Nanosponge 可调大小和交联密度
Summary
这篇文章描述一个调优共价交联纳米粒子从线性聚酯含吊坠功能的大小和交联密度的过程。由裁缝合成参数 （聚合物的分子量、 吊坠功能纳入和交联剂等价物），所需的纳米颗粒大小和交联密度可以实现对药物释放的应用。
Abstract
我们描述一种协议包含吊坠环氧功能并将其纳入到 nanosponge 与控制尺寸的线性聚酯的合成。这种做法始于官能化的内酯是关键吊坠官能化聚合物的合成。内酯 (VL) 和烯丙基-内酯 (AVL) 进行共聚反应使用开环聚合。后聚合改性然后用于在某些或所有吊坠烯丙基组上安装环氧化合物基团。环氧-胺化学被受雇于窗体纳米粒子在稀溶液中聚合物和基于所需的 nanosponge 大小和交联密度的小分子二胺交联剂。Nanosponge 大小可以通过透射电镜 (TEM) 成像以确定尺寸和分布特点。此方法提供了高度可调谐聚酯可以创建可调谐纳米粒子，可以用于小分子药物封装路径。由于骨干的性质，这些粒子是水解和酶降解的缓释作用的疏水性小分子的种类繁多。
Introduction
精确地调整基于分子间交联的纳米粒子的大小和交联密度是非常重要的影响与导向药物释放的这些纳米技术1。设计 nanosponge 的可调性，即，准备不同的网络密度的颗粒，是依赖于前体聚合物的吊坠功能和成立为法团的亲水性交联剂的等价物。在此方法中，前体和交联剂在溶剂中的浓度是重要形式纳米粒子的离散的大小，而不是一种大容量的凝胶。利用定量核磁共振 (NMR) 作为一种表征技术允许注册挂件功能和聚合物相对分子质量的精确测定。一旦形成纳米粒子，它们可以集中和可溶性有机物中没有的纳米凝胶具有的特点。
纳米药物载体在最近的工作重点在使用聚 (乳酸-co-乙醇酸) (PLGA) 自组装纳米粒子2，，34，，56。PLGA 具有可降解的酯键，使其适合药物释放的应用和经常结合 poly(ethylene glycol) (PEG) 由于其隐身性能7。然而，由于 PLGA 颗粒形成的自组装性质，粒子不能溶解的有机物为进一步功能化。与 PLGA 纳米粒，提出的方法提供共价交联形成纳米粒子和定义的大小和形态，这是稳定中有机物和降解水溶液1中。这种方法的优点是能够进一步化学功能覆层表面的 nanosponge8，和其在有机溶剂中的稳定性可以用于与药物化合物1，9颗粒后加载。使用此方法，封装，疏水性小分子可以通过沉淀到水介质。疏水性和亲水性短交联剂的聚酯骨干给这些粒子在体温非晶结构特征。此外后载药量、, 颗粒可形成细悬浮液在水介质要随时注入的体内。它是我们在这项工作来评估这些聚酯 nanosponges 合成参数，并确定那些是非常重要的设计和控制大小和形态的目标。
Protocol
1. Synthesis and Characterization of AVL Place a magnetic stir bar inside a 2 neck 500 mL round bottom flask (Flask 1) and seal with an appropriate sized rubber septum and steel wire. Flame dry the flask to remove moisture by purging with nitrogen gas connected through an inlet needle and open outlet needle in the septum, while using a butane flame torch to gently heat the outside of the flask by moving the flame along the surface. Continue heating the entire flask by running the …
Representative Results
为了评估 nanosponge 的合成参数和其合力的大小之间的关系，每个聚合物前驱体的浓度及吊坠功能是重要的。在图 1中，nanosponges successfulsynthetic 计划进行回流条件下结合这两种前体聚合物和二元胺交联剂在 DCM 12 小时后。环氧化合物在溶液中的浓度，对形成离散粒子至关重要。一旦合成了 nanosponges，透射电镜成像用来确定粒子一套精确的尺寸。在<strong…
Discussion
获得重现性好的 nanosponge 大小对药物释放的应用至关重要。在聚合和 nanosponge 合成的多个参数影响产生粒子的大小和交联密度。在我们的分析，确定了三个重要参数： 聚合物的分子量、 环氧吊坠功能和交联剂等价物。为了生产出一系列的分子量和 nanosponge 合成的环氧功能，必须改变共聚物组成及 VL-co-AVL。在环氧化反应的共聚物的烯丙基官能团浓度可用于 epoxidize allyls 所需的百分比或所有?…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
LK 是感谢来自全国科学基金会研究生研究奖学金项目 (DGE-1445197) 和范德比尔特大学化学系的资金。LK 和 EH 要感谢欧西里斯瞬变电磁法仪器 (NSF EPS 1004083) 的资金。
Materials
| 2,2'-(Ethylenedioxy)bis(ethylamine) | Sigma-Aldrich | 385506-100ML | |
| 3-methyl-1-butanol | Sigma-Aldrich | 309435-100ML | anhydrous, ≥99% |
| Acetone | Sigma-Aldrich | 179124-4L | |
| Allyl bromide | Sigma-Aldrich | A29585-5G | ≥99% |
| Ammonium chloride | Fisher Scientific | A661-500 | saturated solution in DI water |
| Cell culture water | Sigma-Aldrich | W3500-500ML | Filtered through 0.45 μm syringe filter |
| Dichloromethane (DCM) | Sigma-Aldrich | 270997-100ML | anhydrous, ≥99%, contains 40-150 ppm amylene as stabilizer |
| Ethyl Acetate | Fisher Scientific | E145SK-4 | |
| EZFlow 0.2 μm Syringe Filter | Foxx Life Sciences | 386-2116-OEM | Hydrophillic PTFE, 13 mm |
| EZFlow 0.45 μm Syringe Filter | Foxx Life Sciences | 386-3126-OEM | Hydrophillic PTFE, 25 mm |
| Fisherbrand Disposable Borosilicate Glass Test Tubes with Plain End | Fisher Scientific | 14-961-31 | |
| Fisherbrand Microcentrifuge Tubes | Fisher Scientific | 14-666-318 | 1.5 mL |
| Hamilton Microliter Syringe, 100 μL | Hamilton Company | 80600 | Model 710 N SYR, Cemented NDL, 22s ga, 2 in, point style 2 |
| Hexamethylphosphoramide | Sigma-Aldrich | H11602-100G | ≥99%, contains ≤1000 ppm propylene oxide as stabilizer |
| Hexanes | Fisher Scientific | H292-4 | |
| Magnesium sulfate anhydrous | Fisher Scientific | M65-500 | |
| Meta-chloroperoxybenzoic acid | Sigma-Aldrich | 273031-100G | Purified to ≥99% by buffer wash |
| Methanol (MeOH) | Sigma-Aldrich | 322415-100ML | anhydrous, ≥99% |
| N-butyllithium solution | Sigma-Aldrich | 230707-100ML | 2.5 M in hexanes |
| N,N-diisopropylethylamine | Sigma-Aldrich | 550043-500ML | ≥99% |
| Parafilm M | Sigma-Aldrich | P7793-1EA | |
| PELCO Pro Reverse (Self-Closing) Tweezers | Ted Pella, Inc. | 5375-NM | |
| Phosphotungstic acid hydrate | Alfa Aesar | 40116 | |
| Q55 Sonicator | Qsonica | Q55-110 | 55 Watts, 20 kHz |
| SiliaMetS Cysteine | Silicycle | R80530B-10g | |
| SnakeSkin Dialysis Clips | Thermo Scientific | 68011 | |
| SnakeSkin Dialysis Tubing, 10K MWCO | Thermo Scientific | 68100 | |
| Sodium bicarbonate | Fisher Scientific | 5233-500 | saturated solution in DI water |
| TEM grid | Ted Pella, Inc. | 01822-F | Ultrathin Carbon Type-A, 400 mesh, Copper, approx. grid hole size: 42µm |
| Tetrahydrofuran (THF) | Sigma-Aldrich | 401757-1L | Anhydrous, ≥99.9%, inhibitor-free |
| Tin(II) trifluoromethanesulfonate | Sigma-Aldrich | 388122-1G | |
| Vortex-Genie 2 | Scientific Industries | SI-0236 | |
| Whatman Filter Paper, Grade 1 | Fisher Scientific | 09-805H | Circles, 185 mm |
| δ-valerolactone | Sigma-Aldrich | 389579-100ML | Purified by vacuum distillation |
Riferimenti
- van der Ende, A. E., Sathiyakumar, V., Diaz, R., Hallahan, D. E., Harth, E. Linear release nanoparticle devices for advanced targeted cancer therapies with increased efficacy. Polym Chem. 1 (1), 93 (2010).
- Sharma, S., Parmar, A., Kori, S., Sandhir, R. PLGA-based nanoparticles: A new paradigm in biomedical applications. Trends Anal Chem. 80, 30-40 (2016).
- Cao, L. B., Zeng, S., Zhao, W. Highly Stable PEGylated Poly(lactic-co-glycolic acid) (PLGA) Nanoparticles for the Effective Delivery of Docetaxel in Prostate Cancers. Nanoscale Res Lett. 11 (1), 305 (2016).
- Chelopo, M. P., Kalombo, L., Wesley-Smith, J., Grobler, A., Hayeshi, R. The fabrication and characterization of a PLGA nanoparticle-Pheroid® combined drug delivery system. J Mater Sci. 52 (6), 3133-3145 (2016).
- Guan, Q., et al. Preparation, in vitro and in vivo evaluation of mPEG-PLGA nanoparticles co-loaded with syringopicroside and hydroxytyrosol. J Mater Sci Mater Med. 27 (2), 24 (2016).
- Cannava, C., et al. Nanospheres based on PLGA/amphiphilic cyclodextrin assemblies as potential enhancers of Methylene Blue neuroprotective effect. RSC Adv. 6, 16720-16729 (2016).
- Locatelli, E., Franchini, M. C. Biodegradable PLGA-b-PEG polymeric nanoparticles: synthesis, properties, and nanomedical applications as drug delivery system. J Nanopart Res. 14 (12), (2012).
- van der Ende, A. E., Croce, T., Hamilton, S., Sathiyakumar, V., Harth, E. M. Tailored polyester nanoparticles: post-modification with dendritic transporter and targeting units via reductive amination and thiol-ene chemistry. Soft Matter. 5 (7), 1417 (2009).
- Lockhart, J. N., Stevens, D. M., Beezer, D. B., Kravitz, A., Harth, E. M. Dual drug delivery of tamoxifen and quercetin: Regulated metabolism for anticancer treatment with nanosponges. J Control Release. 220 (Pt. B), 751-757 (2015).
- Shriner, R. L., Hermann, C. K. F., Morrill, T. C., Curtin, D. Y., Fuson, R. C. . The Systematic Identification of Organic Compounds. , (2004).
- Derome, A. E. . Modern NMR Techniques for Chemistry Research. , (1987).
- Williams, D. B., Barry Carter, C. . Transmission Electron Microscopy: A Textbook for Materials Science. , (2009).
- van der Ende, A. E., Kravitz, E. J., Harth, E. M. Approach to Formation of Multifunctional Polyester Particles in Controlled Nanoscopic Dimensions. J Am Chem. Soc. 130 (27), 8706-8713 (2008).
- Stevens, D. M., Watson, H. A., LeBlanc, M. A., Wang, R. Y., Chou, J., Bauer, W. S., Harth, E. M. Practical polymerization of functionalized lactones and carbonates with Sn(OTf)2 in metal catalysed ring- opening polymerization methods. Polym Chem. 4, 2470-2474 (2013).
Citazione di questo articolo
Kendrick-Williams, L. L., Harth, E. Nanosponge Tunability in Size and Crosslinking Density. J. Vis. Exp. (126), e56073, doi:10.3791/56073 (2017).