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目的制备载盐酸小檗碱(berberine hydrochloride,BH)的叶酸(folic acid,FA)偶联壳聚糖(chitosan,CTS)纳米粒(BH/FA-CTS-NPs),优化制备工艺并考察BH/FA-CTS-NPs的理化特性及其对CNE-1细胞的抑制作用。方法利用FA活性酯与CTS分子上的氨基反应,制得FA偶联CTS(FA-CTS);BH为模型药物,通过离子交联法制备BH/FA-CTS-NPs。考察其形态、粒径、包封率、载药量及体外释放等理化特性;分别通过MTT法、划痕实验和Annexin-V-FITC单染法进行检测,验证BH/FA-CTS-NPs对CNE-1细胞的抑制作用、抗增殖侵袭能力以及诱导凋亡作用。结果透射电镜下观察所得BH/FA-CTS-NPs形态外观圆整,大小均匀,无粘连,平均粒径为(282.4±4.5)nm;包封率为(89.82±2.91)%;载药量为(9.16±1.01)%;5 h内累积释药率为(80.32±3.24)%,随后缓慢释放,24 h内累积释药率为(90.92±5.21)%。CNE-1细胞实验显示,BH/FA-CTS-NPs能够显著抑制CNE-1细胞的生存和增殖能力,诱导细胞CNE-1凋亡,具有剂量和时间依赖性。结论离子交联法成功制备具有体外缓释作用的BH/FA-CTS-NPs,形态圆整,粒径大小均一,对抑制CNE-1细胞增殖及促进凋亡效果好,为开发抗肿瘤给药系统提供了理论依据。
OBJECTIVE To prepare folic acid (FA) coupled chitosan (CTS) nanoparticles (BH / FA-CTS-NPs) loaded with berberine hydrochloride (BH) Physical and chemical properties of FA-CTS-NPs and their inhibitory effect on CNE-1 cells. Methods The FA-CTS (FA-CTS) was synthesized by reacting the active ester of FA with the amino group of CTS molecule. BH was used as a model drug to prepare BH / FA-CTS-NPs by ion-crosslinking method. The morphology, particle size, entrapment efficiency, drug loading and release in vitro were investigated. MTT assay, scratch assay and Annexin-V-FITC single staining assay were used to verify the effect of BH / FA-CTS-NPs CNE-1 cells, antiproliferative and invasive ability, and apoptosis induction. Results The morphology of the BH / FA-CTS-NPs observed by transmission electron microscopy was round and uniform in size with no adhesions. The average particle size was (282.4 ± 4.5) nm. The encapsulation efficiency was (89.82 ± 2.91) (9.16 ± 1.01)%. The cumulative release rate within 5 h was (80.32 ± 3.24)%, and then slowly released. The cumulative release rate within 24 h was (90.92 ± 5.21)%. CNE-1 cells showed that BH / FA-CTS-NPs could significantly inhibit the survival and proliferation of CNE-1 cells and induce apoptosis of CNE-1 cells in a dose- and time-dependent manner. Conclusion The ionic crosslinking method has been successfully used to prepare BH / FA-CTS-NPs with sustained release in vitro. The morphology of the BH / FA-CTS-NPs is uniform and the size of the particles is uniform. It is effective in inhibiting the proliferation and promoting apoptosis of CNE- System provides a theoretical basis.