【摘要】 实验以聚天冬氨酸(PASP)为载体,以甲硝唑为模型药物,用较温和简便的方法制备了新型聚天冬氨酸-甲硝唑(PASP-MTI)纳米前药.采用红外光谱及核磁氢谱分析、透射电镜观察、激光粒度、透析和紫外分光光度测定、MTT比色、荧光显微镜和流式细胞术等方法,观察到甲硝唑以酯键方式键合于聚天冬氨酸高分子链上.PASP-MTI纳米前药呈球型,平均粒径404.8nm,载药量30%,体外释药明显延缓;MTT结果显示,PASP-MTI显著提高甲硝唑对滴虫的抑杀作用,经纳米甲硝唑作用后,部分滴虫胞核出现染色质浓集、核固缩、核碎裂等一系列凋亡改变.流式细胞术检测可见凋亡峰,凋亡率由对照组的11.5%上升到纳米前药组的35.69%.上述实验结果表明:PASP是一种非常有潜力的前药载体,制得的新型PASP-MTI纳米前药具有载药量高、缓释和杀虫作用强等特点,可能的杀虫机制除与高聚物纳米前药促吞噬作用有关外,还与诱导滴虫凋亡有关.更多还原

【Abstract】 Poly aspartic acid or its salts is a kind of newly innocuous, environmental friendly biodegradable polymer, recognized as green material, and widely applied in such areas as agriculture, medicine, commodity, water treatment. In order to prepare new poly aspartic acid-metronidazole (PASP-MTI) nanoprodrug, observe its trichomonacidal effect in vitro, and explore its probable mechanism, the nanoprodrug PASP-MTI was synthesized by DL-aspartic acid and metronidazole in a relatively mild condition. Its structural characteristics were attested by means of infrared spectrum (IR), 1H nuclear magnetic resonance (1H-NMR), transmission electron microscope (TEM) etc measurements. The release behavior of metronidazole in PASP-MTI nanoprodrug was investigated by the dialysis method. The MTT assay was employed to test PASP-MTI’s trichomonacidal effect in vitro and the apoptotic-like processes of trichomonas were detected by fluorescence microscope and flow cytometer (FCM). The IR and 1H-NMR showed that the metronidazole was conjugated to poly aspartic acid high molecular chains by ester bonds. Synthesized PASP-MTI nanoprodrug was globular and had a mean diameter 404.8 nm with favorable dispersity. The drug loading was about 30% and the cumulative release profile was delayed (about 38.01% of MTI loading in the PASP-MTI nanoprodrug was released within the first 24 h and 86.64% in 30 d, while the release of free metronidazole was almost complete by 24 h). The trichomonacidal activity of PASP-MTI nanoprodrug increased remarkably{IC50 of PASP-MTI and free metronidazole was 1.22 mg/L and 2.85 mg/L, respectively}. Observed by fluorescence microscope, the nuclei of trichomonas presented a series of changes similar to apoptosis such as chromatin agglutination, nuclear fragmentation, etc. The FCM study indicated that the apoptotic rate in the PASP-MTI group was the highest of all groups, up to 35.69%. It is concluded that PASP can be effectively used in drug delivery systems, the synthetic novel poly aspartic acid-metronidazole nanoprodrug has higher drug loading, better the role of prolonged-release and higher trichomonacidal effect in vitro. Its mechanism may be related with enhancing the polymer prodrug’s internalization and inducing the apoptosis of trichomonas.更多还原