Preparation And Preliminary Pharmacokinetics Study Of Pulsatilla Saponin D Albumin Nanoparticles

Pulsatilla Saponin D is a kind of saponin separated from Pulsatilla chinensis,which has a strong anti-tumor activity.Due to its low solubility,and the hemolysis is strong,which seriously affects the formation of the drug.In order to solve the problems of low solubility and strong hemolysis of Pulsatilla Saponin D,albumin nanoparticles were prepared from Pulsatilla Saponin D.The preparation process,quality evaluation,hemolytic characteristic test and pharmacokinetic test of the preparation were preliminarily studied.The specific research contents and conclusions are as follows:1.High performance liquid chromatography（HPLC）method was used to study the content of Pulsatilla Saponin D.The solubility of Pulsatilla Saponin D in different concentrations of ethanol was determined.The effects of light,humidity and heat on the Pulsatilla Saponin D were studied.The factors influencing the preparation of blank albumin nanoparticles were investigated.The method of determining the content of Pulsatilla Saponin D is accurate,effective and reliable.The results showed that the solubility of Pulsatilla Saponin D in different ethanol concentrations increased with the increase of the nonpolarity of the solvent,which provided a basis for the rational selection of the solvent for the following preparations.In the experiment of high humidity,it shows that the Pulsatilla Saponin D has certain hygroscopicity and volatility,so it should be sealed and preserved.The results showed that the ionic strength and p H had some influence on the size and distribution of the nanoparticles.It laid a foundation for the preparation of follow-up preparation,reduced the risk of follow-up preparation research,and improved the success rate of the experiment.2.HPLC was used to determine the content of Pulsatilla Saponin D in Pulsatilla Saponin D albumin nanoparticles.Albumin nanoparticles of Pulsatilla Saponin D were prepared by desolvent method.The formulation and preparation technology of the drug loaded nanoparticles were investigated by single factor experiment.The Box-Behnken experimental design and the response surface were used to optimize the preparation of Pulsatilla Saponin D albumin nanoparticles.HPLC method for the determination of Pulsatilla Saponin D in albumin nanoparticles is simple,accurate and feasible.It can be used to determine the drug loading and entrapment efficiency of albumin nanoparticles,which lays a foundation for the optimization of subsequent preparation prescription and process,as well as the quality evaluation and control of preparations.Single factor experiment was carried out to investigate the influence of related prescription and process factors on drug loaded nanoparticles.The dosage of albumin,the dosage of organic phase and the dosage of drug were screened out.Through optimization,the optimal preparation conditions were determined as follows:the dosage of albumin was 100mg,the dosage of organic phase was 1m L,and the dosage of Pulsatilla Saponin D was 22.78mg.3.Study on the characterization and quality evaluation of Pulsatilla Saponin D albumin nanoparticles prepared with the optimal formulation.Scanning electron microscope and transmission electron microscope were used to observe the morphology of Pulsatilla Saponin D albumin nanoparticles.The particle size and distribution of Pulsatilla Saponin D albumin nanoparticles were determined by Malvern laser particle size analyzer.The drug loading and entrapment efficiency of Pulsatilla Saponin D albumin nanoparticles were determined by HPLC.The formation of nanoparticles was characterized by IR and X-powder diffraction.The solubility of nanoparticles in water was determined by shaking table method.The release behavior of nanoparticles in vitro was measured by dynamic dialysis.The hemolysis of nanoparticles was evaluated by enzyme labeling instrument,and the preliminary stability was investigated.The morphology of albumin nanoparticles was round,well dispersed and well distributed.The drug loading and encapsulation efficiency were17.5%and 94.5%.The results of FTIR and X-powder diffraction showed that there was no obvious crystal character of albumin nanoparticles,and the drugs were mainly encapsulated in carrier materials.After the preparation of albumin nanoparticles,the hemolysis of Pulsatilla Saponin D was lower than that of the drug substance,higher solubility than drug substance.There was no sudden release effect in 2 hours,and it had a slow-release effect.4.Preliminary study on the pharmacokinetics of Pulsatilla Saponin D albumin nanoparticles.LC-MS/MS method was established for the determination of Pulsatilla Saponin D in biological samples of rats.The blood concentration of Pulsatilla Saponin D albumin nanoparticles was measured at different time points with different doses were injected into tail vein.Data fitting analysis with Das 2.0 software.LC-MS/MS quantitative detection method conforms to the guiding principle of biological sample analysis.The analytical time of this method is within 2.5min,and the linear range is 10～1 500 ng/m L.The sample processing method is simple and specific.The time of T_1/2 was longer than that of drug substance and MRT takes longer than drug substance.It shows that the preparation has a sustained-release effect.AUC shows that the preparation is 5 times of the drug substance.It shows that the preparation is more efficient than the drug substance.