目的 制备叶酸（FA）修饰的pH值响应型去甲斑蝥素（Nor）脂质体（Nor@LP-CHS-FA），初步评价其促HepG2细胞凋亡活性。方法 应用薄膜分散法制备Nor@LP-CHS-FA，采用单因素实验结合星点设计-效应面法设计和优化制剂处方；研究该载药系统的粒径、ζ电位、多分散系数（PDI）、包封率、载药量，应用透射电镜、傅里叶变换红外光谱、差式扫描量热仪、X射线仪进行物理表征；评价其稳定性及在模拟人工胃液、肠液和肿瘤微环境下的释药情况；通过溶血性实验考察生物安全性，CCK-8法评估Nor@LP-CHS-FA对HepG2细胞的增殖抑制作用，结合流式细胞术分析其对HepG2细胞凋亡、周期的影响。结果 成功制备外观呈黄色的Nor@LP-CHS-FA，其最优处方为脂药比5.05∶ 1、磷脂占膜材总质量的52.85%、FA用量13.90 mg；透射电镜下Nor@LP-CHS-FA呈规则球形，粒径为（55.48±0.67） nm、ζ电位为（-18.15±0.54） mV、PDI为0.42±0.02、包封率（82.72±0.84）%、载药量（12.25±0.13）%； Nor@LP-CHS-FA在4℃条件下储存，包封率、载药量更高，泄漏率更低。与游离Nor相比，Nor@LP-CHS-FA在模拟胃液、肠液和肿瘤微环境释放速度更快，以模拟肿瘤微环境中最快，表明其有pH值响应性，释放行为遵循Weibull模型。溶血性实验结果表明Nor@LP-CHS-FA有较小的溶血率，CCK-8结果显示，给药24、48、72 h后Nor@LP-CHS-FA对HepG2细胞的半数抑制浓度（IC50）分别为8.56、5.89、4.77 μg·mL^-1，流式细胞术结果表明Nor@LP-CHS-FA可诱导HepG2细胞发生凋亡和G2/M期阻滞。结论 Nor@LP-CHS-FA处方工艺简单，具备良好的pH值响应性，能增强药物促HepG2细胞凋亡效果。

Objective To prepare pH-responsive norcantharidin (Nor) liposomes modified with folic acid (FA) (Nor@LP-CHS-FA) and preliminarily evaluate its activity in promoting apoptosis of HepG2 cells. Methods Nor@LP-CHS-FA was prepared by the thinfilm dispersion method. The formulation recipe was designed and optimized through single-factor experiments combined with central composite design-response surface methodology. The particle size, ζ potential, polydispersity index (PDI), encapsulation rate, and drug loading capacity were studied. Physical characterization examinations were conducted using transmission electron microscopy, fourier transform infrared spectroscopy, differential scanning calorimetry, and X-ray. The stability of the system was evaluated, as well as its drug release behavior in simulated artificial gastric fluid, intestinal fluid, and tumor microenvironment. The biological safety was evaluated through hemolysis experiments. The proliferation inhibitory effect of Nor@LP-CHS-FA on HepG2 cells was assessed using the CCK-8 method, and the effects on HepG2 cell apoptosis and cell cycle were analyzed by flow cytometry. Results The yellowcolored Nor@LP-CHS-FA was successfully prepared. The optimal formulation was a lipid-to-drug ratio of 5.05∶ 1, phospholipids accounting for 52.85% of the total mass of the membrane material, and a FA dosage of 13.90 mg. Under transmission electron microscopy, Nor@LP-CHS-FA presented a regular spherical shape, with a particle size of (55.48 ±0.67) nm, a ζ potential of (-18.15 ±0.54) mV, a PDI of 0.42 ±0.02, a encapsulation rate of (82.72 ±0.84)%, and a drug loading of (12.25 ±0.13)%. Nor@LP-CHS-FA had a higher encapsulation rate and drug loading and a lower leakage rate when stored at 4 ℃. Compared with free Nor, Nor@LP-CHS-FA released more rapidly in simulated gastric fluid, intestinal fluid, and tumor microenvironment, and was the fastest in the simulated tumor microenvironment, indicating its pH responsiveness and release behavior following the Weibull model. The hemolysis experiment results showed that Nor@LP-CHS-FA had a lower hemolysis rate. The CCK-8 results indicated that the half-maximal inhibitory concentration (IC₅₀) of Nor@LP-CHS-FA on HepG2 cells after 24, 48, and 72 hours of administration was 8.56, 5.89, 4.77 μg·mL^-1, respectively. The flow cytometry results showed that Nor@LP-CHS-FA could induce HepG2 cells to undergo apoptosis and G2/M phase arrest. Conclusion Nor@LP-CHS-FA prescription process is simple, has good pH responsiveness, and can enhance the effect of promoting HepG2 cell apoptosis by the drug.

R283.6

国家卫生健康委医院管理研究所项目（ YLZLXZ24G039）；四川省自然科学基金面上项目（2025ZNSFSC0684）；云南省科学技术厅地方高校联合专项-面上项目（202001BA070001-041）；云南省天然药物药理重点实验室开放基金重点项目（ YKLPNP-K2302)；四川省中医药管理局科研课题（ 2023MS057, 2023MS207）；四川省医院协会青年药师科研专项资金项目（ 22009, YP2202419）；攀枝花市科学技术局项目（2024ZD-S-28）