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Química

内部自组装脂质颗粒的简易制备碳纳米管稳定

Published: February 19, 2016
doi:
10.3791/53489
, , ,
1Centre for Materials Science, School of Physical Sciences and Computing,University of Central Lancashire, 2School of Food Science & Nutrition,University of Leeds

Summary

We report on a smart application of carbon nanotubes for kinetic stabilization of lipid particles that contain self-assembled nanostructures in their cores. The preparation of lipid particles requires rather low concentrations of carbon nanotubes permitting their use in biomedical applications such as drug delivery.

Abstract

我们提出了一种简便的方法，制备由碳纳米管（CNT）稳定的纳米结构的脂质颗粒。单壁（原始）和多壁（官能化）碳纳米管被用作稳定剂，以产生皮克林类型油包水（O / W）乳剂。脂质即的Dimodan U和植烷三醇用作乳化剂，其在过量的水自组装成的双连续立方Pn3m相。高粘度的阶段被分成使用传统的表面活性剂稳定剂或为这里所做的碳纳米管的存在探头超声波发生器更小的粒子。最初，碳纳米管（粉末形式）分散在水中，然后进一步超声与熔化的脂类以形成最终乳液。在此过程中的CNT得到涂覆有脂质分子，这反过来又被假定为包围脂滴，以形成微粒乳状液是稳定数月。 CNT-中稳定的纳米结构的脂质颗粒的平均尺寸是在亚微米ř法兰，这与颗粒比较好使用常规的表面活性剂稳定化。相比于纯脂质相（散装状态）小角X射线散射数据证实原始Pn3m立方相在CNT-中稳定的脂质分散液的滞留。蓝移和在特性G和拉曼光谱观察到碳纳米管的G'带的强度的降低表征碳纳米管表面和脂质分子之间的相互作用。这些结果表明，在碳纳米管和脂质之间的相互作用是负责在水溶液中相互稳定。作为稳定化所用的碳纳米管的浓度是非常低的和脂质分子能够官能化碳纳米管，碳纳米管的毒性预计是微不足道，而他们的生物相容性大大提高。因此，本方法发现在各种生物医学应用的巨大潜力，例如用于开发混合纳米载体系统m的输送ultiple功能分子如在联合治疗中或polytherapy。

Introduction

在过去的几十年中，纳米技术已成为一个强大的工具，特别是在医学打击臭名昭著疾病如癌症1的临床前开发的领域。在这种情况下，用尺寸<1000nm的广泛探索作为各种活性生物分子如药物，蛋白质，核酸，基因和诊断显像剂1-4的递送载体的纳米级结构。这些生物分子的纳米颗粒内的任一胶囊或缀合到纳米粒子的表面和由触发器如pH或温度5,6-在作用部位被释放。尽管在尺寸非常小，这些纳米颗粒的表面积大证明是活性生物分子的靶向递送大大有利的。在粒度和生物相容性的控制是最重要的，以优化治疗效果，因此纳米颗粒7,8的适用性。9-13脂质，聚合物14,15，16,17的金属和碳纳米管18,19已被广泛用作纳米载体各种生物医学和制药应用。

此外，基于脂质的自组装纳米结构纳米载体的应用在许多其它领域，包括食品和化妆品行业20,21宽的意义。例如，它们在蛋白质结晶22，生物分子23的分离的使用，如食品稳定剂例如在甜点24，和在活性分子如营养物质，风味剂和香料25-31的交付。自组装的脂质纳米结构不仅必须释放生物活性分子在受控和靶向方式32-38的能力，但他们也能保护功能分子从化学和酶降解39,40。虽然平面流体双层是最通讯由两亲脂质分子在水存在下形成的纳米结构，其他的结构，例如六方和立方通常也观察到20,41,42。纳米结构体的类型取决于脂质“分子形状的结构，在水中的脂质组合物，以及对温度和压力43采用这样的物理化学条件。非平面脂质纳米结构的适用性特别是立方相的，是因为它们的高粘度和非均相域一致性的限制。这些问题是由分散在大量的水中的脂质纳米结构，以形成油包水（O / W）含有微米或亚微米大小的脂质颗粒乳剂克服。在这种方式下，低粘度的合适的产品能够在保持分散颗粒内的原始脂质自组装结构来制备。这些内部自组装的颗粒的形成（简称为ISAsomes 44 </sup> 例如，从立方相和六角相hexosomes）cubosomes通常需要高能量输入步骤以及加入稳定剂，例如表面活性剂或聚合物的组合。在这个方向最近的研究表明，包括二氧化硅纳米颗粒46，粘土47-49和碳纳米管50前述乳液，适当称为皮克林51或拉姆斯登-Pickering乳液52的稳定各种固体颗粒45的应用程序。

近年来，碳基纳米结构如单壁碳纳米管（单壁碳纳米管），多壁碳纳米管（多壁碳纳米管）和富勒烯已收到的极大关注作为新型生物材料53,54。主要关注的是其毒性55-58，不溶于水，59和因此他们的生物相容性56。解决这些问题的有效方法是在表面功能alization采用无毒和生物相容分子如脂类。在水存在，脂类的方式与碳纳米管相互作用，碳纳米管的疏水表面由极性水性介质屏蔽而脂质的亲水性头部基团有助于在水中60,61它们的溶解度或分散体。脂质是细胞器以及一些食品材料的组成成分，因此其装饰应理想降低CNT的体内毒性 。在碳纳米管18,19和脂质纳米结构9-13基于独立医学领域的应用正在广泛开展，但是，结合两者的性能应用尚未充分探讨。

在这项工作中，我们使用两种不同类型的脂质和三种类型的碳纳米管，其中单壁碳纳米管是在纯净的形式，而多壁碳纳米管与羟基和羧酸基团官能化。我们已经使用非常低浓度的碳纳米管制备的分散体稳定性取决于几个因素如脂质的类型，碳纳米管的种类，脂质至CNT使用的，以及对电力和持续时间采用这种超声处理参数比。此视频协议提供的动力学稳定使用各种CNT-稳定脂质纳米粒的方法的技术细节。

Protocol

注意：在此工作中使用的CNT是在相比其散装同行可具有额外的危害纳米颗粒形式。石墨吸入，天然和合成，可引起尘肺62相似，煤工尘肺。此外，已经有关于碳纳米结构的毒性和一些以前的研究的关注建议与碳纳米管63-68的吸入有关的急性和慢性毒性。因此，避免了细CNT粉吸入，并且十分小心处理。如吸入，移至空气新鲜处。如呼吸困难，用纯氧代替，并就医。碳纳米管的溶液/分散液的配方是相?…

Representative Results

下面的结果表示）的分散体的稳定性，二）脂质颗粒的尺寸分布，三）自组装以及d的类型）对于CNT的脂质包衣的证据。分散体的稳定性（ 图2）用一个500万像素的摄像头，支持自动对焦和LED闪光灯监控。 图2.得到仅在一定区域内的稳定乳液的CNT…

Discussion

脂质颗粒的稳定
三种不同的碳纳米管用于稳定脂质分散体;其中两个是多壁和用-OH和-COOH基团官能化的，一种是单壁和非官能（原始）。碳纳米管的尺寸变化如下（直径×长度）：MWCNT-COOH：9.5纳米×1.5微米;碳纳米管-OH：8-15纳米×50微米; SWCNT：1-2纳米点¯x1-3微米。粉状碳纳米管通过探针的超声波处理分散在水中。碳纳米管的上述大小可能进一步下降，由于超超声，尽管并不均衡。在…

Declarações

The authors have nothing to disclose.

Acknowledgements

我们要感谢马修·贝克J.博士，现在斯特拉斯克莱德大学，格拉斯哥与拉曼实验和尼克·冈特先生的支持，他之前这个项目的工作。

Materials

Dimodan U Danisco 15312 Store at 4°C, Non-hazardous. Irritant to eyes and skin
Phytantriol (> 95%, GC) TCI Europe N.V. P1674 Store at 4°C, Non-hazardous. Irritant to eyes and skin
Single walled Carbon Nanotubes (90%) Nanostructured & Amorphous Materials, Inc.  1246YJS Store at room temperature. Away from direct light. Irritating to eyes, skin and respiratory system
Multi-walled carboxylic acid functionalised Carbon Nanotubes (> 80% Caron basis, > 8% carboxylic acid functionalized) Sigma-Aldrich Co. LLC  755125 Store at room temperature. Away from direct light. Causes serious eye irritation. May cause respiratory irritation
Graphitized Multi-walled hydroxy functionalised Carbon Nanotubes (99.9%) Nanostructured & Amorphous Materials, Inc. (NanoAmor)  1224YJF Store at room temperature. Away from direct light. Irritating to eyes, skin and respiratory system
Pluronic F127 Sigma-Aldrich Co. LLC  P2443 BioReagent, suitable for cell culture. Not a hazardous substance or mixture. Store at room temperature.
Acetone (99.5%) Fisher Scientific  10134100 Highly flammable liquid. Causes serious eye irritation. May cause drowsiness or dizziness
Scintillation Vial VWR International Ltd 548‐0704 Soda‐lime glass vial with low background count Fitted with foil lined urea cap, 20 ml
Jars with loose, enfolding lids (375ml) VWR International Ltd 216-3308
Beaker , 1000mL Fisher Scientific  12942161 heavy duty, low form, with spout and graduations
Pasteur glass pipette (150 mm length) with latex bulb Fisher Scientific  10006021
Microcentrifuge tube conical snap cap 1.5mL Fisher Scientific  11558232
Spatula Fisher Scientific  11352204
Heating magnetic stirrer Fisher Scientific  11715704
Magnetic stirrer bars (cylindrical, opaque PTFE, 30mm x 7mm (l x diameter)) Fisher Scientific  10011792
Needle (0.9 mm x 40 mm cannula length) Terumo UK Ltd MN-2038MQ
Retort Stand Set – With stand, clamp, base, rod, rubber 3 jaw and bosshead Camlab Ltd, UK 1177157
Millipore water equipment Barnstead Nanopure, Thermoscientific, USA
Progen Genfuge 24D Digital Microcentrifuge Progen Scientific C-2400
Probe ultra-sonicator, with 13 mm  SONICS, Vibracell,  USA
5MP camera with auto-focus and LED flash Samsung Galaxy Fame Mobile camera
Raman Spectrometer Horiba Jobin-Yvon LabRAM HR800 spectrometer
Mastersizer 3000  Malvern Instruments Ltd, Malvern, United Kingdom
Small angle X-ray scattering (SAXS) SAXSpace camera (Anton Paar, Graz, Austria), X-ray generating equipment (ISO-DEBYEFLEX3003, GE Inspection Technologies GmbH), closed water circuit (Chilly 35, HYFRA, Germany). 
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Keywords: Carbon NanotubesLipid ParticlesSelf-assemblyEmulsionNanostructureSurfactantSonicationDimodan UPhytantriolF127Hydroxyl-functionalized CNTsCarboxyl-functionalized CNTs
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Patil-Sen, Y., Sadeghpour, A., Rappolt, M., Kulkarni, C. V. Facile Preparation of Internally Self-assembled Lipid Particles Stabilized by Carbon Nanotubes. J. Vis. Exp. (108), e53489, doi:10.3791/53489 (2016).
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