Cationic oligo-p-phenylene ethynylenes have shown much promise as broad-spectrum light-activated antimicrobial compounds against both Gram-positive and Gram-negative bacteria. The anionic varieties, however, have weak biocidal activity. In this study, a complex is formed between a weakly biocidal anionic oligomer and a cationic surfactant, and the effects on their biocidal activity against Gram-negative E. coli and Gram-positive S. aureus are explored. The enhancement in biocidal activity that is observed when the complex is irradiated suggests that interfacial surfactant gives the complex a net-positive charge, allowing it to associate strongly with the bacterial membrane. The results of this study demonstrate a method for the enhancement of biocidal activity of singlet-oxygen sensitizers and corroborate the use of surfactants as trans-membrane drug-delivery agents.

Antibiotic-resistant bacterial infections have become a serious public health threat. In an effort to address this threat, we develop an imidazolium-functionalized conjugated polyelectrolyte that exhibits profound light-activated biocidal activity. Here we report the synthesis, photophysical properties, and biocidal activity of a regioregular head-to-tail polythiophene substituted with cationic imidazolium units (P3HT-Im) prepared by Grignard metathesis controlled polymerization. In water, P3HT-Im has a broad absorption in the visible region and exhibited a remarkably high biocidal efficiency with both Gram-positive and Gram-negative bacteria at sub-microgram per milliliter concentrations. Moreover, mammalian cell studies suggest that P3HT-Im is nontoxic to mammalian cells at concentrations of ≤20 μg/mL over a short time scale (≤1 h) in the dark and light, and the targeting rate-dependent selective mechanism is revealed. This study demonstrates the capability of P3HT-Im to achieve selective imaging and inactivation of bacteria over mammalian cells, suggesting that the polymer has significant potential for ameliorating antibiotic-resistant bacteria in a clinical setting.

A wide range of oligo-p-phenylene ethynylenes has been shown to exhibit good biocidal activity against both Gram-negative and Gram-positive bacteria. While cell death may occur in the dark, these biocidal compounds are far more effective in the light as a result of their ability to sensitize the production of cell-damaging reactive oxygen species. In these studies, the interactions of a specific cationic oligo-p-phenylene ethynylene with spore-forming Bacillus atrophaeus and Bacillus anthracis Sterne have been investigated. Flow cytometry assays are used to rapidly monitor cell death as well as spore germination. This compound effectively killed Bacillus anthracis Sterne vegetative cells (over 4 log reduction), presumably by severe perturbations of the bacterial cell wall and cytoplasmic membrane, while also acting as an effective spore germinant in the dark. While 2 log reduction of B. anthracis Sterne spores was observed, it is hypothesized that further killing could be achieved through enhanced germination.

Chemical and biological sensors are sought for their ability to detect enzymes as biomarkers for symptoms of various disorders, or the presence of chemical pollutants or poisons. p-Phenylene ethynylene oligomers with pendant charged groups have been recently shown to have ideal photophysical properties for sensing. In this study, one anionic and one cationic oligomer are combined with substrates that are susceptible to enzymatic degradation by phospholipases or acetylcholinesterases. The photophysical properties of the J-aggregated oligomers with the substrate are ideal for sensing, with fluorescence quantum yields of the sensors enhanced between 30 and 66 times compared to the oligomers without substrate. The phospholipase sensor was used to monitor the activity of phospholipase A1 and A2 and obtain kinetic information, though phospholipase C did not degrade the sensor. The acetylcholinesterase sensor was used to monitor enzyme activity and was also used to detect the inhibition of acetylcholinesterase by three different inhibitors. Phospholipase A2 is a biomarker for heart and circulatory disease, and acetylcholinesterase is a biomarker for Alzheimer’s, and indicative of exposure to certain pesticides and nerve agents. This work shows that phenylene ethynylene oligomers can be tailored to enzyme-specific sensors by careful selection of substrates that induce formation of a molecular aggregate, and that the sensing of enzymes can be extended to enzyme kinetics and detection of inhibition. Furthermore, the aggregates were studied through all-atom molecular dynamics, providing a molecular-level view of the formation of the molecular aggregates and their structure.Keywords: biomarkers; biosensing; chemical sensing; enzyme sensing; fluorescence quenching; molecular aggregation

Mitigation of bacterial adhesion and subsequent biofilm formation is quickly becoming a strategy for the prevention of hospital-acquired infections. We demonstrate a basic strategy for surface modification that combines the ability to control attachment by microbes with the ability to inactivate microbes. The surface consists of two active materials: poly(p-phenylene ethynylene)-based polymers, which can inactivate a wide range of microbes and pathogens, and poly(N-isopropylacrylamide)-based polymers, which can switch between an hydrophobic “capture” state and a hydrophilic “release” state. The combination of these materials creates a surface that can both bind microbes in a switchable way and kill surface-bound microbes efficiently. Considerable earlier work with cationic poly(p-phenylene ethynylene) polyelectrolytes has demonstrated and characterized their antimicrobial properties, including the ability to efficiently destroy or deactivate Gram-negative and Gram-positive bacteria, fungi, and viruses. Similarly, much work has shown (1) that surface-polymerized films of poly(N-isopropylacrylamide) are able to switch their surface thermodynamic properties from a swollen, relatively hydrophilic state at low temperature to a condensed, relatively hydrophobic state at higher temperature, and (2) that this switch can control the binding and/or release of microbes to poly(N-isopropylacrylamide) surfaces. The active surfaces described herein were fabricated by first creating a film of biocidal poly(p-phenylene ethynylene) using layer-by-layer methods, and then conferring switchable adhesion by growing poly(N-isopropylacrylamide) through the poly(p-phenylene ethynylene) layer, using surface-attached polymerization initiators. The resulting multifunctional, complex films were then characterized both physically and functionally. We demonstrate that such films kill and subsequently induce widespread release of Gram-negative and Gram-positive bacteria.Keywords: bacteria; conjugated polyelectrolyte; kill; PNIPAAm; release

This article reports an investigation of the photophysical properties and the light- and dark-biocidal activity of two poly(phenyleneethynylene) (PPE)-based conjugated polyelectrolytes (CPEs) bearing cationic imidazolium solubilizing groups. The two polymers feature the same PPE-type backbone, but they differ in the frequency of imidazoliums on the chains: PIM-4 features two imidazolium units on every phenylene repeat, whereas PIM-2 contains two imidazolium units on every other phenylene unit. Both polymers are very soluble in water and polar organic solvents, but their propensity to aggregate in water differs with the density of the imidazolium units. The polymers are highly fluorescent, and they exhibit the amplified quenching effect when exposed to a low concentration of anionic electron-acceptor anthraquinone disulfonate. The CPEs are also quenched by a relatively low concentration of pyrophosphate by an aggregation-induced quenching mechanism. The biocidal activity of the cationic imidazolium CPEs was studied against both Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus bacteria in the dark and under blue-light illumination. Both polymers are effective biocides, exhibiting greater than 3 log kill with 30–60 min of light exposure at concentrations of ≤10 μg mL–1.Keywords: antimicrobial polymers; biocides; conjugated polyelectrolytes; ionic liquids; singlet oxygen; triplet state

Finding new optical probes to detect and track amyloid protein aggregates is key to understanding and defeating the myriad of neurodegenerative and other diseases associated with these misfolded proteins. Herein we report that a series of fluorescent, soluble oligo(p-phenylene ethynylene)s (OPEs) are able to detect amyloids in vitro by massive binding-activated superluminescence, with low micromolar affinity and high selectivity for the amyloid conformer. The OPEs track the kinetics of amyloid fibril formation from native hen egg white lysozyme (HEWL) similarly to thioflavin T (ThT), and the dependence of binding affinity on OPE length supports the theory of a linear binding groove. We hypothesize, based on spectral properties, induced circular dichroism, and previous work in analogous systems, that the fluorescence turn-on mechanism is a combination of the reduction of static solvent-mediated quenching at the ethyl ester end groups of the phenylene ethynylene fluorophore and the formation of chiral J-type aggregates templated on the amyloid fibril surface.Keywords: Alzheimer’s disease; amyloid diseases; amyloid fibril staining; conjugated oligoelectrolytes; fluorescent optical probes; J aggregates; protein aggregate detection

Cationic oligo-p-phenylene ethynylenes are highly effective light-activated biocides that deal broad-spectrum damage to a variety of pathogens, including bacteria. A potential problem arising in the long-term usage of these compounds is photochemical breakdown, which nullifies their biocidal activity. Recent work has shown that these molecules complex with oppositely-charged surfactants, and that the resulting complexes are protected from photodegradation. In this manuscript, we determine the biocidal activity of an oligomer and a complex formed between it and sodium dodecyl sulfate. The complexes are able to withstand prolonged periods of irradiation, continuing to effectively kill both Gram-negative and Gram-positive bacteria, while the oligomer by itself loses its biocidal effectiveness quickly in the presence of light. In addition, damage and stress responses induced by these biocides in both E. coli and S. aureus are discussed. This work shows that complexation with surfactants is a viable method for long-term light-activated biocidal applications.

Cationic oligo-p-phenylene ethynylenes have shown much promise as broad-spectrum light-activated antimicrobial compounds against both Gram-positive and Gram-negative bacteria. The anionic varieties, however, have weak biocidal activity. In this study, a complex is formed between a weakly biocidal anionic oligomer and a cationic surfactant, and the effects on their biocidal activity against Gram-negative E. coli and Gram-positive S. aureus are explored. The enhancement in biocidal activity that is observed when the complex is irradiated suggests that interfacial surfactant gives the complex a net-positive charge, allowing it to associate strongly with the bacterial membrane. The results of this study demonstrate a method for the enhancement of biocidal activity of singlet-oxygen sensitizers and corroborate the use of surfactants as trans-membrane drug-delivery agents.

The antifungal activities of poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPEs) and oligo-phenylene ethynylenes (OPEs) were investigated using Saccharomyces cerevisiae (S. cerevisiae) as a model pathogen. The effect of the CPE and OPE materials on the vegetative cells and ascospores were tested in the dark or with UV-irradiation. A number of the tested polymers and oligomers significantly reduced the viability of the vegetative yeast cells in the dark, with activities exceeding the commonly used antibiotic Amphotericin B. With UV-irradiation, all of the tested CPEs and OPEs exhibited potent antifungal activities and completely inactivated the yeast cells. In particular, the oligomeric EO-OPE-1(Th, C2) strongly inactivates ascospores with UV-light at a dose level lower than sporicidal agents reported in the literature. Under conditions that promote spore germination, the CPEs and OPEs show efficient activities against the germinated spores. The protein-enriched outer envelope of yeast cells and germinated ascospores appears to serve as a main target for the CPE and OPE antimicrobial materials.Keywords: antifungal; antimicrobial; cationic conjugated polyelectrolytes; cationic oligo-phenylene ethynylenes; sporicidal;

A multiscale investigation was carried out to study the dark and light-enhanced bactericidal mechanisms of poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPEs) and oligo-phenylene ethynylenes (OPEs). On the morphological scale, Gram-negative E. coli cells exposed to CPE and OPE compounds in the dark show damage to the cell envelope, plasma membrane, and in some cases the cytoplasm, while with UV-irradiation, E. coli sustained catastrophic damages to both the cell envelope and cytoplasm. In contrast, the Gram-positive S. epi bacteria appeared intact when exposed to CPE and OPE compounds in the dark but showed damages to the cell envelope with UV-irradiation. To better understand the molecular basis of CPE- and OPE-induced morphological changes and damages to bacteria, we investigated the effect of these compounds on model bacterial plasma membrane and bacterial proteins and plasmid DNA. Measurements of dark membrane perturbation activity of the CPEs and OPEs using model lipid membranes support a carpet or detergent-like mechanism by which the antimicrobial compounds induce membrane collapse and phase transitions. Under UV-irradiation, E. coli bacteria exposed to CPEs and OPEs showed covalent modifications and damages to both cellular protein and plasmid DNA, likely through oxidative pathways mediated by singlet oxygen and subsequent reactive oxygen species sensitized by the CPE and OPE compounds. Our finding thus show that the antimicrobial polymers and oligomers exert toxicity toward Gram-negative bacteria by disrupting the morphology and structures of cell envelope and cytoplasm, including cellular components such as proteins and DNA, while exert toxicity toward Gram-positive bacteria by binding to and disrupting just the cell wall.

This Feature Article focuses on recent progress made in elucidating the intermolecular interactions between a novel class of synthetic phenylene ethynylene (PPE)-based conjugated polyelectrolyte polymers (CPEs) and oligomers (OPEs) and multiscale cellular targets that give rise to their remarkable broad spectrum biocidal activity. We first review the interactions and self-assembly behaviors of the CPEs and OPEs with a set of vital biomolecules, including lipids, proteins, and nucleic acids, that reveal the potential pathways by which synthetic biocidal agents could exert toxicity. An overview of the antimicrobial effects and mechanisms of the CPEs and OPEs on multiple clinically relevant pathogens is then presented, with an emphasis on the morphological damage induced by the biocidal compounds toward the pathogens. Finally, we discuss the cytotoxicity of these materials against mammalian cells and human tissues to explore the potential applications of the CPEs and OPEs as antiseptics. We also pose some unanswered questions about their antimicrobial mechanisms, which provide direction for a future study.

Poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPEs) and cationic phenylene ethynylene oligomers (OPEs) exhibit broad-spectrum antimicrobial activity and one of their main targets is believed to be the bacterial cell wall and membrane. This review focuses on recent progress made in elucidating the membrane perturbation mechanism of these antimicrobial molecules. We first review the membrane action models proposed for naturally occurring antimicrobial peptides (AMPs) because the membrane activities of CPEs and OPEs and AMPs are believed to be mediated by the same physical and chemical driving forces. An overview of the current research progress on the interactions between CPE and OPE compounds and model membranes is then presented, including main parameters controlling the membrane activity of these synthetic antimicrobial compounds. Combined with direct visualization of CPE and OPE induced changes in bacterial cell morphology, evidence to date points to a carpet or detergent-like pathway by which these compounds perturb the lipid membrane. Lastly, general trends between the membrane activity and dark antimicrobial activity of the CPEs and OPEs are reviewed to shed light on the structure–function relationship of this novel class of biocidal compounds.

Cationic end-only-functionalized oligo(arylene-ethynylene)s (EO-OPEs) have recently been found to be broad-spectrum and effective antimicrobial agents because of their unique structure and optical properties. In this study, we investigated their potential use for preventing and reducing Escherichia coli (E. coli) biofilms. The Calgary biofilm device (CBD) was used to form bacterial biofilms of E. coli; in these studies, the minimum inhibitory concentration (MIC) and the minimum biofilm eradication concentration (MBEC) were determined. E. coli biofilms uniformly grow on pegs of the CBD device lid. The MIC values determined for EO-OPEs are comparable to those found for standard antibiotics such as kanamycin (MIC = 11.2 μg/mL). About 10–30 times the concentration of EO-OPEs was required to eradicate E. coli biofilms and prevent regrowth in the dark. Near-UV irradiation of EO-OPEs enhanced their efficacy in killing biofilms.

The bactericidal mechanisms of poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPE) and oligo-phenylene ethynylenes (OPE) were investigated using electron/optical microscopy and small-angle X-ray scattering (SAXS). The ultrastructural analysis shows that polymeric PPE-Th can significantly remodel the bacterial outer membrane and/or the peptidoglycan layer, followed by the possible collapse of the bacterial cytoplasm membrane. In contrast, oligomeric end-only OPE (EO-OPE) possesses potent bacteriolysis activity, which efficiently disintegrates the bacterial cytoplasm membrane and induces the release of bacterial cell content. Using single giant vesicles and SAXS, we demonstrated that the membrane perturbation mechanism of EO-OPE against model bacterial membranes results from a 3D membrane phase transition or perturbation.

The antiviral activities of poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPE) and oligo-phenylene ethynylenes (OPE) were investigated using two model viruses, the T4 and MS2 bacteriophages. Under UV/visible light irradiation, significant antiviral activity was observed for all of the CPEs and OPEs; without irradiation, most of these compounds exhibited high inactivation activity against the MS2 phage and moderate inactivation ability against the T4 phage. Transmission electron microscopy (TEM) and SDS polyacrylamide gel electrophoresis (SDS-PAGE) reveal that the CPEs and OPEs exert their antiviral activity by partial disassembly of the phage particle structure in the dark and photochemical damage of the phage capsid protein under UV/visible light irradiation.Keywords: antiviral; cationic conjugated polyelectrolytes; cationic oligo-phenylene ethynylenes; MS2 phage; photoinduced antimicrobial activity; T4 phage;

We demonstrate herein a method for chemically modifying cotton fibers and cotton-containing fabric with a light-activated, cationic phenylene–ethynylene (PPE-DABCO) conjugated polyelectrolyte biocide. When challenged with Pseudomonas aeruginosa and Bacillus atropheaus vegetative cells from liquid suspension, light-activated PPE-DABCO effects 1.2 and 8 log, respectively, losses in viability of the exposed bacteria. These results suggest that conjugated polyelectrolytes retain their activity when grafted to fabrics, showing promise for use in settings where antimicrobial textiles are needed.Keywords: Bacillus atrophaeus; biocidal fibers; light activation; poly(phenylene−ethynylene) conjugated polyelectrolytes; Pseudomonas aeruginosa; XTT;

This Spotlight on Applications provides an overview of a research program that has focused on the development and mechanistic study of cationic conjugated polyelectrolytes (CPEs) that function as light- and dark-active biocidal agents. Investigation has centered on poly-(phenylene ethynylene) (PPE) type conjugated polymers that are functionalized with cationic quaternary ammonium solubilizing groups. These polymers are found to interact strongly with Gram-positive and Gram-negative bacteria, and upon illumination with near-UV and visible light act to rapidly kill the bacteria. Mechanistic studies suggest that the cationic PPE-type polymers efficiently sensitize singlet oxygen (1O2), and this cytotoxic agent is responsible for initiating the sequence of events that lead to light-activated bacterial killing. Specific CPEs also exhibit dark-active antimicrobial activity, and this is believed to arise due to interactions between the cationic/lipophilic polymers and the negatively charged outer membrane characteristic of Gram-negative bacteria. Specific results are shown where a cationic CPE with a degree of polymerization of 49 exhibits pronounced light-activated killing of E. coli when present in the cell suspension at a concentration of 1 μg mL–1.Keywords: amplified quenching; antibacterial; antimicrobial; conjugated polyelectrolytes; light activated biocides; singlet oxygen;

A poly(phenylene ethynylene) conjugated polymer (PPE-NMe3+-COO−) containing tetraalkylammonium groups and carboxylate groups has been synthesized by Sonogashira coupling. Due to the presence of the strong cationic and weak anionic pendant units, the polymer undergoes a pH-induced transition from cationic polyelectrolyte to polyampholyte due to deprotonation of the carboxylic acid units in basic solution. Studies of the pH dependence of the polymers’ optical properties reveal changes in absorption oscillator strength and fluorescence quantum efficiency that are triggered by the transition from cationic polyelectrolyte to polyampholyte nature. Stern−Volmer fluorescence quenching of PPE-NMe3+-COO− with a negatively charged quencher 1,4,5,8-naphthalenediimide-N,N-bis(methylsulfonate) (NDS) shows that the polymer fluorescence quenching is amplified at low pH where the polymer is a polycation, whereas the quenching efficiency is considerably less at high pH.

The interactions of poly(phenylene ethynylene)- (PPE-) based cationic conjugated polyelectrolytes (CPEs) and oligo(phenylene ethynylene)s (OPEs) with different model lipid membrane systems were investigated to gain insight into the relationship between molecular structure and membrane perturbation ability. The CPE and OPE compounds exhibit broad-spectrum antimicrobial activity, and cell walls and membranes are believed to be their main targets. To better understand how the size, in terms of the number of repeat units, of the CPEs and OPEs affects their membrane disruption activities, a series of PPE-based CPEs and OPEs were synthesized and studied. A number of photophysical techniques were used to investigate the interactions of CPEs and OPEs with model membranes, including unilamellar vesicles and lipid monolayers at the air/water interface. CPE- or OPE-induced dye leakage from vesicles reveals that the CPEs and OPEs selectively perturb model bacterial membranes and that their membrane perturbation abilities are highly dependent on molecular size. Consistent with dye-leakage assay results, the CPEs and OPEs also exhibit chain-length-dependent ability to insert into 1,2-dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DPPG) monolayers. Our results suggest that, for PPE-based CPE and OPE antimicrobials, chain length can be tuned to optimize their membrane perturbation ability.

Cationic poly(phenylene ethynylene)- (PPE-) based conjugated polyelectrolytes (CPEs) with six different chain lengths ranging in degree of polymerization from ∼7 to ∼49 were synthesized from organic-soluble precursor polymers. The molecular weight of the precursor polymers was controlled by the amount of a monofunctional “end-capping” agent added to the polymerization reaction. Cationic CPEs were prepared by quaternization of amine groups to tetraalkylammonium groups. Their structure–property relationships were investigated by observing their photophysical properties and antibacterial activity. The polymers were found to exhibit a chain-length dependence in their photophysical properties. It has also been observed that the polymers exhibit effective antibacterial activity against both Gram-positive and Gram-negative bacteria under UV irradiation, whereas they show little antibacterial activity in the dark. An effect of chain length on the light-activated antibacterial activity was also found: The shortest polymer (n = 7) exhibited the most effective antibacterial activity against both Gram-positive and Gram-negative bacteria.

It is essential to develop alternative strategies to treat infections, especially those infections caused by Staphylococcus aureus, which is responsible for most skin infections. Among those strategies, light-induced inactivation of pathogens appears to be a promising candidate. We present four novel “end only” oligo(phenylene ethynylene)s (EO-OPE-1s) that have the ends functionalized with cationic groups and are powerful light-activated biocides against Escherichia coli, Staphylococcus epidermidis, and S. aureus. We have correlated the light-induced biocidal activities with singlet oxygen quantum yields Φ (1O2) of EO-OPE-1s, and a higher Φ (1O2) correlates with a better light-induced biocidal activity. Coupled with our previous work on the interactions of EO-OPE-1s with dioleoyl-sn-glycero-3-phosphocholine (DOPC)/cholesterol vesicles, we believe the biocidal process involves the following: (1) EO-OPE-1s penetrate the bacterial membrane, (2) EO-OPE-1s photosensitize the generation of singlet oxygen and/or other reactive oxygen species, and (3) singlet oxygen and/or reactive oxygen species trigger the cytotoxicity.Keywords (keywords): antimicrobials; biocidal activity; oligo(phenylene ethynylene)s; OPE derivatives; photodynamic activity; reactive oxygen species; singlet oxygen;

The interactions of antimicrobial cationic conjugated polyelectrolytes (CPEs) with two model membranes, liposomes and lipid monolayers at the air−water interface, have been investigated by fluorescence emission, fluorescence quenching, pressure−area isotherm, and dynamic light scattering measurements. This study continues the evaluation of the antimicrobial mechanism of a cationic poly(phenylene ethynylene) (PPE)-based CPE (polymer 1), which contains a 2,5-thienylene moiety in the repeat unit. To this end, the interactions of polymer 1 with lipids with varying headgroup charge and acyl chain length have been examined. Our results show that the catioic polymer 1 can efficiently associate with and insert into anionic phosphatidylglycerol (PG) membranes. However, polymer 1 does not exhibit any interactions with zwitterionic lipid membranes composed of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) lipids. Polymer 1’s selective affinity toward anionic lipids over zwitterionic lipids makes it an attractive antimicrobial agent with low toxicity. The interactions of polymer 1 with lipid membranes of different fluidity were studied by varying the surface pressure of lipid monolayers and by adjusting the temperature of liposomes. We observe that increasing membrane fluidity enhances both the conformational changes of polymer 1 upon associating with lipid membranes and the extent of polymer 1 insertion into lipid monolayers. We also find that the thickness of the lipid bilayers, modulated by acyl chain length, affects the extent of polymer 1 incorporation into the lipid bilayer.

Microcapsules consisting of alternating layers of oppositely charged poly(phenylene ethynylene)-type conjugated polyelectrolytes (CPEs) were prepared via layer-by-layer deposition onto MnCO3 template particles followed by dissolution of the template particles using an ethylenediaminetetraacetate solution. The resulting microcapsules exhibit bright-green fluorescence emission characteristics of the CPEs. Strong antimicrobial activity was observed upon mixing of polyelectrolyte capsules with Cobetia marina or Pseudomonas aeruginosa followed by white-light irradiation. It was demonstrated that the materials act as highly effective light-activated micro “Roach Motels” with greater than 95% kill after exposure to ∼1 h of white light.Keywords: biocide; confocal microscopy; conjugated polymer; polyelectrolyte capsule; singlet oxygen

A study of the synthesis, photophysical behavior and self-assembly of a series of symmetrical cationic oligophenylene ethynylenes is reported. Aqueous solutions of all these compounds exhibit strong and structured absorption in the ultraviolet (maxima near 305 nm and 355 nm) with a broad fluorescence in the range 370–600 nm. While all of these compounds show strong fluorescence in methanol, the fluorescence yields in water are variable and substituent dependent. Transient absorption, presumably attributable to a triplet is found for all of the oligomers studied thus far. All of the compounds having a net positive charge exhibit strong complex formation when they are added to aqueous solutions containing the anionic biopolymer carboxymethylcellulose. The complexes are characterized by a pronounced red shift in the oligomer absorption spectrum and a red shifted and generally intense fluorescence. We attribute the spectral shifts due to the formation of “J-dimers” in each case. The results of this preliminary study suggest that these compounds may be useful in sensing due to their strong tendency to associate with anionic biomacromolecules.

A pair of cationic phenylene ethynylene oligomers (OPEs) have been synthesized, and their optical properties have been studied in solution with and without added scaffold materials, including carboxymethylcellulose, carboxymethylamylose, and Laponite. The OPEs are strongly fluorescent in methanol solution, but in water, the fluorescence yield is suppressed. The addition of scaffolds to aqueous solutions of OPEs leads to a red shift in the absorption and in most cases a significant increase in the fluorescence quantum yield. The effects most likely arise because of template-induced formation of linear J-dimers or possibly because of planarization, which give rise to an effective increase in the conjugation length of OPEs.

The interactions of antimicrobial poly(phenylene ethynylene) (PPE)-based cationic conjugated polyelectrolytes (CPEs) with lipid membranes were investigated to gain insight into the mechanism of their biocidal activity. Three model membrane systems comprising negatively charged phosphatidylglycerol (PG) lipids were used to mimic the bacterial cell membrane, including unilamellar lipid vesicles in aqueous solution, lipid bilayer coated silica microspheres, and lipid monolayers at the air−water interface. Two PPE CPEs, one containing a thiophene moiety on the PPE repeat unit and the second containing a diazabicyclooctane (DABCO) moiety on the pendant side chain, were chosen, since the former exhibits distinct dark biocidal activity and the latter shows strong light-activated antimicrobial activity but little dark biocidal activity. The interactions of these two PPE polymers with lipid membranes were characterized in detail by CPE fluorescence spectral changes, fluorescence resonance energy transfer (FRET), fluorescence quenching, monolayer insertion, and dynamic light scattering assays. Both PPE polymers exhibit affinity for the anionic lipid membrane systems. Their concomitant association and insertion into the membrane leads to conformational changes of the PPE polymer from an aggregated state to a more extended state, as evidenced by the polymer’s enhanced fluorescence and FRET between the polymer and rhodamine incorporated in the lipid membrane. In comparison, the thiophene polymer exhibits stronger interactions with PG lipid membranes than the DABCO-containing polymer. The former induces a larger fluorescence enhancement, shows faster transfer across the lipid membrane, and inserts more readily and to a higher extent into lipid monolayers. The observed differences between the two PPE polymers in their interactions with the lipid membrane may stem from their structural differences, as the DABCO-containing polymer has a much bulkier and larger pendant group on its side chain. The higher degree of membrane interaction and insertion, and subsequent membrane disorganization, of the thiophene polymer may account for its dark biocidal activity.

A series of water soluble, cationic conjugated polyelectrolytes (CPEs) with backbones based on a poly(phenylene ethynylene) repeat unit structure and tetraakylammonium side groups exhibit a profound light-induced biocidal effect. The present study examines the biocidal activity of the CPEs, correlating this activity with the photophysical properties of the polymers. The photophysical properties of the CPEs are studied in solution, and the results demonstrate that direct excitation produces a triplet excited-state in moderate yield, and the triplet is shown to be effective at sensitizing the production of singlet oxygen. Using the polymers in a format where they are physisorbed or covalently grafted to the surface of colloidal silica particles (5 and 30 μm diameter), we demonstrate that they exhibit light-activated biocidal activity, effectively killing Cobetia marina and Pseudomonas aeruginosa. The light-induced biocidal activity is also correlated with a requirement for oxygen suggesting that interfacial generation of singlet oxygen is the crucial step in the light-induced biocidal activity.

A fluorescence based assay for human serum-derived phospholipase activity has been developed in which cationic conjugated polyelectrolytes are supported on silica microspheres. The polymer-coated beads are overcoated with an anionic phospholipid (1,2-dimyristoyl-sn-glycero-3-[phospho-rac-(1-glycerol)) (DMPG) to provide “lipobeads” that serve as a sensor for PLA2. The lipid serves a dual role as a substrate for PLA2 and an agent to attenuate quenching of the polymer fluorescence by the external electron transfer quencher 9,10-anthraquinone-2,6-disulfonic acid (AQS). In this case quenching of the polymer fluorescence by AQS increases as the PLA2 digests the lipid. The lipid can also be used itself as a quencher and substrate by employing a small amount of energy transfer quencher substituted lipid in the DMPG. In this case the fluorescence of the polymer is quenched when the lipid layer is intact; as the enzyme digests the lipid, the fluorescence of the polymer is restored. The sensing of PLA2 activity has been studied both by monitoring fluorescence changes in a multiwell plate reader and by flow cytometry. The assay exhibits good sensitivity with EC50 values in the nanomolar range.

We describe the development of an optical sensing system for the high-throughput screening (HTS) of a broad range of biological molecules, whole cells, organisms and pathogens, and illustrate the technology applications by a hyaluronidase enzyme activity assay as a specific example. At the core of the technology described in this paper, is the exciton concept that is relevant to molecular aggregation. J-aggregates of cyanine dyes have a narrower, red-shifted absorption band compared to monomer. We demonstrate that self-assembly may be driven by the helicogenic nature of the cyanine dye, converting the linear polymers of hyaluronic acid or carboxymethyl cellulose into supramolecular helical assemblies. This self-assembly is accompanied by an intense, sharp, red-shifted J-aggregate fluorescence. We utilized this property to develop an assay for the enzyme hyaluronidase, based upon the concept of “scaffold destruction,” whereby the disruption/destruction of the hyaluronic acid polymer by hyaluronidase is accompanied by an attenuation of light emission from the J-aggregate. The extent of light attenuation provides an index of hyaluronidase activity. Other polymers of carbohydrates, proteins, nucleic acids and chemical polymers (such as the carbon nanotube) might provide a similar scaffold for helicogenic dyes upon which molecular aggregation can occur. A key feature of these assays is that they are label-free.