Water-soluble extracellular vesicle probes based on conjugated oligoelectrolytes.

We developed a series of transmembrane conjugated oligoelectrolytes (COEs) with tunable optical emissions from the UV to the near IR to address the false-positive problem when detecting nanometer-sized extracellular vesicles (EVs) by flow cytometry. The amphiphilic molecular framework of COEs is defined by a linear conjugated structure and cationic charged groups at each terminal site. Consequently, COEs have excellent water solubility and the absence of nanoaggregates at concentrations up to 50 µM, and unbound COE dyes can be readily removed through ultrafiltration. These properties enable unambiguous and simple detection of COE-labeled small EVs using flow cytometry with negligible background signals. We also demonstrated the time-lapsed tracking of small EV uptake into mammalian cells and the endogenous small EV labeling using COEs. Briefly, COEs provide a class of membrane-targeting dyes that behave as biomimetics of the lipid bilayer and a general and practical labeling strategy for nanosized EVs.

Membrane-intercalating conjugated oligoelectrolytes.

By acting as effective biomimetics of the lipid bilayers, membrane-intercalating conjugated oligoelectrolytes (MICOEs) can spontaneously insert themselves into both synthetic lipid bilayers and biological membranes. The modular and intentional molecular design of MICOEs enable a range of applications, such as bioproduction, biocatalysis, biosensing, and therapeutics. This tutorial review provides a structural evolution of MICOEs, which originated from the broader class of conjugated molecules, and analyses the drivers behind this evolutionary process. Various representative applications of MICOEs, accompanied by insights into their molecular design principles, will be reviewed separately. Perspectives on the current challenges and opportunities in research on MICOEs will be discussed at the end of the review to highlight their potential as unconventional and value-added materials for biological systems.

Conjugated Oligoelectrolytes for Long-Term Tumor Tracking with Incremental NIR-II Emission.

The design and synthesis of the near-infrared (NIR)-II emissive conjugated oligoelectrolyte COE-BBT are reported. COE-BBT has a solubility in aqueous media greater than 50 mg mL-1 , low toxicity, and a propensity to intercalate lipid bilayers, wherein it exhibits a higher emission quantum yield relative to aqueous media. Addition of COE-BBT to cells provides two emission channels, at ≈500 and ≈1020 nm, depending on the excitation wavelength, which facilitates in vitro confocal microscopy and in vivo animal imaging. The NIR-II emission of COE-BBT is used to track intracranial and subcutaneous tumor progression in mice. Of relevance is that the total NIR-II intensity increases over time. This phenomenon is attributed to a progressive attenuation of a COE-BBT self-quenching effect within the cells due to the expected dye dilution per cell as the tumor proliferates.

Bacterial lipopolysaccharide core structures mediate effects of butanol ingress.

The outer membrane (OM) is the first defence for Gram-negative bacteria against their environments making it important in strain improvement for sustainable biobutanol production. While modifying the OM structure using chemical additives could enhance microbial viability, there are currently no model systems that accurately describe OM responses to butanol. Here, we experimentally determined that reducing the lipopolysaccharide (LPS) core length and charge increased Escherichia coli sensitivity to butanol. In silico models were built to describe how OM structure contributes to its ability to withstand butanol under conditions of exposure and production. Consistent with experiments, resistance to ingress of butanol into OMs correlates with both core length and charge, where a lower charge density is more conducive to butanol assimilation. The core length and branching correlate with the lateral spacing of the lipids, suggestive of a role of them in maintaining OM fluidity. In contrast to systems with short-length LPS cores, butanol intercalation into OMs with longer LPS cores increases membrane order and rigidity, which might be due to their more porous internal structure. These findings will assist the development of more butanol-tolerant biobutanol-producing bacteria, where thicker, more compact and less polar LPS-core surfaces reinforce the integrity of OMs and further improve resilience in extreme environments.

A Chain-Elongated Oligophenylenevinylene Electrolyte Increases Microbial Membrane Stability.

A novel conjugated oligoelectrolyte (COE) material, named S6, is designed to have a lipid-bilayer stabilizing topology afforded by an extended oligophenylenevinylene backbone. S6 intercalates biological membranes acting as a hydrophobic support for glycerophospholipid acyl chains. Indeed, Escherichia coli treated with S6 exhibits a twofold improvement in butanol tolerance, a relevant feature to achieve within the general context of modifying microorganisms used in biofuel production. Filamentous growth, a morphological stress response to butanol toxicity in E. coli, is observed in untreated cells after incubation with 0.9% butanol (v/v), but is mitigated by S6 treatment. Real-time fluorescence imaging using giant unilamellar vesicles reveals the extent to which S6 counters membrane instability. Moreover, S6 also reduces butanol-induced lipopolysaccharide release from the outer membrane to further maintain cell integrity. These findings highlight a deliberate effort in the molecular design of a chain-elongated COE to stabilize microbial membranes against environmental challenges.

Informed Molecular Design of Conjugated Oligoelectrolytes To Increase Cell Affinity and Antimicrobial Activity.

Membrane-intercalating conjugated oligoelectrolytes (COEs) are emerging as potential alternatives to conventional, yet increasingly ineffective, antibiotics. Three readily accessible COEs, belonging to an unreported series containing a stilbene core, namely D4, D6, and D8, were designed and synthesized so that the hydrophobicity increases with increasing side-chain length. Decreased aqueous solubility correlates with increased uptake by E. coli. The minimum inhibitory concentration (MIC) of D8 is 4 µg mL-1 against both E. coli and E. faecalis, with an effective uptake of 72 %. In contrast, the MIC value of the shortest COE, D4, is 128 µg mL-1 owing to the low cellular uptake of 3 %. These findings demonstrate the application of rational design to generate efficacious antimicrobial COEs that have potential as low-cost antimicrobial agents.