Preparation And Quality Evaluation Of Novel Evodiamine Nanoparticles

Evodiamine（EVO）is an alkaloid which isolated from the Chinese medicine Evodia,it has various pharmacological activities,such as anti-inflammatory,lower blood pressure,loss weight,anti-tumor and so on.Because of poor water solubility and low bioavailability,the clinical application of EVO is limited.In this study,novel evodiamine nanoparticles（NENP）were prepared by ionic gelation method,and it was hoped to increase the oral bioavailability of EVO.We established HPLC analysis method to determine the content of EVO in vitro and in vivo,characterized the physicochemical properties of the NENP,and investigated the pharmacokinetics of NENP and EVO.The first part,EVO content determination method in vitro.Objective:To establish a method for determination and analysis of EVO in NENP.Method:The separation was performed on COSMOSIL 5C18-MS-II（250mm×4.6 mm,5μm）column,10 mmol·L^-1ammonium acetate-acetonitrile（40:60,v/v）as mobile phase,the detection wavelength was 225 nm and flow rate was 1.0 mL·min^-1,the column temperature was 30°C,the sample volume was 20μL.Results:Different excipients in NENP did not affect the content determination of EVO.The concentration of EVO at the range of 0.2²0μg·mL^-1,it had a good linear relationship between the peak area and concentration.The linear regression equation was A=241702C+21970,r=0.9999.The limit of detection was 1 ng·mL^-1.The intra-day and inter-day relative standard deviations in different concentrations were at the range of0.13%².04%.The average recoveries were at the range of 97%¹00%（RSD were less than 0.8%）.Conclusion:The method of determination of EVO in NENP was reliable.The second part,preparation and physicochemical properties of NENP.Objective:To prepare NENP and characterize the physicochemical properties of NENP.Methods:NENP was prepared by ionic gelation method.The physicochemical properties of NENP,such as particle size and Zeta potential were characterized.Results:The size of NENP was435.30±24.70 nm,PDI was 0.30±0.02,the Zeta potential was 43.43±0.50mV,and the entrapment efficiency was 89.35±1.02%.Conclusion:The particle size distribution of NENP was uniform and the prepared NENP was stable.The third part,content determination of EVO in vivo.Objective:To establish an analytical method for the determination of EVO in plasma of Sprague Dawley rats.Method:Using Honokiol（HK）as internal standard,the separation was performed on COSMOSIL 5C18-MS-II（250 mm×4.6mm,5μm）column,10 mmol·L^-1ammonium acetate solution-acetonitrile（40:60,v/v）as mobile phase,detection wavelength was 225 nm and flow rate was 1.0 mL·min^-1,the column temperature was 30°C,the sample volume was 40μL.Results:The separation degree of EVO and HK was more than 1.5.Endogenous substances in plasma did not influence the content determination of EVO.At the range of 8⁶40 ng·mL^-1,the linear relationship between the ratio of the EVO peak area to the internal standard HK peak area（Y）and concentration（C）of EVO in plasma was good.The linear regression equation of EVO was Y=0.0124C+0.0083,r=0.9994.The limit of detection was 3 ng·mL^-1.The extraction recoveries of different concentrations of EVO(8,80,640 ng·mL^-1)were at the range of 80⁹3%（RSD were less than 5.5%）;the method recoveries were at the range of 94%⁹9%（RSD were less than 6.5%）.Conclusion:The method of determination of EVO in NENP is reliable.The fourth part,pharmacokinetics evaluation of NENP.Objective:To investigate the pharmacokinetic characteristics of NENP in Sprague Dawley（SD）rats.Methods:SD rats were orally administered of NENP and EVO,respectively.After getting blood sample,we measured the concentration of NENP and EVO.The main pharmacokinetic parameters of NENP and EVO were calculated by DAS 2.1.1 pharmacokinetic software,at the same time evaluated bioequivalence of NENP and EVO.Results:The C_max of NENP was about 1.49 times than EVO.The relative bioavailability of the NENP in non-compartment model was 285%than EVO.Conclusion:The bioavailability of EVO was improved after preparing into NENP,and the bioavailability between NENP and EVO were not equivalent.NENP was better than EVO.