摘要
本文评价了两亲性p H敏感聚合物聚(2-乙基-2-噁唑啉)-胆固醇氯甲酸酯[poly(2-ethyl-2-oxazoline)-cholesteryl methyl carbonate,PEOZ-CHMC]的缓冲能力。采用硫酸铵梯度法制备盐酸阿霉素脂质体(doxorubicin hydrochloride liposomes,DOX-L),后插入法制备PEOZ-CHMC和聚乙二醇-二硬脂酰磷脂酰乙醇胺(polyethylene glycol-distearoyl phosphatidyl ethanolamine,PEG-DSPE)修饰的盐酸阿霉素脂质体(PEOZ-DOX-L和PEG-DOX-L),对盐酸阿霉素脂质体的理化性质、体外释放、细胞抑制作用和细胞内定位情况进行评价。结果显示,PEOZ-CHMC对微酸环境有一定的缓冲能力。利用葡聚糖凝胶微柱离心法测定脂质体的包封率为(97.3±1.4)%,动态光散射法测定脂质体的粒径在120 nm左右,经PEG-DSPE和PEOZ-CHMC修饰后脂质体的包封率和粒径均无明显改变。Zeta电位分析仪结果显示3种脂质体表面电荷均为负值。通过透析实验考察制剂的体外释药行为,结果表明,在p H 7.4的释放介质中,PEOZ-DOX-L释放较为缓慢,而在p H 5.0时,PEOZ-DOX-L快速释药,显示出良好的p H敏感性。激光共聚焦实验表明核内体的微酸环境可以促进脂质体发生核内体逃逸,直接释放盐酸阿霉素进入细胞核,而DOX-L和PEG-DOX-L均无此作用。细胞抑制作用结果显示,PEOZ-DOX-L对细胞的抑制作用随p H降低而增强,但p H的降低对PEG-DOX-L和DOX-L的细胞抑制作用没有影响。因此PEOZ-CHMC构建的p H敏感脂质体能克服PEG链抑制脂质体的细胞摄取和妨碍p H敏感脂质体的核内体逃逸问题。
In this study, the buffering capacity of amphiphilic p H-sensitivity copolymer poly（2-ethyl-2- oxazoline）-cholesteryl methyl carbonate（PEOZ-CHMC） was evaluated. The ammonium sulfate gradient method was used to prepare doxorubicin hydrochloride（DOX·HCl）-loaded liposomes（DOX-L）, and then the post-insertion method was used to prepare PEOZ-CHMC and polyethylene glycol-distearoyl phosphatidyl ethanolamine（PEG-DSPE） modified DOX·HCl-loaded liposomes（PEOZ-DOX-L and PEG-DOX-L）. The physico-chemical properties, in vitro drugs release behavior, cellular toxicity and intracellular delivery of liposomes were evaluated, separately. The results showed that PEOZ-CHMC has a satisfactory buffering capacity. The sephadex G-50 column centrifugation method and dynamic light scattering were used to determine the encapsulation efficiency（EE） and particle size of liposomes. The EE and particle size of DOX-L were（97.3 ± 1.4） % and 120 nm, respectively, and the addition of PEOZ-CHMC or PEG-DSPE had no influence on EE and particle size. The zeta potentials of three kinds of liposomes were negative. The release behavior of various DOX liposomes in vitro was investigated by dialysis method. In phosphate buffer solution（PBS） at p H 7.4, DOX·HCl was released from PEOZ-DOX-L in a sustained manner. While in PBS at p H 5.0, the release rate of DOX·HCl from PEOZ-DOX-L increased significantly, which suggested DOX·HCl was released from PEOZ-DOX-L in a p H-dependent manner. The intracellular delivery of liposomes was investigated by confocal laser scanning microscopy（CLSM）. The CLSM images indicated that PEOZ-DOX-L showed efficient intracellular trafficking including endosomal escape and release DOX·HCl into nucleus, as well as the DOX-L and PEG-DOX-L had no this effect. The cytotoxicity of liposomes against MCF-7 cells was detected by using MTT assay. The results showed that antiproliferative effects of PEOZ-DOX-L enhanced with p H value decreased, whereas DOX-L and PEG-DOX-L did not have any significa
出处
《药学学报》
CAS
CSCD
北大核心
2015年第9期1174-1179,共6页
Acta Pharmaceutica Sinica
基金
国家自然科学基金资助项目(81102394)
中国博士后科学基金资助项目(2013M530899)
