.75±0.42?h,?2.25±0.27?h,?2.83±0.26?h,and the half-life(t_1/2)were?4.61±1.84?h,?5.28±1.78?h,?6.38±1.22?h,the intravenous injection of t_1/2is?0.76±0.11?h.The results show that the buddlejasaponin IVb absorption is slower,the elimination rate is faster.3.The binding rate of Buddlejasaponin IVb in blood plasma with rat plasma proteinIn this study,the binding of buddlejasaponin IVb with rat plasma protein was investigated,and A method for the determination of buddlejasaponin IVb in interrupted phosphate buffer?PBS?by UPLC-MS/MS was established.Glibenide was used as the internal standard and electrospray?ESI?was used as the ion source.The results showed that there was a good linear relationship in the range of 0.1-716.8 ng·m L-1,all r²were greater than 0.99,the quantitative lower limit was 0.1 ng·m L-1,the recovery rate was greater than 85%,and the specificity,precision,accuracy and stability of the method all met the requirements of the experiment.The results showed that the binding rate of buddlejasaponin IVb to plasma proteins was?43.86-50.79?%.4.Tissue distribution of Buddlejasaponin IVbIn this study,UPLC-MS/MS was first used to determine the content of buddlejasaponin IVb in the homogenate of rat tissues?heart,liver,spleen,lung,kidney,brain,stomach,uterus,small intestine and large intestine?.The internal standard was glibenide,and the ion source was electrospray?ESI?.The results showed that the mass concentration of buddlejasaponin IVb in each tissue homogenate?heart,liver,spleen,lung,kidney,uterus,brain?ranged from 0.8 to 307.2 ng·m L-1,showing A good linear relationship.The mass concentration of buddlejasaponin IVb in each tissue homogenate?stomach,small intestine,large intestine?ranged from 2 to 3500 ng·m L-1,showing good linearity,with r²greater than 0.99,and the lower limit of quantification being 2 ng·m L-1.The recovery rate of>was 85%,and the results of methodological investigation all met the requirements of biological sample analysis,and there was no obvious matrix effect.After intragastric administration,the rats were given 0.72 g·Kg^-1of interrupted blood flow extract?dry extract??crude dose/body weight of the rats?(equivalent to 38.66 mg·kg^-1of interrupted blood flow buddlejasaponin IVb),and the distribution of interrupted blood flow buddlejasaponin IVb in different tissues was determined.The results showed that buddlejasaponin IVb was widely and rapidly distributed in rats,but it was difficult to cross the blood-brain barrier and reach the brain tissue.At 0.5 h after administration,the concentration of buddlejasaponin IVb in all tissues reached the maximum value,and then the concentration of buddlejasaponin IVb in all tissues showed A decreasing trend with the extension of time.The concentration of buddlejasaponin IVb in the stomach and intestine was significantly higher than that in other tissues,indicating that the buddlejasaponin IVb in the broken blood stream could be quickly absorbed by the stomach and intestine.Moreover,7 h after administration,the concentration of buddlejasaponin IVb in the gastrointestinal tract was still higher than that in other tissues and organs.This result suggested that the distribution of buddlejasaponin IVb in the gastrointestinal tract may have gastrointestinal accumulation,which may easily lead to low blood drug concentration and low bioavailability.