海参皂苷nobiliside A脂质体及其溶血行为的初步研究

Preliminary study on sea cucumber saponin-nobiliside A liposome and its hemolytic activity

本文旨在制备海参皂苷nobiliside A(Nob A)脂质体,建立脂质体中Nob A的含量和包封率的测定方法,考察该脂质体的体内外溶血行为。以高效液相色谱-蒸发光散射检测法(HPLC-ELSD)测定脂质体中Nob A含量,微柱离心法测定其包封率。以溶血率和包封率、粒径为指标,单因素考察了制备方法、磷脂用量、胆固醇用量和药脂比。根据单因素的考察结果,以薄膜超声法制备3批脂质体,与同浓度的药物溶液进行体内外溶血行为的比较。结果显示:用于含量测定和包封率测定的HPLC-ELSD和微柱离心法准确、快速,采用薄膜超声法制备的脂质体形态圆整,粒径较为均匀。当磷脂与胆固醇比例为2∶1,药脂比为1∶40时,Nob A脂质体的平均包封率为95.7%,平均粒径为87.6nm。Nob A以脂质体作为给药载体,其体内外溶血行为大大降低,有望成为静脉注射用脂质体。

Nobiliside A （Nob A） liposomes were prepared. Its assay method of content and encapsulation efficiency （EE） were established, and hemolytic activity with Nob A solution in vivo and in vitro were compared. Preparative method, phospholipid content, ratio of phospholipid to cholesterol and ratio of drug to lipids were optimized by single factor exploration. According to the optimized results, 3 batches of Nob A liposomes were prepared, then high performance liquid chromatography coupled with evaporative light scattering detector （HPLC-ELSD） method was used to determine the content of Nob A and minicolumn centrifugation method to determine EE, transmission electron microscope was used to detect the morphology and laser scatter analysis to evaluate particle sizes of the liposomes. The hemolytic activity was studied both in vivo and in vitro. The results indicated that HPLC-ELSD method and minicolumn centrifugation method used in this study are simple, applicable and accurate for the determination of the content and EE of Nob A liposome respectively .Nob A liposomes have a high EE with spherical shape and uniform size by using the film ultrasonication technique. When the ratio of phospholipid to cholesterol was 2∶1 and the ratio of Nob A to lipids was 1∶40, the mean EE of Nob A liposomes was 95.7% and the mean diameter was 87.6 nm. Liposomes inhibited the hemolytic activity of Nob A in vivo and in vitro sharply. As for its low hemolytic activity in vivo and in vitro, Nob A liposomes are optimistic to be used by intravenous injection.