叶酸修饰的斑蝥素/黄芩苷共载脂质体处方工艺优化及其评价

Optimization and evaluation of preparation process of folate-modified cantharidin/baicalin co-loaded liposomes

目的筛选二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000-叶酸(DSPE-PEG 2000 folate,DSPE-PEG2k-FA)修饰的斑蝥素/黄芩苷共载脂质体(DSPE-PEG2k-FA-cantharidin&baicalin-lipsomes,FA-Can&Bai-Lips)的处方工艺并对其进行表征及评价。方法由薄膜分散超声法制备FA-Can&Bai-Lips,以药脂比、胆脂比、水化温度及水化时间为考察因素,以两药包封率、粒径、多分散指数(polydispersity index,PDI)和ζ电位为评价指标,采用单因素考察和Box-Behnken设计-响应面法(BoxBehnken design-response surface methodology,BBD-RSM)优化其处方工艺。采用电位粒径仪和透射电子显微镜(TEM)分别测定其粒径、ζ电位及形态。通过溶血性实验、细胞摄取实验、CCK-8法实验及活体成像等方法进行安全性、靶向性评价。结果FA-Can&Bai-Lips的最佳制备工艺为药脂比1∶11、胆脂比1∶6、DSPE-PEG2k-FA用量为10%、溶剂为50 mL、旋转蒸发及水化温度为55℃、水化介质为pH 6.8 PBS缓冲液、水化体积为10 mL、水化时间为94 min,超声时间为8 min。经BBD-RSM优化制得的FA-Can&Bai-Lips形态规整,为类球形粒子,稳定性及安全性好,斑蝥素与黄芩苷的包封率分别为(90.19±0.67)%、(81.24±0.72)%,粒径为(172.47±4.83)nm,PDI为0.268±0.018,ζ电位为(−1.07±0.07)mV。体外评价结果表明,FA-Can&Bai-Lips能有效抑制HepG2细胞增殖,具良好的肝肿瘤靶向性。结论FA-Can&Bai-Lips具有增效减毒作用,能高效靶向肿瘤组织,是一种极具开发潜力的抗肝肿瘤纳米递送系统。

Objective To screen the prescription process of DSPE-PEG 2000 folate(DSPE-PEG2K-FA)modified cantharidin/baicalin co-loaded liposomes(FA-Can&Bai-Lips),and to characterize and evaluate it.Methods FA-Can&Bai-Lips were prepared by thinfilm dispersion sonication,and their prescription process was optimized by one-factor investigation and the Box-Behnken designresponse surface methodology(BBD-RSM)using drug-lipid ratio,bile-lipid ratio,hydration temperature,and hydration time as the investigating factors,and encapsulation rate of the two drugs,particle size,polydispersity index(PDI),andζpotential as the evaluating indexes.The particle size,potential,and morphology were determined by potentiometric particle sizer and transmission electron microscopy(TEM).Safety and targeting were evaluated by hemolytic assay,cell uptake assay,CCK-8,and in vivo imaging.Results The optimal preparation process of FA-Can&Bai-Lips was as follows:the drug-lipid ratio was 1:11,the bile-lipid ratio was 1:6,the amount of DSPE-PEG2k-FA was 10%,solvent was 50 mL,rotary evaporation and hydration temperature was 55℃,hydration medium was pH 6.8 PBS buffer,hydration volume was 10 mL,hydration time was 94 min,ultrasonication time was 8 min.The FA-Can&BaiLips optimized by the BBD-RSM had regular morphology,spherical particles,good stability and safety,and the encapsulation rates of cantharidin and baicalin were(90.19±0.67)%and(81.24±0.72)%,respectively,and the particle sizes were(172.47±4.83)nm,with a PDI of 0.268±0.018,and aζpotential of(−1.07±0.07)mV.The in vitro evaluation results showed that FA-Can&Bai-Lips could effectively inhibit the proliferation of HepG2 cells,bearing good liver tumor targeting.Conclusion FA-Can&Bai-Lips has potentiation and toxicity reduction effects,can efficiently target tumor tissues,and is an anti-hepatic tumor nano-delivery system with great potential for development.