油溶性Fe_3O_4纳米颗粒—脂质体的结构、性质及功能研究
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摘要
由脂质体(liposomes)和磁性纳米颗粒(magnetic nanoparticles, MNPs)组合而成的磁性脂质体(magnetoliposomes, MLs)被认为是生物医学领域最具应用前景的药物载体之一在以往的研究中,包载亲水性磁性纳米颗粒的脂质体是研究的主要对象,而关于磷脂膜中包载油溶性磁性纳米颗粒脂质体的研究则鲜见报导。本文的工作则是围绕磷脂膜中包载油溶性磁性纳米颗粒的脂质体展开,主要内容包括：
1本文运用微乳法合成Fe3O4磁性纳米颗粒,并通过一系列操作使其具有疏水性表面。这种油溶性磁性纳米颗粒直径为6nm,能长期稳定分散于氯仿中,且具有良好的超顺磁性。在此基础上,采用优化的成膜-超临界二氧化碳法制得磷脂膜中包夹油溶性磁性纳米颗粒的脂质体。通过对脂质体形貌,磷脂膜表面性质及微观流动性的研究,可以确定油溶性磁性纳米颗粒已成功包载于磷脂双分子层的疏水区域中。
2本文综合运用原子力显微镜,静态/瞬态荧光探针技术及计算机模拟方法对油溶性磁性纳米颗粒与磷脂双分子层之间的相互作用进行研究。实验观察到当油溶性磁性纳米颗粒的尺寸比磷脂膜厚度稍大时,其包载会导致磷脂膜出现扭曲现象。疏水性磁性纳米颗粒扰乱了磷脂膜中原本紧密的分子排列。其不仅提高磷脂膜的微观流动性,还扩大了膜中的非极性区域和自由体积。此外,本文还就磁性纳米颗粒浓度对磷脂膜性质的影响开展了一系列研究。结果表明,磷脂双分子层对油溶性磁性纳米颗粒的包载能力有限,当纳米颗粒与磷脂质量比为0.002时,膜中纳米颗粒己达饱和。通过计算机模拟可以发现油溶性磁性纳米颗粒的包载使磷脂膜面积发生收缩,且纳米颗粒的浓度会对收缩程度产生影响。
3本文还重点研究了该磁性脂质体的粒度分布,包封率,磁场响应性和释放行为等性质,旨在揭示其潜在应用价值。由于磁性纳米颗粒对磷脂膜微结构的扰乱作用,脂质体的相变温度明显降低。此外,疏水性磁性纳米颗粒的包载使磁性脂质体的包封率增加,这是因为更多比例的磷脂自发形成了包封率较高的大单层囊泡。本文制备的磁性脂质体矫顽力和剩磁都为零,具有超顺磁性。因此,它在磁场中表现出良好的可控磁响应性。在具备靶向性的同时,该磁性脂质体可通过两种机制促发释放,分别是温度和交变电磁场(alternating current electromagnetic field, AMF)。磁性脂质体具有温敏特性,当环境温度高于其相变温度时,脂质体发生释放,且磁性脂质体的最终释放率基本能达到90%左右。外加交变电磁场也能促发磁性脂质体的释放,脂质体释放的开始/结束可通过施加/撤销磁场往复进行。结果表明,磁场促使的脂质体释放并不破坏脂质体结构,其为一个可逆变化。在不引起本体溶液大幅升温的情况下,磁致热现象导致磁性纳米颗粒周围的磷脂膜升温至相变温度,磷脂膜转变为结构较为松散的液晶态,这有利于内含物的向外扩散。此外,交变磁场中纳米颗粒的扰动作用会导致磷脂膜流动性增加,渗透性提高,也有助于内含物的释放。
本文研究成果进一步丰富了国内外磁性脂质体的研究内容,在磁性脂质体开发方面作出了有益的探索。
Magnetoliposomes(MLs), consisting of liposomes and magnetic nanoparticles (MNPs), have been tailored as very promising delivery vehicles in biotechnology and biomedicine applications. Liposomes with hydrophilic MNPs in their inner water are the major targets of numerous studies, while liposomes with hydrophobic MNPs in membrane were scarcely reported. In this paper, researches were carried out on magnetoliposomes with the latter structure. The main contents are as follows:
1Hydrophobic MNPs coated with AOT was synthesized by microemulsion method, which was followed by rotary evaporation treatment in order to coat the synthesized MNPs with hydrophobic surface. The superparamagnetic MNPs are spherical in shape with the diameter of about6nm, and it can be dispersed stably in chloroform. Liposomes with hydrophobic MNPs in membrane were prepared by modified supercritical carbon dioxide method. The hydrophobic MNPs were successfully embedded in the lipid bilayer, which was proved by the distorted lipid bilayer and the changed membrane fluidity.
2The interaction between hydrophobic MNPs and lipid bilayer was investigated applying atomic force microscope, steady-state/time-resolved fluorescence probe method and computer simulation method. The microstructure research indicated that the hydrophobic MNPs in the lipid bilayer not only improved membrane fluidity but also enlarged the nonpolar domain and the free volume in membrane. Moreover, systematic researches were carried out to investigate the effects of hydrophobic MNPs concentration on the morphology and microstructure of liposomes. The results show that the lipid bilayer was saturated with the hydrophobic MNPs when the mass ratio of MNPs to lipid reached0.002. The computer simulation results showed that the membrane area shrank after the MNPs embedment, because the diameter of MNPs was larger than the thickness of lipid bilayer.
3In addition, the size distribution, encapsulation efficiency, release properties and magnetic response were also studied, which shed light on the potential applications of the magnetoliposomes. The phase transition temperature(Tm) of lipid bilayer was decreased due to the disordered membrane microstructure. The hydrophobic MNPs improved the encapsulation efficiency of MLs, because the portion of large unilamelar vesicle with higher encapsulation efficiency was enlarged. The hydrophobic MNPs exhibited superparamagnetic behavior with zero coercivity and zero remanence. As a result, they showed high and controllable magnetic responses. Cargo can be released from MLs by two triggered agents, temperature and alternating current electromagnetic field (AMF). The content release from liposomes could be triggered when the temperature was higher than Tm. The cargo could be repetitively released from liposomes controlled by switching on and off the AMF. The results indicated that the release from the liposomes is due to the magnetocaloric effect resulting in the liposome phase transition and the magnetic-impelled motions leading to the improved bilayer permeability, rather than the destruction of the liposome structure.
The results of this study enriches the research of magnetoliposomes at home and abroad, and they make a useful exploration in the development of magnetoliposomes.
1本文运用微乳法合成Fe3O4磁性纳米颗粒,并通过一系列操作使其具有疏水性表面。这种油溶性磁性纳米颗粒直径为6nm,能长期稳定分散于氯仿中,且具有良好的超顺磁性。在此基础上,采用优化的成膜-超临界二氧化碳法制得磷脂膜中包夹油溶性磁性纳米颗粒的脂质体。通过对脂质体形貌,磷脂膜表面性质及微观流动性的研究,可以确定油溶性磁性纳米颗粒已成功包载于磷脂双分子层的疏水区域中。
2本文综合运用原子力显微镜,静态/瞬态荧光探针技术及计算机模拟方法对油溶性磁性纳米颗粒与磷脂双分子层之间的相互作用进行研究。实验观察到当油溶性磁性纳米颗粒的尺寸比磷脂膜厚度稍大时,其包载会导致磷脂膜出现扭曲现象。疏水性磁性纳米颗粒扰乱了磷脂膜中原本紧密的分子排列。其不仅提高磷脂膜的微观流动性,还扩大了膜中的非极性区域和自由体积。此外,本文还就磁性纳米颗粒浓度对磷脂膜性质的影响开展了一系列研究。结果表明,磷脂双分子层对油溶性磁性纳米颗粒的包载能力有限,当纳米颗粒与磷脂质量比为0.002时,膜中纳米颗粒己达饱和。通过计算机模拟可以发现油溶性磁性纳米颗粒的包载使磷脂膜面积发生收缩,且纳米颗粒的浓度会对收缩程度产生影响。
3本文还重点研究了该磁性脂质体的粒度分布,包封率,磁场响应性和释放行为等性质,旨在揭示其潜在应用价值。由于磁性纳米颗粒对磷脂膜微结构的扰乱作用,脂质体的相变温度明显降低。此外,疏水性磁性纳米颗粒的包载使磁性脂质体的包封率增加,这是因为更多比例的磷脂自发形成了包封率较高的大单层囊泡。本文制备的磁性脂质体矫顽力和剩磁都为零,具有超顺磁性。因此,它在磁场中表现出良好的可控磁响应性。在具备靶向性的同时,该磁性脂质体可通过两种机制促发释放,分别是温度和交变电磁场(alternating current electromagnetic field, AMF)。磁性脂质体具有温敏特性,当环境温度高于其相变温度时,脂质体发生释放,且磁性脂质体的最终释放率基本能达到90%左右。外加交变电磁场也能促发磁性脂质体的释放,脂质体释放的开始/结束可通过施加/撤销磁场往复进行。结果表明,磁场促使的脂质体释放并不破坏脂质体结构,其为一个可逆变化。在不引起本体溶液大幅升温的情况下,磁致热现象导致磁性纳米颗粒周围的磷脂膜升温至相变温度,磷脂膜转变为结构较为松散的液晶态,这有利于内含物的向外扩散。此外,交变磁场中纳米颗粒的扰动作用会导致磷脂膜流动性增加,渗透性提高,也有助于内含物的释放。
本文研究成果进一步丰富了国内外磁性脂质体的研究内容,在磁性脂质体开发方面作出了有益的探索。
Magnetoliposomes(MLs), consisting of liposomes and magnetic nanoparticles (MNPs), have been tailored as very promising delivery vehicles in biotechnology and biomedicine applications. Liposomes with hydrophilic MNPs in their inner water are the major targets of numerous studies, while liposomes with hydrophobic MNPs in membrane were scarcely reported. In this paper, researches were carried out on magnetoliposomes with the latter structure. The main contents are as follows:
1Hydrophobic MNPs coated with AOT was synthesized by microemulsion method, which was followed by rotary evaporation treatment in order to coat the synthesized MNPs with hydrophobic surface. The superparamagnetic MNPs are spherical in shape with the diameter of about6nm, and it can be dispersed stably in chloroform. Liposomes with hydrophobic MNPs in membrane were prepared by modified supercritical carbon dioxide method. The hydrophobic MNPs were successfully embedded in the lipid bilayer, which was proved by the distorted lipid bilayer and the changed membrane fluidity.
2The interaction between hydrophobic MNPs and lipid bilayer was investigated applying atomic force microscope, steady-state/time-resolved fluorescence probe method and computer simulation method. The microstructure research indicated that the hydrophobic MNPs in the lipid bilayer not only improved membrane fluidity but also enlarged the nonpolar domain and the free volume in membrane. Moreover, systematic researches were carried out to investigate the effects of hydrophobic MNPs concentration on the morphology and microstructure of liposomes. The results show that the lipid bilayer was saturated with the hydrophobic MNPs when the mass ratio of MNPs to lipid reached0.002. The computer simulation results showed that the membrane area shrank after the MNPs embedment, because the diameter of MNPs was larger than the thickness of lipid bilayer.
3In addition, the size distribution, encapsulation efficiency, release properties and magnetic response were also studied, which shed light on the potential applications of the magnetoliposomes. The phase transition temperature(Tm) of lipid bilayer was decreased due to the disordered membrane microstructure. The hydrophobic MNPs improved the encapsulation efficiency of MLs, because the portion of large unilamelar vesicle with higher encapsulation efficiency was enlarged. The hydrophobic MNPs exhibited superparamagnetic behavior with zero coercivity and zero remanence. As a result, they showed high and controllable magnetic responses. Cargo can be released from MLs by two triggered agents, temperature and alternating current electromagnetic field (AMF). The content release from liposomes could be triggered when the temperature was higher than Tm. The cargo could be repetitively released from liposomes controlled by switching on and off the AMF. The results indicated that the release from the liposomes is due to the magnetocaloric effect resulting in the liposome phase transition and the magnetic-impelled motions leading to the improved bilayer permeability, rather than the destruction of the liposome structure.
The results of this study enriches the research of magnetoliposomes at home and abroad, and they make a useful exploration in the development of magnetoliposomes.
引文
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