摘要
建立siRNA含量测定方法,用于阳离子脂质体中siRNA的含量和包封率测定。利用荧光染料SYBR Gold和Ribogreen与siRNA结合后能发出强烈荧光的原理,分别采用电泳法和荧光分光光度法测定脂质体中的siRNA含量,计算阳离子脂质体的包封率。SYBR Gold电泳法的检测线性范围为0.2～2.0μmol·L-1(R=0.9930),高、中、低3个浓度的回收率分别为96.35%、96.92%和100.74%(n=3);日内日间精密度的RSD均小于5%(n=5)。Ribogreen荧光分光光度法的检测线性范围为10～50nmol·L-1(R=0.9971),高、中、低3个浓度的回收率分别为98.22%、99.88%和99.64%(n=3);日内日间精密度的RSD均小于5%(n=5)。利用这两种方法所测得3批阳离子脂质体中siRNA平均含量分别为98.52%和97.85%,平均包封率分别为99.20%和96.45%,经方差分析,两种方法无显著性差异。两种方法准确可靠、稳定性高和方便实用,均可用于阳离子脂质体中siRNA的含量和包封率测定。
To develop different methods for determining siRNA content and the entrapment efficiency of siRNA loaded liposomes, SYBR Gold electrophoresis method and Ribogreen fluorospectrophotometry method were used respectively. SYBR Gold electrophoresis method has a good linear relation in a range at 0.2-2.0 μmol·L^-1 （R = 0.993 0）, and the recovery at the high, middle and low concentrations were 96.35%, 96.92%, and 100.74%, respectively （n = 3）. The intra-day and inter-day RSD were far below 5% （n = 5）. Ribogreen fluorospectrophotometry method has a good linear relation in a range at 10-50 nmol·L^-1 （R = 0.997 1）, and the recovery at the high, middle and low concentrations were 98.22%, 99.88% and 99.64%, respectively （n = 3）. The intra-day and inter-day RSD were far below 5% （n = 5）. The content and the entrapment efficiency of three batches of siRNA cationic liposomes were 98.52%, 97.85% and 99.20%, 96.45%, respectively, with these two methods. And there is no significant difference by ANOVA. Both of the two methods are accurate, sensitive, convenient method for determination of the siRNA content and the entrapment efficiency of siRNA loaded cationic liposomes.
出处
《药学学报》
CAS
CSCD
北大核心
2009年第4期430-435,共6页
Acta Pharmaceutica Sinica
基金
国家科技支撑计划(2008BA155B03)
