Joseph R. Lovett

pH-Responsive Non-Ionic Diblock Copolymers: Ionization of Carboxylic Acid End-Groups Induces an Order–Order Morphological Transition†

A carboxylic acid based reversible additionfragmentation transfer (RAFT) agent is used to prepare gels composed of worm-like diblock copolymers using two non-ionic monomers, glycerol monomethacrylate (GMA) and 2-hydroxypropyl methacrylate (HPMA). Ionization of the carboxylic acid end-group on the PGMA stabilizer block induces a worm-to-sphere transition, which in turn causes immediate degelation. This morphological transition is fully reversible as determined by TEM and rheology studies and occurs because of a subtle change in the packing parameter for the copolymer chains. A control experiment where the methyl ester derivative of the RAFT agent is used to prepare the same diblock copolymer confirms that no pH-responsive behavior occurs in this case. This end-group ionization approach is important for the design of new pH-responsive copolymer nano-objects as, unlike polyacids or polybases, only a minimal amount of added base (or acid) is required to drive the morphological transition.

Combining Biomimetic Block Copolymer Worms with an Ice‐Inhibiting Polymer for the Solvent‐Free Cryopreservation of Red Blood Cells

Abstract The first fully synthetic polymer‐based approach for red‐blood‐cell cryopreservation without the need for any (toxic) organic solvents is reported. Highly hydroxylated block copolymer worms are shown to be a suitable replacement for hydroxyethyl starch as a extracellular matrix for red blood cells. When used alone, the worms are not a particularly effective preservative. However, when combined with poly(vinyl alcohol), a known ice‐recrystallization inhibitor, a remarkable additive cryopreservative effect is observed that matches the performance of hydroxyethyl starch. Moreover, these block copolymer worms enable post‐thaw gelation by simply warming to 20 °C. This approach offers a new solution for both the storage and transport of red blood cells and also a convenient matrix for subsequent 3D cell cultures.

pH-Responsive non-ionic diblock copolymers: protonation of a morpholine end-group induces an order–order transition

A new morpholine-functionalised, trithiocarbonate-based RAFT agent, MPETTC, was synthesised with an overall yield of 80% and used to prepare a poly(glycerol monomethacrlyate) (PGMA) chain transfer agent. Subsequent chain extension with 2-hydroxypropyl methacrylate (HPMA) using a RAFT aqueous dispersion polymerisation formulation at pH 7.0–7.5 resulted in the formation of morpholine-functionalised PGMA-PHPMA diblock copolymer worms via polymerisation-induced self-assembly (PISA). These worms form soft, free-standing aqueous hydrogels at 15% w/w solids. Acidification causes protonation of the morpholine end-groups, which increases the hydrophilic character of the PGMA stabiliser block. This causes a subtle change in the copolymer packing parameter which induces a worm-to-sphere morphological transition and hence leads to in situ degelation at pH 3. This order–order transition was characterised by dynamic light scattering, transmission electron microscopy and gel rheology studies. On returning to pH 7, regelation is observed at 15% w/w solids, indicating the reversible nature of the transition. However, such diblock copolymer worm gels remain intact when acidified in the presence of electrolyte, since the terminal cationic charge arising from the protonated morpholine end-groups is screened under these conditions. Moreover, regelation is also observed in relatively acidic solution (pH < 2), because the excess acid acts as a salt under these conditions and so induces a sphere-to-worm transition.

A Robust Cross-Linking Strategy for Block Copolymer Worms Prepared via Polymerization-Induced Self-Assembly

A poly(glycerol monomethacrylate) (PGMA) chain transfer agent is chain-extended by reversible addition–fragmentation chain transfer (RAFT) statistical copolymerization of 2-hydroxypropyl methacrylate (HPMA) with glycidyl methacrylate (GlyMA) in concentrated aqueous solution via polymerization-induced self-assembly (PISA). A series of five free-standing worm gels is prepared by fixing the overall degree of polymerization of the core-forming block at 144 while varying its GlyMA content from 0 to 20 mol %. 1H NMR kinetics indicated that GlyMA is consumed much faster than HPMA, producing a GlyMA-rich sequence close to the PGMA stabilizer block. Temperature-dependent oscillatory rheological studies indicate that increasing the GlyMA content leads to progressively less thermoresponsive worm gels, with no degelation on cooling being observed for worms containing 20 mol % GlyMA. The epoxy groups in the GlyMA residues can be ring-opened using 3-aminopropyltriethoxysilane (APTES) in order to prepare core cross-linked worms via hydrolysis-condensation with the siloxane groups and/or hydroxyl groups on the HPMA residues. Perhaps surprisingly, 1H NMR analysis indicates that the epoxy–amine reaction and the intermolecular cross-linking occur on similar time scales. Cross-linking leads to stiffer worm gels that do not undergo degelation upon cooling. Dynamic light scattering studies and TEM analyses conducted on linear worms exposed to either methanol (a good solvent for both blocks) or anionic surfactant result in immediate worm dissociation. In contrast, cross-linked worms remain intact under such conditions, provided that the worm cores comprise at least 10 mol % GlyMA.

Bespoke cationic nano-objects via RAFT aqueous dispersion polymerisation

A range of cationic diblock copolymer nanoparticles are synthesised via polymerisation-induced self-assembly (PISA) using a RAFT aqueous dispersion polymerisation formulation. The cationic character of these nanoparticles can be systematically varied by utilising a binary mixture of two macro-CTAs, namely non-ionic poly(glycerol monomethacrylate) (PGMA) and cationic poly[2-(methacryloyloxy)ethyl]trimethylammonium chloride (PQDMA), with poly(2-hydroxypropyl methacrylate) (PHPMA) being selected as the hydrophobic core-forming block. Thus a series of cationic diblock copolymer nano-objects with the general formula ([1 − n] PGMAx + [n] PQDMAy) − PHPMAz were prepared at 20% w/w solids, where n is the mol fraction of the cationic block and x, y and z are the mean degrees of polymerisation of the non-ionic, cationic and hydrophobic blocks, respectively. These cationic diblock copolymer nanoparticles were analysed in terms of their chemical composition, particle size, morphology and cationic character using 1H NMR spectroscopy, dynamic light scattering (DLS), transmission electron microscopy (TEM), and aqueous electrophoresis, respectively. Systematic variation of the above PISA formulation enabled the formation of spheres, worms or vesicles that remain cationic over a wide pH range. However, increasing the cationic character favors the formation of kinetically-trapped spheres, since it leads to more effective steric stabilisation which prevents sphere–sphere fusion. Furthermore, cationic worms form a soft free-standing gel at 25 °C that undergoes reversible degelation on cooling, as indicated by variable temperature oscillatory rheology studies. Finally, the antimicrobial activity of this thermo-responsive cationic worm gel towards the well-known pathogen Staphylococcus aureus is examined via direct contact assays.

H2O2 Enables Convenient Removal of RAFT End-Groups from Block Copolymer Nano-Objects Prepared via Polymerization-Induced Self-Assembly in Water

RAFT-synthesized polymers are typically colored and malodorous due to the presence of the sulfur-based RAFT end-group(s). In principle, RAFT end-groups can be removed by treating molecularly dissolved copolymer chains with excess free radical initiators, amines, or oxidants. Herein we report a convenient method for the removal of RAFT end-groups from aqueous dispersions of diblock copolymer nano-objects using H2O2. This oxidant is relatively cheap, has minimal impact on the copolymer morphology, and produces benign side products that can be readily removed via dialysis. We investigate the efficiency of end-group removal for various diblock copolymer nano-objects prepared with either dithiobenzoate- or trithiocarbonate-based RAFT chain transfer agents. The advantage of using UV GPC rather than UV spectroscopy is demonstrated for assessing both the kinetics and extent of end-group removal.

Stimulus-responsive non-ionic diblock copolymers: protonation of a tertiary amine end-group induces vesicle-to-worm or vesicle-to-sphere transitions

A well-defined poly(glycerol monomethacrylate) (PGMA) macromolecular chain transfer agent (macro-CTA) with a mean degree of polymerisation (DP) of 43 was prepared by reversible addition–fragmentation chain transfer (RAFT) polymerisation using a morpholine-functionalised trithiocarbonate-based chain transfer agent (MPETTC). Chain extension of this macro-CTA by RAFT aqueous dispersion polymerisation of 2-hydroxypropyl methacrylate (HPMA) at pH 7.0–7.5 produced a series of four MPETTC-PGMA43-PHPMAy vesicles (where y = 190, 200, 220 or 230). Protonation of the morpholine end-group increases the hydrophilic character of the PGMA stabiliser block, which leads to a reduction in the packing parameter for the diblock copolymer chains. However, such pH-responsive behaviour critically depends on the value of y. For y = 190 or 200, lowering the solution pH to pH 3 induces a vesicle-to-worm transition at 20 °C according to dynamic light scattering, aqueous electrophoresis, transmission electron microscopy and turbidimetry studies. This order–order transition is suppressed in the presence of added electrolyte, which screens the cationic end-groups. In addition, no change in copolymer morphology was observed on lowering the solution temperature at neutral pH, regardless of the y value. The diblock copolymer nano-objects obtained at pH 3 were also cooled to 4 °C to examine their dual stimulus-responsive behaviour to both pH and temperature triggers. In all four cases, a change in morphology from either worms or vesicles to afford spheres (or spheres plus relatively short worms) was observed. Temperature-dependent oscillatory rheology experiments performed on cationic worms at pH 3 indicated a worm-to-sphere transition on cooling from 20 °C to 4 °C, which leads to reversible degelation. In summary, spheres, worms or vesicles can be obtained for MPETTC-PGMA-PHPMA diblock copolymers on first lowering the solution pH to pH 3, followed by cooling from 20 °C to 4 °C.

Synthesis of well-defined epoxy-functional spherical nanoparticles by RAFT aqueous emulsion polymerization

The environmentally-friendly synthesis of epoxy-functional spherical nanoparticles has been achieved using polymerization-induced self-assembly (PISA) in aqueous solution. Firstly, a non-ionic hydrophilic stabilizer block, poly(glycerol monomethacrylate) (PGMA), was prepared by reversible addition–fragmentation chain transfer (RAFT) solution polymerization in ethanol. This water-soluble precursor was subsequently chain-extended via RAFT aqueous emulsion polymerization of glycidyl methacrylate (GlyMA) at 50 °C and neutral pH to ensure maximum retention of the epoxy functionality. PISA leads to the formation of well-defined PGMA-PGlyMA spherical diblock copolymer nanoparticles at up to 35% w/w solids and 1H NMR spectroscopy studies indicated that virtually all of the epoxy groups survive such relatively mild conditions. DMF GPC studies confirmed that relatively low dispersities (Mw/Mn < 1.30) were obtained if the mean degree of polymerization of the core-forming PGlyMA block remained below 100. Well-defined triblock copolymer nanoparticles could also be prepared via seeded RAFT emulsion polymerization of n-butyl methacrylate, with DMF GPC analysis indicating a relatively narrow molecular weight distribution (Mw/Mn < 1.20). The epoxy groups within the nanoparticle cores were ring-opened by adding sodium azide to a 10% w/w aqueous copolymer dispersion at 50 °C, as confirmed by FT-IR spectroscopy. PGMA45-PGlyMA100 diblock copolymer nanoparticles could be conveniently converted into cationic nanogels by utilizing water-soluble diamines as crosslinkers. These nanogels were characterized by DLS and aqueous electrophoresis and remained intact when dispersed in DMF; in contrast, the corresponding linear precursor nanoparticles dissociated to form molecularly-dissolved copolymer chains under the same conditions.

Characterization of diblock copolymer order-order transitions in semidilute aqueous solution using fluorescence correlation spectroscopy

The temperature and pH-dependent diffusion of poly(glycerol monomethacrylate)-block-poly(2-hydroxypropyl methacrylate) nanoparticles prepared via polymerization-induced self-assembly in water is characterized using fluorescence correlation spectroscopy (FCS). Lowering the solution temperature or raising the solution pH induces a worm-to-sphere transition and hence an increase in diffusion coefficient by a factor of between four and eight. FCS enables morphological transitions to be monitored at relatively high copolymer concentrations (10% w/w) compared to those required for dynamic light scattering (0.1% w/w). This is important because such transitions are reversible at the former concentration, whereas they are irreversible at the latter. Furthermore, the FCS data suggest that the thermal transition takes place over a very narrow temperature range (less than 2 °C). These results demonstrate the application of FCS to characterize order-order transitions, as opposed to order-disorder transitions.

Epoxy-Functional Sterically Stabilized Diblock Copolymer Nanoparticles via RAFT Aqueous Emulsion Polymerization: Comparison of Two Synthetic Strategies

Polymerization-induced self-assembly (PISA) is a powerful and versatile technique for the synthesis of a wide range of sterically stabilized diblock copolymer nano-objects. Recently, PISA has been used to prepare epoxy-functional diblock copolymer worms and spheres directly in aqueous solution by incorporating glycidyl methacrylate (GlyMA) into the core-forming hydrophobic block. Herein, the synthesis of diblock copolymer spheres via reversible addition-fragmentation chain transfer (RAFT) aqueous emulsion polymerization of benzyl methacrylate is examined, in which the epoxy groups are exclusively located within a non-ionic poly(glycerol monomethacrylate)-based stabilizer block. Two synthetic strategies are explored: i) using an epoxy-functional RAFT chain transfer agent (CTA) to place an epoxy group at the terminus of every stabilizer block and ii) incorporation of ≈1 epoxy group per stabilizer chain via copolymerization of GlyMA with glycerol monomethacrylate (GMA). The epoxy groups conferred by the GlyMA comonomer are significantly more resistant to hydrolysis than those introduced using the epoxy-functional RAFT CTA. The epoxy-functional nanoparticles are subsequently reacted with various water-soluble thiols to modify their electrophoretic behavior. Such nanoparticles are expected to offer potential applications in the context of mucoadhesion.