脑靶向脂质体构建及其靶分子Ac_4MAN体外结构稳定性研究

Preparation of braining-targeting liposomes and the structural stability of Ac_4MAN on the surface of liposome in vitro

目的构建修饰的Ac_4MAN(p-羧基苯-α-D-乙酰甘露糖)脑靶向脂质体,并探讨Ac_4MAN结构稳定性。方法将Ac_4MAN与猪肝酯酶(PLE)于37℃、pH 7.4条件下孵育,于不同时间点取样;采用高效液相色谱(HPLC)和超高效液相色谱-质谱(UPLC-MS)联用技术对其进行分析。制备Ac_4MAN脂质体并对其进行表征,用流式细胞仪检测脑胶质瘤细胞U87摄取Ac_4MAN脂质体和Ac_4MAN脂质体加PLE的情况。结果 HPLC结果显示,孵育0～3 h,Ac_4MAN峰面积从578.4减少到37.9;孵育4～7 h,Ac_4MAN峰面积接近于0;孵育0～7 h,生成的MAN(p-氨基苯-α-D-吡喃甘露糖苷)的峰面积从0增长到363.0,表明随孵育时间延长,Ac_4MAN逐渐减少,MAN逐渐增多。结论本研究制备的Ac_4MAN脂质体粒径约为(120.3±2.0)nm,电位约为(-15.76±1.23)mV。Ac_4MAN脂质体中加入酶后,U87细胞对其摄取能力有一定的增强。

Objective To construct a modified Ac4MAN (p-carboxybenzene-α-D-acetylmannose) brain-targeted liposome and investigate the structural stability of Ac4MAN. Methods Ac4MAN was incubated with porcine liver esterase (PLE) at 37°C, pH 7.4 and sampled at different time points. High performance liquid chromatography (HPLC) and ultra performance liquid chromatography-mass spectrometry (UPLC-MS) were used to analyze it. Ac4MAN liposomes were prepared and characterized. Flow cytometry was used to detect the uptake of Ac4MAN liposomes and Ac4MAN liposomes plus PLE by glioma cells U87. Results The results of HPLC showed that the area of Ac4MAN decreased from 578.4 to 37.9 after incubation for 0-3 h. The area of Ac4MAN peak was close to 0 after incubation for 4-7 h. The peak area of the formed MAN (p-aminobenzene-α-D-pyran mannoside) increased from 0 to 363.0, indicating that with the prolongation of incubation time, Ac4MAN gradually decreased and MAN gradually increased. Conclusion The Ac4MAN liposome prepared in this study has a particle size of (120.3±2.0) nm and a potential of (-15.76±1.23) mV. The uptake of AC4MAN liposomes by U87 cells is enhanced after adding enzymes.