THE PROTECTIVE EFFECT OF FREE AND MEMBRANE-BOUND CRYOPROTECTANTS DURING FREEZING AND FREEZE-DRYING OF LIPOSOMES
THE PROTECTIVE EFFECT OF FREE AND MEMBRANE-BOUND CRYOPROTECTANTS DURING FREEZING AND FREEZE-DRYING OF LIPOSOMES
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DOI：
10.1016/0168-3659(94)90257-7
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发表时间：
1994-05-01
影响因子：
10.8
通讯作者：
NUHN, P
中科院分区：
文献类型：
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作者：
AUSBORN, M;SCHREIER, H;NUHN, P
脂质体由天然（EPC）和氢化（HEPC）卵磷脂（含和不含胆固醇（CHOL））、蔗糖脂肪酸酯（SPS 7;蔗糖棕榈酸酯/硬脂酸酯）与CHOL和磷酸二鲸蜡酯（DCP）或EPC和HEPC与SPS 7的单酯、二酯和三酯制备。蔗糖或膜结合的蔗糖脂肪酸酯的冷冻保护活性进行了评估。将囊泡冷冻和解冻，或冷冻干燥和重构，并监测包封的标记物5，6-羧基荧光素（CF）的保留。CF保留率随冷冻温度的降低而降低，而蔗糖浓度的增加在冷冻和解冻过程中提供了更好的冷冻保护。SPS 7囊泡被0.6 M蔗糖完全保护，而EPC和HEPC与SPS 7的等摩尔混合物需要1 M蔗糖才能完全保护。在最高蔗糖浓度下，EPC/CHOL脂质体最多保留85%标记物，HEPC/CHOL脂质体保留95%标记物。不含蔗糖或与SPS单酯或二酯混合的冻干脂质体保留<10%CF。EPC和HEPC脂质体在0.4 M蔗糖存在下的冻干导致原始包封的标记物的75%保留。差示扫描量热法显示，在蔗糖和SPS单酯的存在下，冻干HEPC脂质体的转变温度（T（c））显著降低。红外光谱表明，蔗糖和SPS单酯形成强氢键与磷酸盐头基团，支持水置换或“假水合”假说。
Liposomes were prepared from natural (EPC) and hydrogenated (HEPC) egg phosphatidylcholine, with and without cholesterol (CHOL), from sucrose fatty acid ester (SPS7; sucrose-palmitate/stearate) with CHOL and dicetylphosphate (DCP) or from EPC and HEPC with the mono-, di- and tri-ester of SPS7. The cryoprotective activity of sucrose or membrane-bound sucrose fatty esters was assessed. Vesicles were frozen and thawed, or freeze-dried and reconstituted, and retention of the encapsulated marker 5,6-carboxyfluorescein (CF) was monitored. CF retention decreased with decreasing freezing temperature, while increasing concentrations of sucrose provided increasing cryoprotection during freezing and thawing. SPS7 vesicles were fully protected by 0.6 M sucrose, whereas equimolar mixtures of EPC and HEPC with SPS7 required 1 M sucrose for complete protection. EPC/CHOL liposomes retained maximally 85% and HEPC/CHOL liposomes 95% marker at the highest sucrose concentration. Lyophilized liposomes without sucrose or in mixture with the SPS mono- or diester retained < 10% CF. Lyophilization of EPC and HEPC liposomes in the presence of 0.4 M sucrose resulted in 75% retention of originally encapsulated marker. Differential scanning calorimetry showed a significant reduction of the transition temperature (T(c)) of lyophilized HEPC liposomes in the presence of sucrose and the SPS monoester. Infrared spectroscopy indicated sucrose and the SPS monoester forming strong hydrogen bonds with phosphate head groups which supports the water replacement or 'pseudohydration' hypothesis.