Toward Synthetic Intrinsically Disordered Polypeptides (IDPs): Controlled Incorporation of Glycine in the Ring-Opening Polymerization of N-Carboxyanhydrides.

Intrinsically disordered proteins (IDPs) do not have a well-defined folded structure but instead behave as extended polymer chains in solution. Many IDPs are rich in glycine residues, which create steric barriers to secondary structuring and protein folding. Inspired by this feature, we have studied how the introduction of glycine residues influences the secondary structure of a model polypeptide, poly(l-glutamic acid), a helical polymer. For this purpose, we carried out ring-opening copolymerization with γ-benzyl-l-glutamate and glycine N-carboxyanhydride (NCA) monomers. We aimed to control the glycine distribution within PBLG by adjusting the reactivity ratios of the two NCAs using different reaction conditions (temperature, solvent). The relationship between those conditions, the monomer distributions, and the secondary structure enabled the design of intrinsically disordered polypeptides when a highly gradient microstructure was achieved in DMSO.

Polypeptide- and Protein-Based Conjugate Nanoparticles via Aqueous Ring-Opening Polymerization-Induced Self-Assembly (ROPISA).

Protein-polymer conjugates and polymeric nanomaterials hold great promise in many applications including biomaterials, medicine, or nanoelectronics. In this work, the first polymerization-induced self-assembly (PISA) approach performed in aqueous medium enabling protein-polymer conjugates and nanoparticles entirely composed of amino acids is presented by using ring-opening polymerization (ROP). It is indeed shown that aqueous ring-opening polymerization-induced self-assembly (ROPISA) can be used with protein or peptidic macroinitiators without prior chemical modification and afford the simple preparation of nanomaterials with protein-like property, for example, to implement biomimetic thermoresponsivity in drug delivery.

Oxygen-Enhanced Atom Transfer Radical Polymerization through the Formation of a Copper Superoxido Complex.

In controlled radical polymerization, oxygen is typically regarded as an undesirable component resulting in terminated polymer chains, deactivated catalysts, and subsequent cessation of the polymerization. Here, we report an unusual atom transfer radical polymerization whereby oxygen favors the polymerization by triggering the in situ transformation of CuBr/L to reactive superoxido species at room temperature. Through a superoxido ARGET-ATRP mechanism, an order of magnitude faster polymerization rate and a rapid and complete initiator consumption can be achieved as opposed to when unoxidized CuBr/L was instead employed. Very high end-group fidelity has been demonstrated by mass-spectrometry and one-pot synthesis of block and multiblock copolymers while pushing the reactions to reach near-quantitative conversions in all steps. A high molecular weight polymer could also be targeted (DPn = 6400) without compromising the control over the molar mass distributions (D < 1.20), even at an extremely low copper concentration (4.5 ppm). The versatility of the technique was demonstrated by the polymerization of various monomers in a controlled fashion. Notably, the efficiency of our methodology is unaffected by the purity of the starting CuBr, and even a brown highly-oxidized 15-year-old CuBr reagent enabled a rapid and controlled polymerization with a final dispersity of 1.07, thus not only reducing associated costs but also omitting the need for rigorous catalyst purification prior to polymerization.

Memory Effect in Thermoresponsive Proline-based Polymers.

We report that synthetic polymers consisting of L-proline monomer units exhibit temperature-driven aggregation in water with unprecedented hysteresis. This protein-like behavior is robust and governed by the chirality of the proline units. It paves the way to new processes, driven by either temperature or ionic strength changes, such as a simple "with memory" thermometer.

Effect of Hydrophilic Monomer Distribution on Self-Assembly of a pH-Responsive Copolymer: Spheres, Worms and Vesicles from a Single Copolymer Composition.

A series of copolymers containing 50 mol % acrylic acid (AA) and 50 mol % butyl acrylate (BA) but with differing composition profiles ranging from an AA-BA diblock copolymer to a linear gradient poly(AA-grad-BA) copolymer were synthesized and their pH-responsive self-assembly behavior was investigated. While assemblies of the AA-BA diblock copolymer were kinetically frozen, the gradient-like compositions underwent reversible changes in size and morphology in response to changes in pH. In particular, a diblock copolymer consisting of two random copolymer segments of equal length (16 mol % and 84 mol % AA content, respectively) formed spherical micelles at pH >5, a mix of spherical and wormlike micelles at pH 5 and vesicles at pH 4. These assemblies were characterized by dynamic light scattering, cryo-transmission electron microscopy and small angle neutron scattering.

Bulk Organocatalytic Synthetic Access to Statistical Copolyesters from l-Lactide and ε-Caprolactone Using Benzoic Acid.

The development of synthetic strategies to produce statistical copolymers based on l-lactide (l-LA) and ε-caprolactone (CL), denoted as P(LA- stat-CL), remains highly challenging in polymer chemistry. This is due to the differing reactivity of the two monomers during their ring-opening copolymerization (ROcP). Yet, P(LA- stat-CL) materials are highly sought after as they combine the properties of both polylactide (PLA) and poly(ε-caprolactone) (PCL). Here, benzoic acid (BA), a naturally occurring, cheap, readily recyclable, and thermally stable weak acid, is shown to trigger the organocatalyzed ring-opening copolymerization (OROcP) of l-LA and CL under solvent-free conditions at 155 °C, in presence of various alcohols as initiators, with good control over molar masses and dispersities (1.11 < D < 1.35) of the resulting copolyesters. Various compositions can be achieved, and the formation of statistical compounds is shown through characterization by 1H, 13C, and diffusion ordered spectroscopy NMR spectroscopies and by differential scanning calorimetry, as well as through the determination of reactivity ratios ( rLA = 0.86, rCL = 0.86), using the visualization of the sum of squared residuals space method. Furthermore, this BA-OROcP process can be exploited to access metal-free PLA- b-P(LA- stat-CL)- b-PLA triblock copolymers, using a diol as an initiator. Finally, residual traces of BA remaining in P(LA- stat-CL) copolymers (<0.125 mol %) do not show any cytotoxicity toward hepatocyte-like HepaRG cells, demonstrating the safety of this organic catalyst.

Deconstructing Oligomer Distributions: Discrete Species and Artificial Distributions.

The separation of an oligo(methyl acrylate) distribution, obtained from reversible addition-fragmentation chain transfer (RAFT) polymerization, in a discrete (dispersity=1) oligomeric library (degree of polymerization between 1 and 22) is presented. The properties of this library in terms of diffusivity, glass transition temperature, and viscosity are determined, filling a significant knowledge gap associated with these materials. The obtained oligomer library is used to construct artificial oligomer distributions on demand. These artificial oligomer distributions are used to highlight the potential to tailor physical properties of a material, while concomitantly demonstrating the limitations associated with size-exclusion chromatography analysis of molecular weight and dispersity in particular.

Asymmetric Copolymers: Synthesis, Properties, and Applications of Gradient and Other Partially Segregated Copolymers.

Asymmetric copolymers are a class of materials with intriguing properties. They can be defined by a distribution of monomers within the polymer chain that is neither strictly segregated, as in the case of block copolymers, nor evenly distributed throughout each chain, as in the case of statistical copolymers. This definition includes gradient copolymers as well as block copolymers that contain segments of statistical copolymer. In this review, different methods to synthesize asymmetric copolymers are first discussed. The properties of asymmetric copolymers are investigated in comparison to those of block and random counterparts of similar composition. Finally, some examples of applications of asymmetric copolymers, both academic and industrial, are demonstrated. The aim of this review is to provide a perspective on the design and synthesis of asymmetric copolymers with useful applications.

Quantifying the Heterogeneity of Chemical Structures in Complex Charged Polymers through the Dispersity of Their Distributions of Electrophoretic Mobilities or of Compositions.

The complexity of synthetic and natural polymers used in industrial and medical applications is expanding; thus, it becomes increasingly important to improve and develop methods for their molecular characterization. Free-solution capillary electrophoresis is a robust technique for the separation and characterization of both natural and synthetic complex charged polymers. In the case of polyelectrolytes, free-solution capillary electrophoresis is in the "critical conditions" (CE-CC): it allows their separation by factors other than molar mass for molar masses typically higher than 20000 g/mol. This method is thus complementary to size-exclusion chromatography (SEC). SEC is widely used to determine molar mass distributions and their dispersities. Utilizing CE-CC, an analogous calculation of dispersity based on the distributions of electrophoretic mobilities was derived and the heterogeneity of composition or branching in different polysaccharides or synthetic polymers was obtained in a number of experimental cases. Calculations are based on a ratio of moments and could therefore be compared to simulations of polymerization processes, in analogy to the work performed on molar mass distributions. Among four possible types of dispersity, the most precise values were obtained with the calculation analogous with the dispersity of molar mass distribution Mw/Mn. In addition, the dispersity value allows conclusions based on a single value: the closer the dispersity is to 1, the more homogeneous the polymer is in terms of composition or branching. This approach allows the analysis of dispersity of important molecular attributes of polymers other than molar mass and aims at improving the overall molecular characterization of both synthetic and natural polymers. The dispersity can also be monitored online while performing a chemical reaction within the CE instrument.

Nitroxide-mediated radical ring-opening copolymerization: chain-end investigation and block copolymer synthesis.

Well-defined, degradable copolymers are successfully prepared by nitroxide-mediated radical ring opening polymerization (NMrROP) of oligo(ethylene glycol) methyl ether methacrylate (OEGMA) or methyl methacrylate (MMA), a small amount of acrylonitrile (AN) and cyclic ketene acetals (CKAs) of different structures. Phosphorous nuclear magnetic resonance allows in-depth chain-end characterization and gives crucial insights into the nature of the copoly-mer terminal sequences and the living chain fractions. By using a small library of P(OEGMA-co-AN-co-CKA) and P(MMA-co-AN-co-CKA) as macroinitiators, chain extensions with styrene are performed to furnish (amphiphilic) block copolymers comprising a degradable segment.

SI-ATRP grafting of polymers from polydopamine-modified cellulose nanocrystals.

This work reports on the possibility of using polydopamine (PDA) as a tool to immobilize bromoisobutyryl moieties at the surface of cellulose nanocrystals (CNCs) and initiate Surface Intitiated Atom Transfer Radical Polymerization (SI-ATRP) reactions from these sites. Two different strategies based on i) the stepwise modification of the CNCs with dopamine (DA) and α-bromoisobutyryl bromide (BiBB) (Protocol 1) and ii) the one-step treatment of the CNCs with a mixture of DA and BiBB-modified DA (Protocol 2), were compared. Only the CNC particles treated according to Protocol 1 guaranteed efficient anchoring of the SI-ATRP initiating sites in our experimental conditions (with limited impact on the CNCs crystalline structure), the coated layer being leached out by certain solvents in the case of Protocol 2. The brominated particles displaying the best performances were subsequently tested as potential ATRP macroinitiators, using methyl methacrylate (MMA) and styrene (St) as model monomers. Polymer-grafted particles were successfully obtained, with a grafting density twice as high for Sty as for MMA, demonstrating the validity of this strategy.

Thermal Solution Depolymerization of RAFT Telechelic Polymers.

Thermal solution depolymerization is a promising low-temperature chemical recycling strategy enabling high monomer recovery from polymers made by controlled radical polymerization. However, current methodologies predominantly focus on the depolymerization of monofunctional polymers, limiting the material scope and depolymerization pathways. Herein, we report the depolymerization of telechelic polymers synthesized by RAFT polymerization. Notably, we observed a significant decrease in the molecular weight (Mn) of the polymers during monomer recovery, which contrasts the minimal Mn shift observed during the depolymerization of monofunctional polymers. Introducing Z groups at the center or both ends of the polymer resulted in distinct kinetic profiles, indicating partial depolymerization of the bifunctional polymers, as supported by mathematical modeling. Remarkably, telechelic polymers featuring R-terminal groups showed up to 68% improvement in overall depolymerization conversion compared to their monofunctional analogues, highlighting the potential of these materials in chemical recycling and the circular economy.

Morphology and thermal transitions of self-assembled NIPAM-DMA copolymers in aqueous media depend on copolymer composition profile.

HYPOTHESIS: There is a lack of understanding of the interplay between the copolymer composition profile and thermal transition observed in aqueous solutions of N-isopropyl acrylamide (NIPAM) copolymers, as well as the correlation between this transition and the formation and structure of copolymer self-assemblies. EXPERIMENTS: For this purpose, we investigated the response of five copolymers with the same molar mass and chemical composition, but with different composition profile in aqueous solution against temperature. Using complementary analytical techniques, we probed structural properties at different length scales, from the molecular scale with Nuclear Magnetic Resonance (NMR) to the colloidal scale with Dynamic Light Scattering (DLS) and Small Angle Neutron Scattering (SANS). FINDINGS: NMR and SANS investigations strengthen each other and allow a clear picture of the change of copolymer solubility and related copolymer self-assembly as a function of temperature. At the molecular scale, dehydrating NIPAM units drag N,N-dimethyl acrylamide (DMA) moieties with them in a gradual collapse of the copolymer chain; this induces a morphological transition of the self-assemblies from star-like nanostructures to crew-cut micelles. Interestingly, the transition spans a temperature range which depends on the monomer distribution profile in the copolymer chain, with the asymmetric triblock copolymer specimen revealing the broadest one. We show that the broad morphological transitions associated with gradient copolymers can be mimicked and even surpassed by the use of stepwise gradient (asymmetric) copolymers, which can be more easily and reproducibly synthesized than linear gradient copolymers.

Polymer prodrug nanoparticles based on naturally occurring isoprenoid for anticancer therapy.

The synthesis of a novel class of polymer prodrug nanoparticles with anticancer activity is reported by using squalene, a naturally occurring isoprenoid, as a building block by the reversible addition-fragmentation (RAFT) technique. The RAFT agent was functionalized by gemcitabine (Gem) as anticancer drug, and the polymerization of squalenyl-methacrylate (SqMA) led to well-defined macromolecular prodrugs comprising one Gem at the extremity of each polymer chain. The amphiphilic nature of the resulting Gem-PSqMA conjugates allowed them to self-assemble into long-term stable and narrowly dispersed nanoparticles with significant anticancer activity in vitro on various cancer cell lines. To confer stealth properties on these nanoparticles, their PEGylation was successfully performed, as confirmed by X-ray photoelectron spectroscopy (XPS) and complement activation assay. It was also shown that the PEGylated nanoparticles could be internalized in cancer cells to a greater extent than their non-PEGylated counterparts.

Degradable and comb-like PEG-based copolymers by nitroxide-mediated radical ring-opening polymerization.

Three cyclic ketene acetals, 2-methylene-1,3-dioxepane (MDO), 5,6-benzo-2-methylene-1,3-dioxepane (BMDO), and 2-methylene-4-phenyl-1,3-dioxolane (MPDL), have been copolymerized with oligo(ethylene glycol) methyl ether methacrylate and a small amount of acrylonitrile (or styrene) at 90 °C by nitroxidemediated radical ring-opening polymerization, as a convenient way to prepare degradable PEG-based copolymers for biomedical applications. MPDL was the best candidate, enabling high monomer conversions to be reached and well-defined PEG-based copolymers with adjustable amount of ester groups in the main chain to be synthesized, leading to nearly complete hydrolytic degradation (5% KOH aqueous solution, ambient temperature). The noncytotoxicity of the obtained copolymers was shown on three different cell lines (i.e., fibroblasts, endothelial cells and macrophages), representing a promising approach for the design of degradable precursors for PEGylation and bioconjugation via the NMP technique.

Use of solvent effects to improve control over nitroxide-mediated polymerization of isoprene.

The use of 1,4-dioxane or pyridine as solvents for the polymerization of isoprene mediated by the acid-functional SG1-based alkoxyamines N-(2-methylpropyl)-N-(1-diethylphosphono-2,2-dimethylpropyl)-O-(2-carboxylprop-2-yl)hydroxylamine (BlocBuilder MA) and N-(2-methylpropyl)-N-(1-diethylphosphono-2,2-dimethylpropyl)-O-(2-carboxyleth-2-yl)hydroxylamine results in an increase in the rate of consumption of the initiator and narrower molecular weight distributions of the resulting polymer. In pyridine, an improved control of the polymerization was also obtained for a non-acid-functional initiator, and the overall rate of polymerization increased. These effects are likely to be the result of the disruption of intramolecular hydrogen bonding and, in the case of pyridine, stabilization of the polar SG1 free radical.

Concurrent control over sequence and dispersity in multiblock copolymers.

Controlling monomer sequence and dispersity in synthetic macromolecules is a major goal in polymer science as both parameters determine materials' properties and functions. However, synthetic approaches that can simultaneously control both sequence and dispersity remain experimentally unattainable. Here we report a simple, one pot and rapid synthesis of sequence-controlled multiblocks with on-demand control over dispersity while maintaining a high livingness, and good agreement between theoretical and experimental molecular weights and quantitative yields. Key to our approach is the regulation in the activity of the chain transfer agent during a controlled radical polymerization that enables the preparation of multiblocks with gradually ascending (Ɖ = 1.16 → 1.60), descending (Ɖ = 1.66 → 1.22), alternating low and high dispersity values (Ɖ = 1.17 → 1.61 → 1.24 → 1.70 → 1.26) or any combination thereof. We further demonstrate the potential of our methodology through the synthesis of highly ordered pentablock, octablock and decablock copolymers, which yield multiblocks with concurrent control over both sequence and dispersity.

Thermo-responsive lipophilic NIPAM-based block copolymers as stabilizers for lipid-based cubic nanoparticles.

The design of drug delivery systems (DDS) for the encapsulation of therapeutic agents and the controlled release to the target site of the disease is one of the main goals of nanomedicine. Although already explored in an extensive number of studies over the years, lipid assemblies, and particularly liposomes, are still considered the most promising and interesting candidates as DDS due to their biocompatibility and structural similarity with plasma membranes. Lately, this research area has been extended to include more complex lipid assemblies, such as cubosomes. Cubosomes are an emerging structural platform for the delivery of molecules with pharmaceutical interest, such as drugs, bioactives and contrast agents. Here we report on the application of a thermo-responsive copolymer poly(N,N-dimethylacrylamide)-block-poly(N-isopropylacrylamide) (PDMA-b-PNIPAM), as a thermoresponsive stabilizer of lipid-based nanoparticles for drug-delivery. First, we assessed the affinity of PDMA-b-PNIPAM towards supported and free-standing bilayers; then, we explored the colloidal and thermoresponsive properties of cubic self-assembled DDS composed of glycerol-monooleate (GMO), where PDMA-b-PNIPAM replaces the conventional stabilizer Pluronic F127 (PEOx-PPOy-PEOx), normally used for cubosomes. We prepared dispersions of cubic lipid nanoparticles with two PDMA-b-PNIPAM block copolymers of different molar mass. The colloidal properties were then assessed and compared to those exhibited by standard lipid cubic dispersions stabilized by Pluronic F-127, combining a series of experimental techniques (Quartz Crystal Microbalance with Dissipation monitoring, Dynamic Light Scattering, Small-Angle X-rays Scattering, Cryo-Transmission Electron Microscopy). Interestingly, PDMA-b-PNIPAM stabilized cubosomes display additional benefits with respect to those stabilized by Pluronic, thanks to the combination of a "sponge " effect for the controlled release of encapsulated molecules and an increased affinity towards lipid bilayer membranes, which is a promising feature to maximize fusion with the target-cellular site.

Hybrid rigid/soft and biologic/synthetic materials: polymers grafted onto cellulose microcrystals.

Rigid nanoscale polymer rods were prepared by grafting preformed amine-terminated poly(styrene) and poly(tert-butyl acrylate) onto oxidized cellulose microcrystals. Low polydispersity polymers, grown using atom transfer radical polymerization, were characterized and purified prior to cellulose attachment. Oxidation of the cellulose microcrystal led to the formation of carboxylic acids on the surface of the microcrystals. Covalent attachment of the polymers onto the cellulose microcrystals was achieved via a carbodiimide-mediated amidation reaction. The length and diameter of the polymer-cellulose composites increased upon surface modification. Typically, polymer-cellulose composites are synthesized by a grafting-from method because it can be difficult to obtain sufficient graft density using a grafting-to preparation. However, the composites reported here comprised 60-64% grafted polymer by mass. This degree of grafting-to allowed the composite to form stable suspensions in organic solvents.

Amphiphilic Asymmetric Diblock Copolymer with pH-Responsive Fluorescent Properties.

Stimuli-responsive polymers with changeable fluorescent properties have numerous applications in sensing, bioimaging, and detection. Here we describe the facile synthesis of a pH-responsive amphiphilic asymmetric diblock copolymer of acrylic acid and butyl acrylate that incorporates a polarity-sensitive fluorophore. The asymmetric structure enhances the stimuli-responsive behavior: as the environmental pH decreases, the fluorescent intensity of the asymmetric diblock copolymer gradually increases, whereas its symmetric block counterpart shows limited and stepwise change. Besides, this remarkable difference was demonstrated to be concentration-independent, as similar emission behavior was found for both polymers at lower concentrations. These results indicate that the fluorescence properties of the copolymer can be adjusted by rationally designing the copolymer structure. This work provides a novel and general strategy for the design and synthesis of polymeric materials with encapsulated structures showing stimuli-responsive fluorescent properties to be applied as fluorescent probes with a smoothly varying response curve rather than the simple on-off switch that is typical of block copolymer systems.