Radical-Mediated Enzymatic Polymerizations.

Polymerization reactions are commonly effected by exposing monomer formulations to some initiation stimulus such as elevated temperature, light, or a chemical reactant. Increasingly, these polymerization reactions are mediated by enzymes--catalytic proteins--owing to their reaction efficiency under mild conditions as well as their environmental friendliness. The utilization of enzymes, particularly oxidases and peroxidases, for generating radicals via reduction-oxidation mechanisms is especially common for initiating radical-mediated polymerization reactions, including vinyl chain-growth polymerization, atom transfer radical polymerization, thiol-ene step-growth polymerization, and polymerization via oxidative coupling. While enzyme-mediated polymerization is useful for the production of materials intended for subsequent use, it is especially well-suited for in situ polymerizations, where the polymer is formed in the place where it will be utilized. Such polymerizations are especially useful for biomedical adhesives and for sensing applications.

Dynamic Covalent Assembly of Peptoid-Based Ladder Oligomers by Vernier Templating.

Dynamic covalent chemistry, in conjunction with template-directed assembly, enables the fabrication of extended nanostructures that are both precise and tough. Here we demonstrate the dynamic covalent assembly of peptoid-based molecular ladders with up to 12 rungs via scandium(III)-catalyzed imine metathesis by employing the principle of Vernier templating, where small precursor units with mismatched numbers of complementary functional groups are coreacted to yield larger structures with sizes determined by the respective precursor functionalities. Owing to their monomer diversity and synthetic accessibility, sequence-specific oligopeptoids bearing dynamic covalent pendant groups were employed as precursors for molecular ladder fabrication. The generated structures were characterized using matrix-assisted laser desorption/ionization mass spectrometry and gel permeation chromatography, confirming successful molecular ladder fabrication.

Effect of Cholesterol on Biomimetic Membrane Curvature and Coronavirus Fusion Peptide Encapsulation.

Biomimetic cubic phases can be used for protein encapsulation in a variety of applications such as biosensors and drug delivery. Cubic phases with a high concentration of cholesterol and phospholipids were obtained herein. It is shown that the cubic phase structure can be maintained with a higher concentration of biomimetic membrane additives than has been reported previously. Opposing effects on the curvature of the membrane were observed upon the addition of phospholipids and cholesterol. Furthermore, the coronavirus fusion peptide significantly increased the negative curvature of the biomimetic membrane with cholesterol. We show that the viral fusion peptide can undergo structural changes leading to the formation of hydrophobic α-helices that insert into the lipid bilayer. This is of high importance, as a fusion peptide that induces increased negative curvature as shown by the formation of inverse hexagonal phases allows for greater contact area between two membranes, which is required for viral fusion to occur. The cytotoxicity assay showed that the toxicity toward HeLa cells was dramatically decreased when the cholesterol or peptide level in the nanoparticles increased. This suggests that the addition of cholesterol can improve the biocompatibility of the cubic phase nanoparticles, making them safer for use in biomedical applications. As the results, this work improves the potential for the biomedical end-use applications of the nonlamellar lipid nanoparticles and shows the need of systematic formulation studies due to the complex interplay of all components.

Nonequilibrium interfacial diffusion across microdroplet interface.

Increases in complexity attainable in molecular self-assembly necessitates both advanced molecular design as well as microenvironmental control. Such control is offered by microfluidics, where precise chemical compositions and gradients can be readily established. A droplet microfluidic platform combining upstream step emulsification with downstream hydrodynamic microtraps has been designed to facilitate molecular self-assembly. The step emulsification rapidly generates uniform droplets which act as reaction chambers. The hydrodynamic microtraps hold droplets against the flow ensuring they are exposed to a continuous supply of fresh fluid for constant reagent extraction and/or delivery. Additionally, the droplet immobilization permits real-time droplet characterization and reaction monitoring. Subsequently, droplets can be released from the traps through flow reversal, allowing post-process characterization. The microfluidic system was demonstrated by the phase separation of lyotropic droplets. Ethanol/water droplets were created in a continuous ambient squalene/monoolein microflow, causing the continuous extraction of ethanol from the droplets and delivery of monoolein from the ambient microflow. Unlike conventional bulk techniques and continuous microfluidics, where finite microchannel lengths necessarily impose limits to the extent to which slow processes can proceed, this approach allows extended duration reactions whilst enabling real time process monitoring.

Microfluidic enhancement of self-assembly systems.

Dynamic, kinetically-controlled, self-assembly processes are commonly observed in nature and are capable of creating intricate, functional architectures from simple precursors. However, notably, much of the research into molecular self-assembly has been performed using conventional bulk techniques where the resultant species are dictated by thermodynamic stability to yield relatively simple assemblies. Whereas, the environmental control offered by microfluidic systems offers methods to achieve non-equilibrium reaction conditions capable of increasingly sophisticated self-assembled structures. Alterations to the immediate microenvironment during the assembly of the molecules is possible, providing the basis for kinetically-controlled assembly. This review examines the key mechanism offered by microfluidic systems and the architectures required to access them. The mechanisms include diffusion-led mixing, shear gradient alignment, spatial and temporal confinement, and structural templates in multiphase systems. The works are selected and categorised in terms of the microfluidic approaches taken rather than the chemical constructs which are formed.

Photopolymerized Thiol-Ene Systems as Shape Memory Polymers.

In this study we introduce the use of thiol-ene photopolymers as shape memory polymer systems. The thiol-ene polymer networks are compared to a commonly utilized acrylic shape memory polymer and shown to have significantly improved properties for two different thiol-ene based polymer formulations. Using thermomechanical and mechanical analysis, we demonstrate that thiol-ene based shape memory polymer systems have comparable thermomechanical properties while also exhibiting a number of advantageous properties due to the thiol-ene polymerization mechanism which results in the formation of a homogenous polymer network with low shrinkage stress and negligible oxygen inhibition. The resulting thiol-ene shape memory polymer systems are tough and flexible as compared to the acrylic counterparts. The polymers evaluated in this study were engineered to have a glass transition temperature between 30 and 40 °C, exhibited free strain recovery of greater than 96% and constrained stress recovery of 100%. The thiol-ene polymers exhibited excellent shape fixity and a rapid and distinct shape memory actuation response.

Sequence-selective dynamic covalent assembly of information-bearing oligomers.

Relatively robust dynamic covalent interactions have been employed extensively to mediate molecular self-assembly reactions; however, these assembly processes often do not converge to a thermodynamic equilibrium, instead yielding mixtures of kinetically-trapped species. Here, we report a dynamic covalent self-assembly process that mitigates kinetic trapping such that multiple unique oligomers bearing covalently coreactive pendant groups are able to undergo simultaneous, sequence-selective hybridization with their complementary strands to afford biomimetic, in-registry molecular ladders with covalent rungs. Analogous to the thermal cycling commonly employed for nucleic acid melting and annealing, this is achieved by raising and lowering the concentration of a multi-role reagent to effect quantitative dissociation and subsequently catalyze covalent bond rearrangement, affording selective assembly of the oligomeric sequences. The hybridization specificity afforded by this process further enabled information encoded in oligomers to be retrieved through selective hybridization with complementary, mass-labeled sequences.

Synthesis of Information-bearing Peptoids and their Sequence-directed Dynamic Covalent Self-assembly.

This protocol presents the use of Lewis acidic multi-role reagents to circumvent kinetic trapping observed during the self-assembly of information-encoded oligomeric strands mediated by paired dynamic covalent interactions in a manner mimicking the thermal cycling commonly employed for the self-assembly of complementary nucleic acid sequences. Primary amine monomers bearing aldehyde and amine pendant moieties are functionalized with orthogonal protecting groups for use as dynamic covalent reactant pairs. Using a modified automated peptide synthesizer, the primary amine monomers are encoded into oligo(peptoid) strands through solid-phase submonomer synthesis. Upon purification by high-performance liquid chromatography (HPLC) and characterization by electrospray ionization mass spectrometry (ESI-MS), sequence-specific oligomers are subjected to high-loading of a Lewis acidic rare-earth metal triflate which both deprotects the aldehyde moieties and affects the reactant pair equilibrium such that strands completely dissociate. Subsequently, a fraction of the Lewis acid is extracted, enabling annealing of complementary sequence-specific strands to form information-encoded molecular ladders characterized by matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS). The simple procedure outlined in this report circumvents kinetic traps commonly experienced in the field of dynamic covalent assembly and serves as a platform for the future design of robust, complex architectures.

Rapid Gel Card Agglutination Assays for Serological Analysis Following SARS-CoV-2 Infection in Humans.

High-throughput and rapid serology assays to detect the antibody response specific to severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) in human blood samples are urgently required to improve our understanding of the effects of COVID-19 across the world. Short-term applications include rapid case identification and contact tracing to limit viral spread, while population screening to determine the extent of viral infection across communities is a longer-term need. Assays developed to address these needs should match the ASSURED criteria. We have identified agglutination tests based on the commonly employed blood typing methods as a viable option. These blood typing tests are employed in hospitals worldwide, are high-throughput, fast (10-30 min), and automated in most cases. Herein, we describe the application of agglutination assays to SARS-CoV-2 serology testing by combining column agglutination testing with peptide-antibody bioconjugates, which facilitate red cell cross-linking only in the presence of plasma containing antibodies against SARS-CoV-2. This simple, rapid, and easily scalable approach has immediate application in SARS-CoV-2 serological testing and is a useful platform for assay development beyond the COVID-19 pandemic.

Volumetric Photopolymerization Confinement through Dual-Wavelength Photoinitiation and Photoinhibition.

Conventional photolithographic rapid prototyping approaches typically achieve reaction confinement in depth through patterned irradiation of a photopolymerizable resin at a wavelength where the resin strongly absorbs, such that only a very thin layer of material is solidified. Consequently, three-dimensional objects are fabricated by progressive, two-dimensional addition of material, curtailing fabrication rates and necessitating the incorporation of support structures to ensure the integrity of overhanging features. Here, we examine butyl nitrite as a UV-active photoinhibitor of blue light-induced photopolymerizations and explore its utilization to confine in depth the region polymerized in a volume of resin. By employing two perpendicular irradiation patterns at blue and near-UV wavelengths to independently effect either polymerization initiation or inhibition, respectively, we enable three-dimensional photopolymerization patterning in bulk resin, thereby complementing emergent approaches to volumetric 3D printing.

Rapid, continuous additive manufacturing by volumetric polymerization inhibition patterning.

Contemporary, layer-wise additive manufacturing approaches afford sluggish object fabrication rates and often yield parts with ridged surfaces; in contrast, continuous stereolithographic printing overcomes the layer-wise operation of conventional devices, greatly increasing achievable print speeds and generating objects with smooth surfaces. We demonstrate a novel method for rapid and continuous stereolithographic additive manufacturing by using two-color irradiation of (meth)acrylate resin formulations containing complementary photoinitiator and photoinhibitor species. In this approach, photopatterned polymerization inhibition volumes generated by irradiation at one wavelength spatially confine the region photopolymerized by a second concurrent irradiation wavelength. Moreover, the inhibition volumes created using this method enable localized control of the polymerized region thickness to effect single-exposure, topographical patterning.

Oxygen-mediated Polymerization Initiated by Oltipraz-derived Thiones.

A pyrrolopyrazine-thione derived from oltipraz, a compound that has been investigated as a chemopreventive agent, affords radicals in the presence of thiols and oxygen via a redox cycle, an attribute that suggests its suitability as an initiator for oxygen-mediated polymerization. Here, we explore the utilization of this pyrrolopyrazine-thione, generated in situ from a precursor, as an initiator for the radical-mediated thiol-ene polymerization. While the pyrrolopyrazine-thione was shown to be capable of generating radicals in the presence of atmospheric oxygen and thiol groups, the reaction extents achievable were lower than desired owing to the presence of unwanted side reactions that would quench radical production and, subsequently, suppress polymerization. Moreover, we found that complex interactions between the pyrrolopyrazine-thione, its precursor, oxygen, and thiol groups determine whether or not the quenching reaction dominates over those favorable to polymerization.

Exploring deformable particles in vascular-targeted drug delivery: Softer is only sometimes better.

The ability of vascular-targeted drug carriers (VTCs) to localize and bind to a targeted, diseased endothelium determines their overall clinical utility. Here, we investigate how particle modulus and size determine adhesion of VTCs to the vascular wall under physiological blood flow conditions. In general, deformable microparticles (MPs) outperformed nanoparticles (NPs) in all experimental conditions tested. Our results indicate that MP modulus enhances particle adhesion in a shear-dependent manner. In low shear human blood flow profiles in vitro, low modulus particles showed favorable adhesion, while at high shear, rigid particles showed superior adhesion. This was confirmed in vivo by studying particle adhesion under venous shear profiles in a mouse model of mesenteric inflammation, where MP adhesion was 127% greater (p < 0.0001) for low modulus particles compared to more rigid ones. Mechanistically, we establish that particle collisions with leukocytes drive these trends, rather than differences in particle deformation, localization, or detachment. Overall, this work demonstrates the importance of VTC modulus as a design parameter for enhanced VTC interaction with vascular walls, and thus, contributes important knowledge for development of successful clinical theranostics with applications for many diseases.

Bactericidal Hydrogels via Surface Functionalization with Cecropin A.

The immobilization of antimicrobial peptides (AMPs) to surfaces, enabling their utilization in biosensor and antibacterial/antifouling coating applications, is typically performed using rigid, solid support materials such as glass or gold and may require lengthy, temperamental protocols. Here, we employ a hydrogel immobilization platform to afford facile fabrication and surface functionalization while offering improved biocompatibility for evaluating the influence of linker length, surface density, and AMP conjugation site on retained peptide activity. Rapid, interfacial photo-polymerization using the radical-mediated thiol-ene addition mechanism was used to generate cross-linked, polymeric coatings bearing residual thiol moieties on prefabricated poly(ethylene glycol) (PEG)-based hydrogel supports. The photo-polymerized coatings were 60 µm thick and contained 0.55 nmol of unreacted free thiols, corresponding to a concentration of 410 µM, for use as cecropin A (CPA) immobilization handles via thiol-maleimide conjugation, where the CPA-bound maleimide moiety was localized at either the carboxyl terminus or midsequence between Ala22 and Gly23. Surface presentation of the thiol handles was controlled by varying the thiolated PEG monomer (PEGSH) used in the photo-polymerizable formulation. Bactericidal activity of CPA functionalized hydrogels against E. coli K235 indicated that CPA immobilized at the carboxyl terminus killed 94 ± 6% of the inoculated pathogens when coatings were prepared with high molecular weight PEGSH and 99 ± 1% when prepared with low molecular weight PEGSH. E. coli cell death demonstrated a stronger dependence on peptide concentration than PEG linker length or degree of thiol functionalization, with activity ranging from 34 ± 13% to 99 ± 1% bacterial cells killed as the prefunctionalization thiol concentration in the coatings was increased from 90 to 990 µM. Finally, the immobilization site on the surface-bound CPA strongly affected antibacterial activity; when midsequence modified CPA was bound to a hydrogel coating bearing 990 µM thiol, only 20 ± 4% of the E. coli population was killed.

Expanding the Alternating Propagation-Chain Transfer-Based Polymerization Toolkit: The Iodo-Ene Reaction.

Analogous to the thiol-ene and phosphane-ene polymerizations, radical-mediated iodo-ene reactions are described here that proceed via alternating propagation and chain transfer (i.e., APT) reactions between perfluoroiodide- and vinyl-bearing monomers. The thermal polymerization of a diiodo/tetraene formulation yielded a cross-linked, homogeneous polymer that was approximately seven times as radiopaque as aluminum owing to its high iodine content. Visible-light photopolymerizations of model iodo-ene monomers were monitored using mid-IR spectroscopy, revealing that the perfluoroiodide functional group consumption exceeded that of the vinyl, a discrepancy that decreased with increasing irradiation intensities and hence polymerization rates. The functional group conversions in resin formulations with a large initial perfluoroiodide excess exacerbated secondary side reactions that led to off-stoichiometric functional group consumption; nevertheless, photopolymerization of resin formulations with excess vinyl stoichiometry proceeded according to the ideal APT mechanism.

Hexaarylbiimidazoles as visible light thiol-ene photoinitiators.

OBJECTIVES: The aim of this study is to determine if hexaarylbiimidazoles (HABIs) are efficient, visible light-active photoinitiators for thiol-ene systems. We hypothesize that, owing to the reactivity of lophyl radicals with thiols and the necessarily high concentration of thiol in thiol-ene formulations, HABIs will effectively initiate thiol-ene polymerization upon visible light irradiation. METHODS: UV-vis absorption spectra of photoinitiator solutions were obtained using UV-vis spectroscopy, while EPR spectroscopy was used to confirm radical species generation upon HABI photolysis. Functional group conversions during photopolymerization were monitored using FTIR spectroscopy, and thermomechanical properties were determined using dynamic mechanical analysis. RESULTS: The HABI derivatives investigated exhibit less absorptivity than camphorquinone at 469nm; however, they afford increased sensitivity at this wavelength when compared with bis(2,4,6-trimethylbenzoyl)-phenylphosphineoxide. Photolysis of the investigated HABIs affords lophyl radicals. Affixing hydroxyhexyl functional groups to the HABI core significantly improved solubility. Thiol-ene resins formulated with HABI photoinitiators polymerized rapidly upon irradiation with 469nm. The glass transition temperatures of the thiol-ene resin formulated with a bis(hydroxyhexyl)-functionalized HABI and photopolymerized at room and body temperature were 49.5±0.5°C and 52.2±0.1°C, respectively. SIGNIFICANCE: Although thiol-enes show promise as continuous phases for composite dental restorative materials, they show poor reactivity with the conventional camphorquinone/tertiary amine photoinitiation system. Conversely, despite their relatively low visible light absorptivity, HABI photoinitiators afford rapid thiol-ene photopolymerization rates. Moreover, minor structural modifications suggest pathways for improved HABI solubility and visible light absorption.

Rapid, Puncture-Initiated Healing via Oxygen-Mediated Polymerization.

Autonomously healing materials that utilize thiol-ene polymerization initiated by an environmentally borne reaction stimulus are demonstrated by puncturing trilayered panels, fabricated by sandwiching thiol-ene-trialkylborane resin formulations between solid polymer panels, with high velocity projectiles; as the reactive liquid layer flows into the entrance hole, contact with atmospheric oxygen initiates polymerization, converting the liquid into a solid plug. Using infrared spectroscopy, we find that formulated resins polymerize rapidly, forming a solid polymer within seconds of atmospheric contact. During high-velocity ballistics experiments, additional evidence for rapid polymerization is provided by high-speed video, demonstrating the immediate viscosity increase when the thiol-ene-trialkylborane resins contact atmospheric oxygen, and thermal imaging, where surface temperature measurements reveal the thiol-ene reaction exotherm, confirming polymerization begins immediately upon oxygen exposure. While other approaches for materials self-repair have utilized similar liquid-to-solid transitions, our approach permits the development of materials capable of sealing a breach within seconds, far faster than previously described methods.

Covalent adaptable networks as dental restorative resins: stress relaxation by addition-fragmentation chain transfer in allyl sulfide-containing resins.

OBJECTIVES: The aim is to demonstrate significant polymerization shrinkage stress reduction in model resins through incorporation of addition-fragmentation chain transfer moieties that promote network stress accommodation by molecular rearrangement. Monomers containing allyl sulfide linkages are incorporated to affect the shrinkage stress that arises during photopolymerization of model resins that contain an initiator and dimethacrylates. Radical-mediated allyl sulfide addition-fragmentation is enabled during polymerization. We hypothesize that allyl sulfide incorporation into methacrylate polymerizations promotes stress relaxation by enabling network adaptation. METHODS: A 1:2 mixture of tetrathiol and allyl sulfide-containing divinyl ethers is formulated with glass-forming dimethacrylates and compared to controls where the allyl sulfide is replaced with a propyl sulfide that is incapable of undergoing addition-fragmentation. Simultaneous shrinkage stress and functional group conversion measurements are performed. The T(g) is determined by DMA. RESULTS: Increasing allyl sulfide concentration reduces the relative stress by up to 75% in the resins containing the maximum amount of allyl sulfide. In glassy systems, at much lower allyl sulfide concentrations, the stress is reduced by up to 20% as compared to propyl sulfide-containing systems incapable of undergoing addition-fragmentation chain transfer. SIGNIFICANCE: Shrinkage stress reduction, typically accompanying free-radical polymerization, is a primary focus in dental materials research and new product development. Allyl sulfide addition-fragmentation chain transfer is utilized as a novel approach to reduce stress in ternary thiol-ene-methacrylate polymerizations. The stress reduction effect depends directly on the allyl sulfide concentration in the given ternary systems, with stress reduction observed even in systems possessing super-ambient T(g)s and low allyl sulfide concentrations.

Stress Relaxation by Addition-Fragmentation Chain Transfer in Highly Crosslinked Thiol-Yne Networks.

Radical mediated addition-fragmentation chain transfer of mid-chain allyl sulfide functional groups was utilized to reduce polymerization-induced shrinkage stress in thiol-yne step-growth photopolymerization reactions. In previous studies, the addition-fragmentation of allyl sulfide during the polymerization of a step-growth thiol-ene network demonstrated reduced polymerization stress; however, the glass transition temperature of the material was well below room temperature (~ -20°C). Many applications require super-ambient glass transition temperatures, such as microelectronics and dental materials. Polymerization reactions utilizing thiol-yne functional groups have many of the advantageous attributes of the thiol-ene-based materials, such as possessing a delayed gel-point, resistant to oxygen inhibition, and fast reaction kinetics, while also possessing a high glass transition temperature. Here we incorporate allyl sulfide functional groups into a highly crosslinked thiol-yne network to reduce polymerization-induced shrinkage stress. Simultaneous shrinkage stress and functional group conversion measurements were performed during polymerization using a cantilever-type tensometer coupled with a FTIR spectrometer. The resulting networks were highly crosslinked, possessed super-ambient glass transition temperatures, and exhibited significantly reduced polymerization-induced shrinkage stress when compared with analogous propyl sulfide-containing materials that are incapable of addition-fragmentation.

Covalent Adaptable Networks (CANs): A Unique Paradigm in Crosslinked Polymers.

Polymer networks possessing reversible covalent crosslinks constitute a novel material class with the capacity for adapting to an externally applied stimulus. These covalent adaptable networks (CANs) represent a trend in polymer network fabrication towards the rational design of structural materials possessing dynamic characteristics for specialty applications. Herein, we discuss the unique attributes of CANs that must be considered when designing, fabricating, and characterizing these smart materials that respond to either thermal or photochemical stimuli. While there are many reversible reactions which to consider as possible crosslink candidates in CANs, there are very few that are readily and repeatedly reversible. Furthermore, characterization of the mechanical properties of CANs requires special consideration owing to their unique attributes. Ultimately, these attributes are what lead to the advantageous properties displayed by CANs, such as recyclability, healability, tunability, shape changes, and low polymerization stress. Throughout this perspective, we identify several trends and future directions in the emerging field of CANs that demonstrate the progress to date as well as the essential elements that are needed for further advancement.