中空纤维离心超滤-HPLC法测定注射用两性霉素B脂质体的包封率

Determination of the entrapment efficiency of liposome amphotericin B by hollow fiber centrifugal ultrafiltration coupled with HPLC

目的建立新型的中空纤维离心超滤法分离脂质体中游离药物,并结合HPLC法用于注射用两性霉素B脂质体的包封率的表征。方法采用新型中空纤维微量离心超滤（HF-CF-UF）装置,在离心力的作用下,利用中空纤维超滤膜分离游离药物和包封药物。色谱条件：色谱柱为Diamonsil C18柱（250 mm×4.6 mm,5μm）,流动相为乙腈-20 mmol·L-1乙二胺四乙酸二钠（体积比38∶62）,流速为1.0 mL·min-1,检测波长为405 nm,柱温为30℃。结果两性霉素B质量浓度在0.670-21.4 mg·L-1内与峰面积呈良好的线性关系（r=0.999 7）,回收率均在97%以上,RSD不大于2%。结论中空纤维离心超滤装置基本保持了脂质体处方的原始的、稳定的存在环境（物理化学环境）,最大限度地减少了脂质体的泄露,结果准确且简便快速,为脂质体包封率的表征提供了一种新方法。

Objective To develop a robust, specific and sensitive method which could effectively separate the free drug and liposomal drug under equilibrium conditions by the hollow fiber centrifugal ultrafiltration （HF- CF-UF）. Methods Briefly ,0.2 mL sample was placed into a centrifugation device, and then a 15 cm piece of hollow fiber was inserted into the glass tube. The filtrate containing free drug was obtained by HF-CF-UF and analyzed by HPLC through direct injection. The accuracy and repeatability of this method have also been validated. Results A good linear relation was obtained in the range of 0. 670-21.4 mg. L -1 （ r = 0. 999 7 ）, and the average recovery was above 97 % with RSD 〈 2 %. Conclusions The apparatus maintains a substantially original liposomal formulation and provides a stability environmental （physical and chemical environment） for liposomes without yielding artificially high free drug concentrations.