The preparation of tissue-type plasminogen activator (t-PA) containing liposomes: Entrapment efficiency and ultracentrifugation damage

被引:60
作者
Heeremans, JLM [1 ]
Gerritsen, HR [1 ]
Meusen, SP [1 ]
Mijnheer, RW [1 ]
Panday, RSG [1 ]
Prevost, R [1 ]
Kluft, C [1 ]
Crommelin, DJA [1 ]
机构
[1] TNO,PG,GAUBIUS LAB,2300 AK LEIDEN,NETHERLANDS
关键词
entrapment; liposomes; t-PA; ultracentrifugation-damage;
D O I
10.3109/10611869509015959
中图分类号
R9 [药学];
学科分类号
1007 ;
摘要
In this study, a method was developed for the efficient entrapment of active tissue-type Plasminogen Activator (t-PA) into liposomes. Experimental conditions were varied to optimize t-PA entrapment: different buffer solutions were used (pH 4 and 7.5), the effect of the incubation concentrations of phospholipid (PL) and t-PA was monitored and the influence of liposome-size was examined. Furthermore, the effect of ultracentrifugation on t-PA containing Liposomes was determined in the presence and absence of Tween 80. t-PA entrapment strongly depended on experimental conditions and ranged from 30 up to 90%. Almost quantitative (90%) entrapment (entrapment percentage defined as absolute entrapment (IU t-PA/mu mol FL) divided by total incubation ratio (IU t-PA/mu mol PL), times 100%) was obtained in Hepes buffer pH 7.5, devoid of arginine, with low ionic strength. Ultracentrifugation, used for removal of non-entrapped t-PA, was shown to have a damaging effect on the liposomes (especially in the presence of 0.05% Tween 80),leading to CPA loss. However, because acceptable alternatives were not available, ultracentrifugation was used during this study. Therefore, the encapsulation-percentage values shown in this study are in fact underestimates for the true entrapment of CPA. In conclusion: almost quantitative CPA entrapment in liposomes can be achieved by selecting the proper milieu and inducing a strong interaction between t-PA and bilayer.
引用
收藏
页码:301 / 310
页数:10
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