Synthesis of bifunctional poly(ethylene oxides) and their use as ligands to nanoparticles Dissertation Submitted to the Faculty of Chemistry, University of Hamburg for the degree of Doctor of Natural Sciences by Charlotta Olsson from Malmö (Sweden) Hamburg 2009 The work described in this thesis was carried out at the Institute of Physical Chemistry, University of Hamburg in the group of Prof. Dr. Stephan Förster. 1. Referee: Prof. Dr. Stephan Förster 2. Referee: Prof. Dr. Horst Weller To my family Content Table of content 1 Introduction ................................................................................................... 1 2 Background.................................................................................................... 3 2.1 Nanoparticles..............................................................................................................3 2.1.1 Semiconductor nanoparticles...............................................................................3 2.1.2 Magnetic particles ...............................................................................................5 2.2 Anionic polymerization ..............................................................................................7 2.2.1 --functionalized poly(ethyleneoxide) ..............................................................9 2.3 Water soluble nanoparticles ......................................................................................10 2.3.1 Ligand exchange ...............................................................................................13 2.4 Self assembly............................................................................................................14 2.5 Methods for characterization.....................................................................................18 2.5.1 Size exclusion chromatography (SEC)...............................................................18 2.5.2 Mass spectrometry (MALDI-TOF)....................................................................19 2.5.3 Dynamic light scattering (DLS).........................................................................21 2.5.4 Light microscopy ..............................................................................................24 2.5.5 Fluorescence microscopy...................................................................................29 2.5.6 Confocal laser scanning microscopy (CLSM)....................................................29 2.5.7 Transmission electron microscopy (TEM).........................................................31 2.5.8 Isothermal titration microcalorimetry (ITC).......................................................32 3 Synthesis and characterization ..................................................................... 34 3.1 - and --functionalized poly(ethylene oxides) by anionic polymerization ............34 3.1.1 Diphenylmethyl- and methoxy-PEO..................................................................35 3.1.2 Acetal-PEO .......................................................................................................37 3.1.3 Acrylate-PEO....................................................................................................40 3.1.4 Carboxylic acid-PEO.........................................................................................40 3.2 Functionalization reactions on poly(ethylene oxide)..................................................43 3.2.1 Amine ...............................................................................................................43 3.2.2 Thiol .................................................................................................................52 3.2.3 Fluorescein........................................................................................................53 I Content 4 Polymer-nanoparticle-conjugates................................................................. 55 4.1 CdSe/CdS Quantum dots ..........................................................................................55 4.1.1 QD with bifunctional PEOs ...............................................................................61 4.1.2 Titration microcalorimetric measurements.........................................................63 4.2 Magnetite .................................................................................................................66 5 Vesicle encapsulation................................................................................... 68 5.1 Vesicle encapsulation of CdSe/CdS ..........................................................................68 5.2 Vesicle encapsulation of magnetic nanoparticles.......................................................74 6 Conclusion................................................................................................... 77 7 Zusammenfassung ....................................................................................... 80 8 Experimental Part ........................................................................................ 83 8.1 Anionic synthesis......................................................................................................83 8.1.1 Synthesis of initiators........................................................................................83 Methoxyethanylpotassium:............................................................................................83 3,3-diethoxypropanylpotassium.....................................................................................83 8.1.2 Synthesis of poly(ethylene oxide)......................................................................84 8.2 Functionalization of poly(ethylene oxide) .................................................................84 8.2.1 Activation of poly(ethylene oxide) ....................................................................84 8.2.2 Coupling of poly(ethylene oxide) with diethylenetriamine, PEO-DETA . ..........86 8.2.3 Coupling of poly(ethylene oxide) with triethylenetetramine, PEO-TETA. .........86 8.2.4 Coupling of poly(ethylene oxide) with tetraethylenepentamine, PEO-TEPA......86 8.2.5 Coupling of poly(ethylene oxide) with pentaethylenehexamine, PEO-PEHA.....87 8.2.6 Coupling of poly(ethylene oxide) with branched poly(ethylene imine), PEO-PEI . 87 8.2.7 Coupling with fluorescein..................................................................................87 8.2.8 Coupling of imidazole activated poly(ethylene oxide) with cysteine ..................88 8.3 Synthesis of magnetic nanoparticles..........................................................................88 8.4 Ligand exchange procedures.....................................................................................88 8.4.1 Ligand exchange on QDs...................................................................................88 8.4.2 Ligand exchange on magnetic nanoparticles......................................................89 8.5 Coupling of aldehyde QDs to hydroxy groups...........................................................89 II Content 8.6 Microcalorimetry......................................................................................................90 8.7 Characterization........................................................................................................90 8.7.1 Nuclear Magnetic Resonance Spectroscopy (NMR) ..........................................90 8.7.2 Size Exclusion Chromatography (SEC) .............................................................90 8.7.3 Matrix Assisted Laser Desorption and Ionization – Time Of Flight (MALDI- TOF) 91 8.7.4 Dynamic light scattering (DLS).........................................................................91 8.7.5 Cryo-TEM.........................................................................................................91 8.7.6 Light microscope...............................................................................................91 8.7.7 Confocal laser scanning microscope..................................................................91 8.7.8 Microcalorimeter...............................................................................................92 8.7.9 Magnetic induction............................................................................................92 9 Appendix ..................................................................................................... 93 9.1 Abbreviations and symbols.......................................................................................93 9.2 Safety precaution information of the used chemicals.................................................96 10 Literature ................................................................................................... 103 III Introduction 1 Introduction Nanoparticles have during the last couple of decades been widely investigated due to the possible applications within the field of biomedicine, optics and electronics. In this thesis special interest is given to nanoparticles within biomedicine, hence to water-soluble nanoparticles. Semiconductor nanoparticles offer the advantage of having size tunable luminescence, which make them suitable for studies of interactions between biomolecules on a cellular level, whereas magnetic nanoparticles offer the possibility for magnetic drug targeting. These nanoparticles have dimensions that are smaller or comparable to those of cells, viruses and genes and can therefore be directed close to a biological entity of interest