CA acknowledges a Discovery grant from the Natural Sciences and Engineering Research Council of Canada. CA acknowledges a Chair in Pharmaceutics and Drug Delivery from GSK. The authors declare no competing financial interest.

<img alt="Figure 1" src="/files/ftp_upload/54422/54422fig1.jpg" /> 
Figure 1. Schematic showing the nine key steps in the preparation of the mPEG-b-PPheGE copolymer. <a href="http://cloudfront.jove.com/files/ftp_upload/54422/54422fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
1. Preparation of the Reagents under Dry Conditions
<ol>
	<li>Preparation of the reagents.
	<ol>
		<li>Weigh 15 g of mPEG-5K...

<ol>
	<li>Dickerson, T. J., Reed, N. N., Janda, K. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soluble+Polymers+as+Scaffolds+for+Recoverable+Catalysts+and+Reagents.">Soluble Polymers as Scaffolds for Recoverable Catalysts and Reagents.</a> Chemical Reviews. 102, 3325-3344 (2002).</li><li>van Heerbeek, R., Kamer, P. C. J., van Leeuwen, P. W. N. M., Reek, J. N. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dendrimers+as+Support+for+Recoverable+Catalysts+and+Reagents.">Dendrimers as Support for Recoverable Catalysts and Reagents.</a> Chemical Reviews. 102 (10), 3717-3756 (2002).</li><li>Knop, K., Hoogenboom, R., Fischer, D., Schubert, U. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Poly(ethylene+glycol)+in+Drug+Delivery:+Pros+and+Cons+as+Well+as+Potential+Alternatives.">Poly(ethylene glycol) in Drug Delivery: Pros and Cons as Well as Potential Alternatives.</a> Angewandte Chemie International Edition. 49 (36), 6288-6308 (2010).</li><li>Eetezadi, S., Ekdawi, S. N., Allen, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+challenges+facing+block+copolymer+micelles+for+cancer+therapy:+In+vivo+barriers+and+clinical+translation.">The challenges facing block copolymer micelles for cancer therapy: In vivo barriers and clinical translation.</a> Advanced Drug Delivery Reviews. 91, 7-22 (2015).</li><li>Attwood, D., Booth, C., Yeates, S. G., Chaibundit, C., Ricardo, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Block+copolymers+for+drug+solubilisation:+Relative+hydrophobicities+of+polyether+and+polyester+micelle-core-forming+blocks.">Block copolymers for drug solubilisation: Relative hydrophobicities of polyether and polyester micelle-core-forming blocks.</a> International Journal of Pharmaceutics. 345 (1-2), 35-41 (2007).</li><li>Matsumura, Y., Kataoka, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preclinical+and+clinical+studies+of+anticancer+agent-incorporating+polymer+micelles.">Preclinical and clinical studies of anticancer agent-incorporating polymer micelles.</a> Cancer Science. 100 (4), 572-579 (2009).</li><li>Chan, A. S., Chen, C. H., Huang, C. M., Hsieh, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+particle+morphology+of+pH-dependent+poly(epsilon-caprolactone)-poly(gamma-glutamic+acid)+micellar+nanoparticles+to+combat+breast+cancer+cells.">Regulation of particle morphology of pH-dependent poly(epsilon-caprolactone)-poly(gamma-glutamic acid) micellar nanoparticles to combat breast cancer cells.</a> Journal of Nanoscience and Nanotechnology. 10 (10), 6283-6297 (2010).</li><li>Diao, Y. Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doxorubicin-loaded+PEG-PCL+copolymer+micelles+enhance+cytotoxicity+and+intracellular+accumulation+of+doxorubicin+in+adriamycin-resistant+tumor+cells.">Doxorubicin-loaded PEG-PCL copolymer micelles enhance cytotoxicity and intracellular accumulation of doxorubicin in adriamycin-resistant tumor cells.</a> International Journal of Nanomedicine. 6, 1955-1962 (2011).</li><li>Mikhail, A. S., Allen, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Poly(ethylene+glycol)-b-poly(&#949;-caprolactone)+Micelles+Containing+Chemically+Conjugated+and+Physically+Entrapped+Docetaxel:+Synthesis,+Characterization,+and+the+Influence+of+the+Drug+on+Micelle+Morphology.">Poly(ethylene glycol)-b-poly(&#949;-caprolactone) Micelles Containing Chemically Conjugated and Physically Entrapped Docetaxel: Synthesis, Characterization, and the Influence of the Drug on Micelle Morphology.</a> Biomacromolecules. 11 (5), 1273-1280 (2010).</li><li>Kataoka, K., Harada, A., Nagasaki, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Block+copolymer+micelles+for+drug+delivery:+design,+characterization+and+biological+significance.">Block copolymer micelles for drug delivery: design, characterization and biological significance.</a> Advanced Drug Delivery Reviews. 47 (1), 113-131 (2001).</li><li>Nakanishi, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+the+polymer+micelle+carrier+system+for+doxorubicin.">Development of the polymer micelle carrier system for doxorubicin.</a> Journal of Controlled Release. 74 (1-3), 295-302 (2001).</li><li>Liu, J., Xiao, Y., Allen, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polymer-drug+compatibility:+A+guide+to+the+development+of+delivery+systems+for+the+anticancer+agent,+ellipticine.">Polymer-drug compatibility: A guide to the development of delivery systems for the anticancer agent, ellipticine.</a> Journal of Pharmaceutical Sciences. 93 (1), 132-143 (2004).</li><li>Flory, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+Size+Distribution+in+Ethylene+Oxide+Polymers.">Molecular Size Distribution in Ethylene Oxide Polymers.</a> Journal of the American Chemical Society. 62 (6), 1561-1565 (1940).</li><li>Kazanskii, K. S., Solovyanov, A. A., Entelis, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polymerization+of+ethylene+oxide+by+alkali+metal-naphthalene+complexes+in+tetrahydrofuran.">Polymerization of ethylene oxide by alkali metal-naphthalene complexes in tetrahydrofuran.</a> European Polymer Journal. 7 (10), 1421-1433 (1971).</li><li>Crothers, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micellization+and+Gelation+of+Diblock+Copolymers+of+Ethylene+Oxide+and+Styrene+Oxide+in+Aqueous+Solution.">Micellization and Gelation of Diblock Copolymers of Ethylene Oxide and Styrene Oxide in Aqueous Solution.</a> Langmuir. 18 (22), 8685-8691 (2002).</li><li>Taboada, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Block+Copolymers+of+Ethylene+Oxide+and+Phenyl+Glycidyl+Ether:+Micellization,+Gelation,+and+Drug+Solubilization.">Block Copolymers of Ethylene Oxide and Phenyl Glycidyl Ether: Micellization, Gelation, and Drug Solubilization.</a> Langmuir. 21 (12), 5263-5271 (2005).</li><li>Taboada, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micellization+and+Drug+Solubilization+in+Aqueous+Solutions+of+a+Diblock+Copolymer+of+Ethylene+Oxide+and+Phenyl+Glycidyl+Ether.">Micellization and Drug Solubilization in Aqueous Solutions of a Diblock Copolymer of Ethylene Oxide and Phenyl Glycidyl Ether.</a> Langmuir. 22 (18), 7465-7470 (2006).</li><li>Attwood, D., Booth, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Colloid Stability. , 61-78 (2010).</li><li>Le Devedec, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postalkylation+of+a+Common+mPEG-b-PAGE+Precursor+to+Produce+Tunable+Morphologies+of+Spheres,+Filomicelles,+Disks,+and+Polymersomes.">Postalkylation of a Common mPEG-b-PAGE Precursor to Produce Tunable Morphologies of Spheres, Filomicelles, Disks, and Polymersomes.</a> ACS Macro Letters. 5 (1), 128-133 (2016).</li><li>Chtryt, V., Ulbrich, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conjugate+of+Doxorubicin+with+a+Thermosensitive+Polymer+Drug+Carrier.">Conjugate of Doxorubicin with a Thermosensitive Polymer Drug Carrier.</a> Journal of Bioactive and Compatible Polymers. 16 (6), 427-440 (2001).</li><li>Kataoka, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doxorubicin-loaded+poly(ethylene+glycol)-poly(&#946;-benzyl-l-aspartate)+copolymer+micelles:+their+pharmaceutical+characteristics+and+biological+significance.">Doxorubicin-loaded poly(ethylene glycol)-poly(&#946;-benzyl-l-aspartate) copolymer micelles: their pharmaceutical characteristics and biological significance.</a> Journal of Controlled Release. 64 (1-3), 143-153 (2000).</li><li>Cammas, S., Matsumoto, T., Okano, T., Sakurai, Y., Kataoka, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+of+functional+polymeric+micelles+as+site-specific+drug+vehicles+based+on+poly+(&#945;-hydroxy+ethylene+oxide-co-&#946;-benzyl+l-aspartate)+block+copolymers.">Design of functional polymeric micelles as site-specific drug vehicles based on poly (&#945;-hydroxy ethylene oxide-co-&#946;-benzyl l-aspartate) block copolymers.</a> Materials Science and Engineering: C. 4 (4), 241-247 (1997).</li><li>Lv, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doxorubicin-loaded+amphiphilic+polypeptide-based+nanoparticles+as+an+efficient+drug+delivery+system+for+cancer+therapy.">Doxorubicin-loaded amphiphilic polypeptide-based nanoparticles as an efficient drug delivery system for cancer therapy.</a> Acta Biomaterialia. 9 (12), 9330-9342 (2013).</li><li>Kim, J. O., Oberoi, H. S., Desale, S., Kabanov, A. V., Bronich, T. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Polypeptide+nanogels+with+hydrophobic+moieties+in+the+cross-linked+ionic+cores:+synthesis,+characterization+and+implications+for+anticancer+drug+delivery.">Polypeptide nanogels with hydrophobic moieties in the cross-linked ionic cores: synthesis, characterization and implications for anticancer drug delivery.</a> Journal of Drug Targeting. 21 (10), 981-993 (2013).</li><li>Zhao, C. L., Winnik, M. A., Riess, G., Croucher, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescence+probe+techniques+used+to+study+micelle+formation+in+water-soluble+block+copolymers.">Fluorescence probe techniques used to study micelle formation in water-soluble block copolymers.</a> Langmuir. 6 (2), 514-516 (1990).</li><li>Wilhelm, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Poly(styrene-ethylene+oxide)+block+copolymer+micelle+formation+in+water:+a+fluorescence+probe+study.">Poly(styrene-ethylene oxide) block copolymer micelle formation in water: a fluorescence probe study.</a> Macromolecules. 24 (5), 1033-1040 (1991).</li><li>Cammas, S., Kataoka, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+poly[(ethylene+oxide)-co-(&#946;-benzyl-L-aspartate)]+polymeric+micelles:+block+copolymer+synthesis+and+micelles+formation.">Functional poly[(ethylene oxide)-co-(&#946;-benzyl-L-aspartate)] polymeric micelles: block copolymer synthesis and micelles formation.</a> Macromolecular Chemistry and Physics. 196 (6), 1899-1905 (1995).</li><li>Kwon, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Micelles+based+on+AB+block+copolymers+of+poly(ethylene+oxide)+and+poly(.beta.-benzyl+L-aspartate).">Micelles based on AB block copolymers of poly(ethylene oxide) and poly(.beta.-benzyl L-aspartate).</a> Langmuir. 9 (4), 945-949 (1993).</li><li>Ahmed, F., Discher, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-porating+polymersomes+of+PEG-PLA+and+PEG-PCL:+hydrolysis-triggered+controlled+release+vesicles.">Self-porating polymersomes of PEG-PLA and PEG-PCL: hydrolysis-triggered controlled release vesicles.</a> Journal of Controlled Release. 96 (1), 37-53 (2004).</li><li>Uhrig, D., Mays, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+techniques+in+high-vacuum+anionic+polymerization.">Experimental techniques in high-vacuum anionic polymerization.</a> Journal of Polymer Science Part A: Polymer Chemistry. 43 (24), 6179-6222 (2005).</li><li>Parker, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+solvation+on+the+properties+of+anions+in+dipolar+aprotic+solvents.">The effects of solvation on the properties of anions in dipolar aprotic solvents.</a> Quarterly Reviews, Chemical Society. 16 (2), 163-187 (1962).</li><li>Cram, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Fundamentals o] Carbanion Chemistry. , (1965).</li><li>Szwarc, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> ACS Symposium Series. 166, 1-15 (1981).</li><li>Cho, Y. W., Lee, J., Lee, S. C., Huh, K. M., Park, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hydrotropic+agents+for+study+of+in+vitro+paclitaxel+release+from+polymeric+micelles.">Hydrotropic agents for study of in vitro paclitaxel release from polymeric micelles.</a> Journal of Controlled Release. 97, 249-257 (2004).</li><li>Dewhurst, P. F., Lovell, M. R., Jones, J. L., Richards, R. W., Webster, J. R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Organization+of+Dispersions+of+a+Linear+Diblock+Copolymer+of+Polystyrene+and+Poly(ethylene+oxide)+at+the+Air&#8722;Water+Interface.">Organization of Dispersions of a Linear Diblock Copolymer of Polystyrene and Poly(ethylene oxide) at the Air&#8722;Water Interface.</a> Macromolecules. 31 (22), 7851-7864 (1998).</li><li>Opanasopit, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Block+Copolymer+Design+for+Camptothecin+Incorporation+into+Polymeric+Micelles+for+Passive+Tumor+Targeting.">Block Copolymer Design for Camptothecin Incorporation into Polymeric Micelles for Passive Tumor Targeting.</a> Pharmaceutical Research. 21 (11), 2001-2008 (2004).</li><li>Allen, G., Booth, C., Price, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=VI-The+physical+properties+of+poly(epoxides).">VI-The physical properties of poly(epoxides).</a> Polymer. 8, 414-418 (1967).</li><li>Jada, A., Hurtrez, G., Siffert, B., Riess, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structure+of+polystyrene-block-poly(ethylene+oxide)+diblock+copolymer+micelles+in+water.">Structure of polystyrene-block-poly(ethylene oxide) diblock copolymer micelles in water.</a> Macromolecular Chemistry and Physics. 197 (11), 3697-3710 (1996).</li><li>Attwood, D., Florence, A. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Surfactant systems : their chemistry, pharmacy, and biology. , (1983).</li><li>Rekatas, C. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+hydrophobe+chemical+structure+and+chain+length+on+the+solubilization+of+griseofulvin+in+aqueous+micellar+solutions+of+block+copoly(oxyalkylene)s.">The effect of hydrophobe chemical structure and chain length on the solubilization of griseofulvin in aqueous micellar solutions of block copoly(oxyalkylene)s.</a> Physical Chemistry Chemical Physics. 3 (21), 4769-4773 (2001).</li></ol>

Due to the good control that anionic polymerization provides over molecular weight it is one of the most applied processes in the industry for the preparation of polymers based on oxirane monomers (PEG and PPG). Optimal and stringent conditions must be used for successful polymerization to be achieved. Rigorous purification of all reagents and appropriate apparatus are essential for the living character of the synthesis. Limitations of the current setup are mostly associated with the transfer technique that relies on can...

In aqueous media, amphiphilic block copolymers assemble to form nano-sized block copolymer micelles (BCMs) that consist of a hydrophobic core surrounded by a hydrophilic shell or corona. The micelle core can serve as a reservoir for the incorporation of hydrophobic drugs; while, the hydrophilic corona provides an interface between the core and the external medium. Poly(ethylene glycol) (PEG) and its derivatives are one of the most important classes of polymers and one of the most widely used in drug formulation.1-3 BCMs have proven to be a worthy drug delivery platform with several formulations relying on this technology now in late stage clinical development.4 Most commonly, the hydrophobic block of the copolymer is comprised of polycaprolactone, poly(D,L-lactide), poly(propylene oxide) or poly(&#946;-benzyl-L-aspartate).5-9
Kataoka&#39;s group investigated spherical micelles formed from PEO-b-PBLA and poly(ethylene oxide)-b-(polyaspartic acid-conjugated doxorubicin) for delivery of doxorubicin (DOX).10,11 In their reports, they put forward that &#960;-&#960; interactions between the polymer-conjugated drug or PBLA and free DOX act to stabilize the micelle core resulting in increases in drug loading and retention. It is established that compatibility or interactions between a drug and the core-forming block are determinants of key performance related parameters.12 In addition to DOX, a number of cancer therapeutics include aromatic rings within their core structure (e.g., methotrexate, olaparib, SN-38).
As a result there is significant interest in synthesis of copolymers that include benzyl rings in their core-forming blocks. Anionic ring-opening polymerization of PEG and its derivatives enable control over molecular weight and result in materials of low polydispersity in good yield.13,14 Ethylene oxide with phenyl glycidyl ether (PheGE) or styrene oxide (SO) can be (co)polymerized to form block copolymers that form micelles for solubilization of hydrophobic drugs.15-18 The current report describes the necessary steps for living anionic polymerization of phenyl glycidyl ether monomer on mPEG-OH as macroinitiator (Figure 1). The resulting block copolymer and its aggregates are then characterized in terms of properties of relevance to use in drug delivery.

<table border="0" cellpadding="0" cellspacing="0" width="742">
	<colgroup>
		<col span="2" />
		<col />
		<col span="2" />
		<col />
		<col span="2" />
		<col />
	</colgroup>
	<tbody>
		<tr height="62">
			<td colspan="3" dir="LTR" height="62" style="height:62px;width:224px;">DMEM/HAMF12</td>
			<td colspan="3" style="width:227px;">Gibco, Life Technologies</td>
			<td colspan="2" style="width:125px;">12500</td>
			<td dir="LTR" style="width:171px;">Supplemented with 10%FBS. Warm in 37 &#176;C water bath 
			&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;&#160;</td>
		</tr>
		<tr height="20">
			<td colspan="3" height="20" style="height:20px;">Trypsin-EDTA(0.25%)</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">T4049</td>
			<td dir="LTR">Warm in 37 &#176;C water bath&#160;</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Fetal bovine serum (FBS)</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">F1051</td>
			<td dir="LTR">Canada origin</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">MDA-MB-468 cell line</td>
			<td colspan="3">ATCC</td>
			<td colspan="2">HTB-132</td>
			<td></td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">MTS tetrazolium reagent</td>
			<td colspan="3">PROMEGA</td>
			<td colspan="2">G111B</td>
			<td></td>
		</tr>
		<tr height="20">
			<td colspan="3" height="20" style="height:20px;">Phenazine ethosulfate (PES)</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">P4544</td>
			<td dir="LTR">&#62;95%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">mPEG5K (Mn 5400 g/mol)</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">81323</td>
			<td>PDI=1.02</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Dimethylsolfoxide (DMSO)</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">D4540</td>
			<td dir="LTR">&#62;99.5%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Naphthalene</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">147141</td>
			<td dir="LTR">&#62;99%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Phenyl glycidyl ether</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">A32608</td>
			<td dir="LTR">&#62;85%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Benzophenone</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">427551</td>
			<td dir="LTR">&#62;99%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Potassium</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">451096</td>
			<td dir="LTR">&#62;98%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Tetrahydrofuran</td>
			<td colspan="3">Caledon Laboratory Chemicals</td>
			<td colspan="2">8900 1</td>
			<td dir="LTR">ACS</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Hexane</td>
			<td colspan="3">Caledon Laboratory Chemicals</td>
			<td colspan="2">5500 1</td>
			<td dir="LTR">ACS</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Calcium hydride (CaH2)</td>
			<td colspan="3">ACP</td>
			<td colspan="2">C-0460</td>
			<td dir="LTR">&#62;99.5%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Diethyl Ether</td>
			<td colspan="3">Caledon Laboratory Chemicals</td>
			<td colspan="2">1/10/4800</td>
			<td dir="LTR">ACS</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Microplate reader</td>
			<td colspan="3">BioTek Instruments</td>
			<td colspan="2"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Differential scanning calorimetry (DSC)</td>
			<td colspan="3">TA Instruments Inc</td>
			<td colspan="2">DSC Q100</td>
			<td></td>
		</tr>
		<tr height="32">
			<td colspan="3" dir="LTR" height="32" style="height:32px;">Gel permeation chromatography (GPC)</td>
			<td colspan="3">Waters</td>
			<td colspan="2" style="width:125px;">2695 separation moldule / 2414 detector&#160;</td>
			<td dir="LTR" style="width:171px;">2 Columns: Agilent Plgel 5&#181;m Mixed-D</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">NMR spectroscopy</td>
			<td colspan="3">Varian Mercury 400MHz</td>
			<td colspan="2"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">Chloroform-d</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">151858</td>
			<td dir="LTR">99.96%</td>
		</tr>
		<tr height="20">
			<td colspan="3" dir="LTR" height="20" style="height:20px;">DMSO-d</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">156914</td>
			<td dir="LTR">99.96%</td>
		</tr>
		<tr height="20">
			<td colspan="3" height="20" style="height:20px;">Vaccum pump&#160;</td>
			<td colspan="3">Gardner Denver Welch Vacuum Tech, Inc.</td>
			<td colspan="2"></td>
			<td>Ultimate&#160; pressure 1.10-4 torr</td>
		</tr>
		<tr height="20">
			<td colspan="3" height="20" style="height:20px;">Drierit with indicator, 8 mesh</td>
			<td colspan="3">Sigma-Aldrich</td>
			<td colspan="2">238988</td>
			<td>Regenerated at 230&#176;C for 2 hrs</td>
		</tr>
	</tbody>
</table>

anionic polymerization of an amphiphilic copolymer for preparation of block copolymer micelles stabilized by &#960;-&#960; stacking interactions

In this study, an amphiphilic copolymer that includes a core-forming block with phenyl groups was synthesized by living anionic polymerization of phenyl glycidyl ether (PheGE) on methoxy-polyethylene glycol (mPEG-b-PPheGE). Characterization of the copolymer revealed a narrow molecular distribution (PDI &#60; 1.03) and confirmed the degree of polymerization of mPEG122-b-(PheGE)15. The critical micelle concentration of the copolymer was evaluated using an established fluorescence method with the aggregation behavior evaluated by dynamic light scattering and transmission electronic microscopy. The potential of the copolymer for use in drug delivery applications was evaluated in a preliminary manner including in vitro biocompatibility, loading and release of the hydrophobic anti-cancer drug doxorubicin (DOX). A stable micelle formulation of DOX was prepared with drug loading levels up to 14% (wt%), drug loading efficiencies &#62; 60% (w/w) and sustained release of drug over 4 days under physiologically relevant conditions (acidic and neutral pH, presence of albumin). The high drug loading level and sustained release is attributed to stabilizing &#960;-&#960; interactions between DOX and the core-forming block of the micelles.

<img alt="Figure 3" src="/files/ftp_upload/54422/54422fig3.jpg" /> 
Figure 3. Illustration of the anionic polymerization of phenyl glycidyl ether on mPEG macroinitiator to produce mPEG-b-(PheGE)15 for preparation of block copolymer micelles for loading of doxorubicin. The schematic illustrates the deprotonation of the hydroxyl group of mPEG using naphthalene potassium as a radical-anion, followed...

Watch this Scientific Journal Video about Anionic Polymerization of an Amphiphilic Copolymer for Preparation of Block Copolymer Micelles Stabilized by Ï€-Ï€ Stacking Interactions at JoVE.com

Anionic Polymerization of an Amphiphilic Copolymer for Preparation of Block Copolymer Micelles Stabilized by &amp;#960;-&amp;#960; Stacking Interactions

The key steps of living anionic polymerization of phenyl glycidyl ether (PheGE) on methoxy-polyethylene glycol (mPEG-b-PPheGE) are described. The resulting block copolymer micelles (BCMs) were loaded with doxorubicin 14% (wt%) and sustained release of drug over 4 days under physiologically relevant conditions was obtained.

Anionic Polymerization of an Amphiphilic Copolymer for Preparation of Block Copolymer Micelles Stabilized by &#960;-&#960; Stacking Interactions

In this study, an amphiphilic copolymer that includes a core-forming block with phenyl groups was synthesized by living anionic polymerization of phenyl ...

The overall goal of this work is to demonstrate the critical steps needed for synthesis of block copolymer materials by ring opening anionic polymerization, and to show the necessary flow of studies for block copolymer micelles formulation of a hydrophobic drug. This protocol demonstrates the proper execution of necessary steps and procedures for the synthesis of block copolymer by anionic polymerization. The big advantage of ring opening an anionic polymerization is the synthesis of materials in good yield, and low pollute dispersity.As well as the scalability of the process. This is an exciting time for polymeric drug delivery. With a number of drugs relying on formulation and block copolymer micelles, now in late stage pre-clinical, or clinical development.Prepare reagents and the materials for the transfer as described in the text protocol. Weigh methoxypolyethylene glycol with a molecular weight of 5, 000. Add the mPEG into a dried flask containing a stir bar, and seal the flushing adapter with a septum at the top.Connect the flask to the manifold, and perch the flask for two to three minutes with Argon. Turn the valve to the vacuum position to perch the flask. Rotate the flask manually, and dry the reaction vessel homogeneously, with a blowdryer or heat gun, until the mPEG melts.After one minute, break the vacuum by turning the valve on the manifold towards the Argon position with several quick snaps to cool down the flask. Keep the polymeric macroinitiator under vacuum for approximately two hours, and then under Argon before the reaction begins. Next, mount two high vacuum distillation apparatuses which have been dried previously in an oven under the hood.One apparatus is for the distillation of DMSO, and one apparatus is for the distillation of the monomer, phenyl glycerol ether. Connect the separate flasks to the two apparatuses, and install each on a hemispherical heating mantel, or in an oil bath. Connect cold water to the top of each apparatus, and to the manifold.Ensure that each apparatus is secure and well sealed. Engage the vacuum via the valve. Set the heating via a temperature controller, and start stirring the solutions.After two hours of circulation distillation of DMSO, close the high vacuum valve to collect approximately 20 milliliters of solution. Then, release the fraction into the flask, and repeat the operation once more, to ensure the purity of the desired fraction that is collected later. Heat the flask containing the mPEG under vacuum with the heat gun until the polymer melts, and then perch again with Argon.After two hours, close the high vacuum valve, and collect the volume of solvent. Stop heating, and break the vacuum from the manifold. Release Argon into the chamber, as described previously.Under a positive pressure of Argon, connect one side of the canula to a graduated cylinder, or directly to the flask containing the mPEG, also under Argon, and immerse the other end carefully into the freshly distilled fraction. Connect an extra bubbler to the flask, close off the Argon flow, and use only the Argon pressure from the distillation apparatus. To avoid any accidents caused by Argon pressure, open the glass stopcock for one to two seconds, and reclose to continue the flow of DMSO until the full transfer is completed.Next, under Argon, connect one side of the canula from the dried graduated cylinder, sealed by a septum, to a stock solution of 0.3 molar naphthalene potassium. Then, add an extra bubbler to the graduated cylinder, close the Argon flow to the cylinder, and under positive Argon pressure, transfer five milliliters of 0.3 molar naphthalene potassium via canulation from the stock solution, to the graduated cylinder. Insert another needle from the manifold into the cylinder, and remove the extra bubbler.Remove the canula connected to the stock solution, carefully, insert it rapidly into the reaction flask, and connect an extra bubbler. Next, as described previously, add the base drop by drop until the solution becomes dark. Following the slow disappearance of color, add another portion until the dark color appears again, and repeat until the full transfer.As described previously, transfer the desired volume of monomer to the reaction to reach a degree of polymerization of polyphenol glycol ether of about 18 to 20 units. Leave the reaction for 48 hours at 80 degrees celsius, under Argon atmosphere with constant stirring, to ensure complete polymerization. Quench the reaction by the addition of drops of one normal hydrochloric acid and methanol, as observed by a color disappearance.Extract the naphthalene from the DMSO solution with hexane. Remove approximately 70 milliliters of the DMSO by distillation under vacuum. Cool down the slurry solution, and add 15 milliliters of THF.Remove the salt from the slurry solution by centrifugation at 5, 000 times g for ten minutes. Transfer the supernatant, and add a dropwise to 500 milliliters of cold diethyl ether. Collect the precipitate by filtration, or centrifugation.After repeating the precipitation twice, dry the precipitate at room temperature under the hood, overnight, and then under vacuum at 30 degrees celsius for 24 to 48 hours, for a yield of 85%The copolymer is now ready for characterization, as described in the text protocol. Dissolve 12 milligrams of doxorubicin in one milliliter of acetonitrile. Add 10 microliters of triethylamine, and let the solution stir in the dark for two hours.Dissolve 45 milligrams of copolymer in one milliliter of THF, and stir for the same period of time. Add the copolymer solution to the doxorubicin solution, and rinse the vile containing residual copolymer with an extra volume of THF. Next, add the copolymer-drug mixture dropwise to a 20 milliliter vile containing 15 milliliters of 0.9%saline with stirring.Transfer the solution to a dialysis bag, and dialyze against 500 milliliters of 0.9%saline. Transfer the dialysate to a 15 milliliter tube and centrifuge at 5, 000 times g, for 15 minutes. Then, transfer the supernatant to an ultrafiltration system that contains a dialysis membrane.Put the stirring adapter into the ultrafiltration system, close the lid, and open to a stream of nitrogen. Concentrate the block copolymer micelles solution to a volume of four milliliters, and add six milliliters of fresh saline. Then, concentrate the block copolymer micelles solution to 4 milliliters again, rinse the chamber with 0.5 milliliters of saline, and add it to the solution.Store the resulting solution in brown viles at room temperature in the dark, prior to further use. The ring opening anionic polymerization of phenyl glycerol ether on the mPEG macroinitiator was used to prepare the amphiphilic diblock copolymer by the deep protonation of the hydroxyl group of mPEG. Using naphthalene potassium as a radical anion, followed by the polymerization of the phenyl glycerol ether monomer.The characterization of the block copolymer by gel permeation chromatography, and proton NMR spectroscopy analysis confirmed a narrow molecular weight distribution with a degree of polymerization of phenyl glycerol ether of 15 units. In acquis media, the amphiphilic block copolymers assemble to form micelles that consist of a hydrophobic core surrounded by a hydrophilic shell. Transmission electron microscopy confirmed a spherical morphology and dynamic light scattering indicated a hydrodynamic diameter of 25 nano meters.Doxorubicin was successfully encapsulated in the block copolymer micelles. Compared to the free protonated drug in solution, self-quenching of the doxorubicin, observed by fluorescence spectroscopy, confirmed the incorporation of the drug in the polymeric micelles, with a drug loading capacity of up to 14%The release profile of doxorubicin, from the block copolymer micelles, indicated good stability for this formulation at neutral pH. And sustained release of the drug over four days under physiologically relevant conditions.Once mastered, this technique can help trainees in pharmaceutical sciences to prepare the synthesis diblock copolymers for application in drug delivery. This procedure described herein, can be applied for synthesis of a wide range of copolymers with PEG as the initiator. Here emphasis has been placed on assuring good laboratory practices, that provide anhydrous conditions, and result in high quality materials.After watching this video, I should have a good understanding of how to work under anhydrous conditions and verse should be able to apply these steps for the synthesis of PEG polyester or PEG polyester materials. In pursuing any of the described research, it is essential to remember that you are working under controlled conditions, with potential hazardous organic solvents, and other agents. Furthermore, all drugs are toxic, and must be handled using necessary precautions.Such as, wearing gloves, a lab coat, close-toed shoes, as well as wearing a mask when you want to weigh the drugs.

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13.8K vistas.  University of Toronto. The overall goal of this work is to demonstrate the critical steps needed for synthesis of block copolymer materials by ring opening anionic polymerization, and to show the necessary flow of studies for block copolymer micelles formulation of a hydrophobic drug. This protocol demonstrates the proper execution of necessary steps and procedures for the synthesis of block copolymer by anionic polymerization. The big advantage of ring opening an anionic polymerization is the synthesis of material...

Video: Anionic Polymerization of an Amphiphilic Copolymer for Preparation of Block Copolymer Micelles Stabilized by &#960;-&#960; Stacking Interactions