Synthesis and Biological Properties of Fluorescent Strigolactone Mimics Derived from 1,8-Naphthalimide.

Strigolactones (SLs) have potential to be used in sustainable agriculture to mitigate various stresses that plants have to deal with. The natural SLs, as well as the synthetic analogs, are difficult to obtain in sufficient amounts for practical applications. At the same time, fluorescent SLs would be useful for the mechanistic understanding of their effects based on bio-imaging or spectroscopic techniques. In this study, new fluorescent SL mimics containing a substituted 1,8-naphthalimide ring system connected through an ether link to a bioactive furan-2-one moiety were prepared. The structural, spectroscopic, and biological activity of the new SL mimics on phytopathogens were investigated and compared with previously synthetized fluorescent SL mimics. The chemical group at the C-6 position of the naphthalimide ring influences the fluorescence parameters. All SL mimics showed effects similar to GR24 on phytopathogens, indicating their suitability for practical applications. The pattern of the biological activity depended on the fungal species, SL mimic and concentration, and hyphal order. This dependence is probably related to the specificity of each fungal receptor-SL mimic interaction, which will have to be analyzed in-depth. Based on the biological properties and spectroscopic particularities, one SL mimic could be a good candidate for microscopic and spectroscopic investigations.

Generalized polarization and time-resolved fluorescence provide evidence for different populations of Laurdan in lipid vesicles.

The solvatochromic dye Laurdan is widely used in sensing the lipid packing of both model and biological membranes. The fluorescence emission maximum shifts from about 440 nm (blue channel) in condensed membranes (So) to about 490 nm (green channel) in the liquid-crystalline phase (Lα). Although the fluorescence intensity based generalized polarization (GP) is widely used to characterize lipid membranes, the fluorescence lifetime of Laurdan, in the blue and the green channel, is less used for that purpose. Here we explore the correlation between GP and fluorescence lifetimes by spectroscopic measurements on the So and Lα phases of large unilamellar vesicles of DMPC and DPPC. A positive correlation between GP and the lifetimes is observed in each of the optical channels for the two lipid phases. Microfluorimetric determinations on giant unilamellar vesicles of DPPC and DOPC at room temperature are performed under linearly polarized two-photon excitation to disentangle possible subpopulations of Laurdan at a scale below the optical resolution. Fluorescence intensities, GP and fluorescence lifetimes depend on the angle between the orientation of the linear polarization of the excitation light and the local normal to the membrane of the optical cross-section. This angular variation depends on the lipid phase and the emission channel. GP and fluorescence intensities in the blue and green channel in So and in the blue channel in Lα exhibit a minimum near 90o. Surprisingly, the intensity in the green channel in Lα reaches a maximum near 90o. The fluorescence lifetimes in the two optical channels also reach a pronounced minimum near 90o in So and Lα, apart from the lifetime in the blue channel in Lα where the lifetime is short with minimal angular variation. To our knowledge, these experimental observations are the first to demonstrate the existence of a bent conformation of Laurdan in lipid membranes, as previously suggested by molecular dynamics calculations.

Concerning the photophysics of fluorophores towards tailored bioimaging compounds: a case study involving S100A9 inflammation markers.

A full understanding concerning the photophysical properties of a fluorescent label is crucial for a reliable and predictable performance in biolabelling applications. This holds true not only for the choice of a fluorophore in general, but also for the correct interpretation of data, considering the complexity of biological environments. In the frame of a case study involving inflammation imaging, we report the photophysical characterization of four fluorescent S100A9-targeting compounds in terms of UV-vis absorption and photoluminescence spectroscopy, fluorescence quantum yields (ΦF) and excited state lifetimes (τ) as well as the evaluation of the radiative and non-radiative rate constants (kr and knr, respectively). The probes were synthesized based on a 2-amino benzimidazole-based lead structure in combination with commercially available dyes, covering a broad color range from green (6-FAM) over orange (BODIPY-TMR) to red (BODIPY-TR) and near-infrared (Cy5.5) emission. The effect of conjugation with the targeting structure was addressed by comparison of the probes with their corresponding dye-azide precursors. Additionally, the 6-FAM and Cy5.5 probes were measured in the presence of murine S100A9 to determine whether protein binding influences their photophysical properties. An interesting rise in ΦF upon binding of 6-FAM-SST177 to murine S100A9 enabled the determination of its dissociation equilibrium constant, reaching up to KD = 324 nM. This result gives an outlook for potential applications of our compounds in S100A9 inflammation imaging and fluorescence assay developments. With respect to the other dyes, this study demonstrates how diverse microenvironmental factors can severely impair their performance while rendering them poor performers in biological media, showing that a preliminary photophysical screening is key to assess the suitability of a particular luminophore.

Fracture Detection in Bio-Glues with Fluorescent-Protein-Based Optical Force Probes.

Glues are being used to bond, seal, and repair in industry and biomedicine. The improvement of gluing performance is hence important for the development of new glues with better and balanced property spaces, which in turn necessitates a mechanistic understanding of their mechanical failure. Optical force probes (OFPs) allow the observation of mechanical material damage in polymers from the macro- down to the microscale, yet have never been employed in glues. Here, the development of a series of ratiometric OFPs based on fluorescent-protein-dye and protein-protein conjugates and their incorporation into genetically engineered bio-glues is reported. The OFPs are designed to efficiently modulate Förster resonance energy transfer upon force application thereby reporting on force-induced molecular alterations independent of concentration and fluorescence intensity both spectrally and through their fluorescence lifetime. By fluorescence spectroscopy in solution and in the solid state and by fluorescence lifetime imaging microscopy, stress concentrations are visualized and adhesive and cohesive failure in the fracture zone is differentiated.

Selectively probing ferric ions in aqueous environments using protonated and neutral forms of 7-azaindole as a multiparametric chemosensor.

7-azaindole (7AI) dimer is a model molecule for DNA study and understanding the mutagenic behavior based on the excited-state proton transfer process in hydrogen-bonded networks. The neutral and protonated forms of 7AI monomer with significant fluorescence (FL) intensity fit the fluorescent sensor strategy to recognize selective metal ions. Out of several metal ions (Fe3+, Al3+, Fe2+, Pb2+, Ba2+, Ni2+, Zn2+, Mg2+, Ca2+, Cu2+, Hg2+ and Cd2+), the absorption, fluorescence and fluorescence lifetime of 7AI in the aqueous medium are selectively sensitive to the ferric (Fe3+) ions. The absolute value of absorption intensity increases linearly with concentration of a particular metal ions. FL intensity of both the forms of 7AI decreases gradually with Fe3+ ions and trails the linear Stern-Volmer relation. The formation of non-fluorescent complexes was confirmed with Benesi-Hildebrand and Job plots, along with FL and FL decays. The FL lifetime of the protonated form of 7AI, which is 0.83 ± 0.01 ns, is nearly constant with Fe3+ ions concentrations, confirming the static quenching mechanism. The limit of detection (LoD) of Fe3+ ions over the long range of 16-363 µM for the neutral and protonated forms of 7AI is 0.46 ± 0.02 and 0.49 ± 0.02 µM, respectively, estimated using FL spectra. Additionally, the linear plot of absorbance with Fe3+ ions of both the forms of 7AI can also act as a calibration curve with very close LoDs, as obtained by FL spectra. Thus, the multi-parameters-based probe for detecting the Fe3+ ions over long-range in real aqueous environments with operational, high sensitivity, fast response (< 5 s), and good selectivity (over 12 metal ions) is undoubtedly a superior approach over other methods.

Contrast-Free FLIM Reveals Metabolic Changes in Pathological Islets of Langerhans.

FLIM (Fluorescence Lifetime Imaging Microscopy) is a powerful tool that could be used in the future to diagnose islet cell recovery after therapy. The identification of appropriate FLIM parameters is required to determine islet quality and islet cell metabolism throughout the organ under various conditions of insulin deficiency. The aim of the work was to identify key FLIM parameters, changes of which are characteristic of pancreatic pathologies. The τm, τ1, τ2, α1, α2 and α1/α2 of free and bound forms of NAD(P)H of the islet cells of animals (rats and pigs) and of humans with and without pathologies were measured and analyzed. The data were confirmed by IHC and histological studies. We identified three FLIM parameters in islet cells from animals with streptozotocin (STZ)-induced diabetes mellitus (DM) and from humans with chronic pancreatitis + type 2 diabetes (T2D), which differ in the same way: τm and α2 take lower values compared to the nonpathological islet cells, while α1/α2 takes higher values. In islet cells from patients with adenocarcinoma (PDAC) and chronic pancreatitis, these parameters had reverse tendency relative to the norm or did not differ. Thus, minimally invasive and non-contrast FLIM methods may, in the future, be used to diagnose pathological islet cells.

Use of Flavin-Related Cellular Autofluorescence to Monitor Processes in Microbial Biotechnology.

Cellular autofluorescence is usually considered to be a negative phenomenon because it can affect the sensitivity of fluorescence microscopic or flow cytometric assays by interfering with the signal of various fluorescent probes. Nevertheless, in our work, we adopted a different approach, and green autofluorescence induced by flavins was used as a tool to monitor fermentation employing the bacterium Cupriavidus necator. The autofluorescence was used to distinguish microbial cells from abiotic particles in flow cytometry assays, and it was also used for the determination of viability or metabolic characteristics of the microbial cells. The analyses using two complementary techniques, namely fluorescence microscopy and flow cytometry, are simple and do not require labor sample preparation. Flavins and their autofluorescence can also be used in a combination with other fluorophores when the need for multi-parametrical analyses arises, but it is wise to use dyes that do not emit a green light in order to not interfere with flavins' emission band (500-550 nm).

Study on Photophysical Properties of Novel Fluorescent Phenanthroimidazole-Thiadiazole Hybrid Derivatives.

Phenanthroimidazole-thiadiazole hybrid derivatives, which are new heterocyclic compounds with fluorescence properties, were synthesized by designing a two-step reaction mechanism and their photophysical properties were investigated. The synthesized derivatives were purified and their structures were elucidated by ATR-IR, 1H-NMR, elemental analysis and HR-MS methods. In the next stage of the study, the absorption and emission spectra of the synthesized new phenanthroimidazole-thiadiazole hybrid derivatives were determined by UV-Vis and fluorescence spectroscopy. Stokes' shifts, molar extinction coefficients (ε), singlet energy levels (Es), quantum yield (ϕf), lifetimes (τ), the radiative (kr) and the nonradiative (knr) constant for new hybrid derivatives were measured in DMSO. In addition, aggregation measurements, which is an important parameter that changes the photophysical properties were performed and for these compounds, structure: photophysical properties were discussed.

Electrostatically induced pKa shifts in oligopeptides: the upshot of neighboring side chains.

pKa values of homorepeat hexapeptides with a 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO) chromophore attached at the peptide C termini, through an asparagine derivative (Dbo), namely His6-Dbo (H6), Lys6-Dbo (K6), and Arg6-Dbo (R6), were determined by a novel fluorescence-based method. The fluorescence lifetime of Dbo in the peptides (τ) was measured as a function of pH. The side chains collide with Dbo intramolecularly and quench it efficiently only when they are deprotonated (i.e., pH ≥ side chain pKa). The pKa values of the H6, K6, and R6 peptides, attributable to side chain ionization, were found to be depressed compared to the pKa values of the His, Lys, and Arg residues in their free amino acid forms. We further looked into the structural changes of the peptides by molecular dynamics (MD) simulations; the peptides were structurally more expanded when their side chains are protonated. The structural expansion of the peptides reflects an electrostatic repulsion between the protonated side chain residues, which also accounts for the observed decrease in pKa values, which corresponds to a facilitated deprotonation, assisted by electrostatic repulsion.

Spectral and lifetime resolution of fundus autofluorescence in advanced age-related macular degeneration revealing different signal sources.

PURPOSE: To determine the fundus autofluorescence (FAF) lifetimes and spectral characteristics of individual drusen and hyperpigmentation independent of those with retinal pigment epithelium (RPE) in geographic atrophy (GA) areas in late-stage age-related macular degeneration (AMD). METHODS: Three consecutive patients with complete RPE and outer retinal atrophy (cRORA) exhibiting drusen that were calcified or associated with hyperpigmentation were investigated with multimodal non-invasive ophthalmic imaging including colour fundus photography (CFP), optical coherence tomography (OCT), near-infrared reflectance (NIR), blue FAF and fluorescence lifetime imaging ophthalmoscopy (FLIO). Fluorescence lifetimes were measured in two spectral channels (short-wavelength spectral channel (SSC): 500-560 nm and long-wavelength spectral channel (LSC): 560-720 nm). RESULTS: Drusen lacking RPE coverage, as confirmed by CFP and OCT, had longer FAF lifetimes than surrounding cRORA by 127 ± 66 ps (SSC) and 113 ± 48 ps (LSC, both p = 0.008 in Wilcoxon test, N = 9) and by 209 ± 100 ps (SSC) and 121 ± 56 ps (LSC, p < 0.001, N = 14) in two patients. Hyperpigmentation in CFP in a third patient shows strong FAF with prolonged lifetimes. In the SSC, persistent FAF was found inside cRORA. A crescent-shaped hyperfluorescence in an area of continuous RPE but lacking outer retina was seen in one eye with a history of anti-VEGF treatment. CONCLUSIONS: Short-wavelength fluorescence in cRORA points to fluorophores beyond RPE organelles. Fluorescence properties of drusen within cRORA differ from in vivo drusen covered by RPE. These limited findings from three patients give new insight into the sources of FAF that can be further elucidated in larger cohorts.

Multispectral LIF-Based Standoff Detection System for the Classification of CBE Hazards by Spectral and Temporal Features.

Laser-induced fluorescence (LIF) is a well-established technique for monitoring chemical processes and for the standoff detection of biological substances because of its simple technical implementation and high sensitivity. Frequently, standoff LIF spectra from large molecules and bio-agents are only slightly structured and a gain of deeper information, such as classification, let alone identification, might become challenging. Improving the LIF technology by recording spectral and additionally time-resolved fluorescence emission, a significant gain of information can be achieved. This work presents results from a LIF based detection system and an analysis of the influence of time-resolved data on the classification accuracy. A multi-wavelength sub-nanosecond laser source is used to acquire spectral and time-resolved data from a standoff distance of 3.5 m. The data set contains data from seven different bacterial species and six types of oil. Classification is performed with a decision tree algorithm separately for spectral data, time-resolved data and the combination of both. The first findings show a valuable contribution of time-resolved fluorescence data to the classification of the investigated chemical and biological agents to their species level. Temporal and spectral data have been proven as partly complementary. The classification accuracy is increased from 86% for spectral data only to more than 92%.

Excited State Lifetimes of Sulfur-Substituted DNA and RNA Monomers Probed Using the Femtosecond Fluorescence Up-Conversion Technique.

Sulfur-substituted DNA and RNA nucleobase derivatives (a.k.a., thiobases) are an important family of biomolecules. They are used as prodrugs and as chemotherapeutic agents in medical settings, and as photocrosslinker molecules in structural-biology applications. Remarkably, excitation of thiobases with ultraviolet to near-visible light results in the population of long-lived and reactive triplet states on a time scale of hundreds of femtoseconds and with near-unity yields. This efficient nonradiative decay pathway explains the vanishingly small fluorescence yields reported for the thiobases and the scarcity of fluorescence lifetimes in the literature. In this study, we report fluorescence lifetimes for twelve thiobase derivatives, both in aqueous solution at physiological pH and in acetonitrile. Excitation is performed at 267 and 362 nm, while fluorescence emission is detected at 380, 425, 450, 525, or 532 nm. All the investigated thiobases reveal fluorescence lifetimes that decay in a few hundreds of femtoseconds and with magnitudes that depend and are sensitive to the position and degree of sulfur-atom substitution and on the solvent environment. Interestingly, however, three thiopyrimidine derivatives (i.e., 2-thiocytidine, 2-thiouridine, and 4-thiothymidine) also exhibit a small amplitude fluorescence component of a few picoseconds in aqueous solution. Furthermore, the N-glycosylation of thiobases to form DNA or RNA nucleoside analogues is demonstrated as affecting their fluorescence lifetimes. In aqueous solution, the fluorescence decay signals exciting at 267 nm are equal or slower than those collected exciting at 362 nm. In acetonitrile, however, the fluorescence decay signals recorded upon 267 nm excitation are, in all cases, faster than those measured exciting at 362 nm. A comparison to the literature values show that, while both the DNA and RNA nucleobase and thiobase derivatives exhibit sub-picosecond fluorescence lifetimes, the 1ππ* excited-state population in the nucleobase monomers primarily decay back to the ground state, whereas it predominantly populates long-lived and reactive triplet states in thiobase monomers.

Fluorescence Lifetime Patterns of Retinal Pigment Epithelium Atrophy in Patients with Stargardt Disease and Age-Related Macular Degeneration.

PURPOSE: To investigate whether autofluorescence lifetime patterns within retinal pigment epithelium (RPE) atrophy differ between age-related macular degeneration (AMD) and Stargardt disease (STGD). METHODS: Mean retinal autofluorescence lifetimes were measured in a short and a long spectral channel (SSC: 498-560 nm; LSC: 560-720 nm). Mean retinal fluorescence lifetimes were analyzed with corresponding clinical features, fundus images, fundus autofluorescence intensity images, and optical coherence tomography. Mean fluorescence lifetime values of atrophic areas were compared between the two cohorts and within the same patient to adjacent nonatrophic regions. RESULTS: Mean fluorescence lifetimes within areas with RPE atrophy of 13 patients with STGD (mean age ± SEM 43.7 ± 5 years) and 30 patients with geographic atrophy (mean age: 78 ± 2 years) were analyzed and compared to age-matched healthy participants. The mean area of RPE atrophy in STGD and AMD was 6.6 ± 2.3 mm2 (range: 0.66-33.17 mm2) and 17.5 ± 3.8 mm2 (range: 0.58-50.02 mm2), respectively. In patients with AMD, atrophic areas revealed significantly longer mean fluorescence lifetime values as compared with patients with STGD (SSC: 997 ± 60 vs. 363 ± 26 ps; LSC: 880 ± 46 vs. 393 ± 23 ps; p < 0.0001). CONCLUSIONS: This study established that RPE atrophy in patients secondary to STGD and AMD display distinctive mean fluorescence lifetime characteristics. As retinal fluorescence lifetimes within areas of RPE atrophy were significantly longer in AMD patients, the analysis of specific lifetime patterns may provide additional insight into the disease processes and the pathogenetic mechanisms in the development of atrophic patches in AMD and STGD.

Fast, Infrared-Active Optical Transistors Based on Dye-Sensitized CdSe Nanocrystals.

We report an optically gated transistor composed of CdSe nanocrystals (NCs), sensitized with the dye zinc ß-tetraaminophthalocyanine for operation in the first telecom window. This device shows a high ON/OFF ratio of 6 orders of magnitude in the red spectral region and an unprecedented 4.5 orders of magnitude at 847 nm. By transient absorption spectroscopy, we reveal that this unexpected infrared sensitivity is due to electron transfer from the dye to the CdSe NCs within 5 ps. We show by time-resolved photocurrent measurements that this enables fast rise times during near-infrared optical gating of 47 ± 11 ns. Electronic coupling and accelerated nonradiative recombination of charge carriers at the interface between the dye and the CdSe NCs are further corroborated by steady-state and time-resolved photoluminescence measurements. Field-effect transistor measurements indicate that the increase in photocurrent upon laser illumination is mainly due to the increase in the carrier concentration while the mobility remains unchanged. Our results illustrate that organic dyes as ligands for NCs invoke new optoelectronic functionalities, such as fast optical gating at sub-bandgap optical excitation energies.

Understanding Local-Field Correction Factors in the Framework of the Onsager-Böttcher Model.

The determination of the appropriate local-field factor for quantifying the response of a molecule to an external electric field is of major importance in optical spectroscopy. Although numerous studies have dealt with the evolution of the optical properties of emitters as a function of their environment, the choice of the model used to quantify local fields is still ambiguous, and sometimes even arbitrary. In this paper, we review the Onsager-Böttcher model, which introduces the polarizability of the probe molecule as the determinant parameter for the local field factor, and we establish a simple conceptual framework encompassing all commonly used models. Finally, a discussion of published experimental research illustrates the potential of the measurement of local electric fields in dense dielectric media, as well as the subtleties involved in their interpretation.

Chemical cross-linking of a variety of green fluorescent proteins as Förster resonance energy transfer donors for Yukon orange fluorescent protein: A project-based undergraduate laboratory experience.

Förster resonance energy transfer (FRET) is the basis for many techniques used in biomedical research. Due to its wide use in molecular sensing, FRET is commonly introduced in many biology, chemistry, and physics courses. While FRET is of great importance in the biophysical sciences, the complexity and difficulty of constructing FRET experiments has resulted in limited usage in undergraduate laboratory settings. Here, we present a practical undergraduate laboratory experiment for teaching FRET using a diverse set of green-emitting fluorescent proteins (FPs) as donors for a cross-linked Yukon orange FP. This laboratory experiment enables students to make the connection of basic lab procedures to real world applications and can be applied to molecular biology, biochemistry, physical chemistry, and biophysical laboratory courses. Published 2018. This article is a U.S. Government work and is in the public domain in the USA., 46(5):516-522, 2018.

Improving Quantum Yield of Upconverting Nanoparticles in Aqueous Media via Emission Sensitization.

We demonstrate a facile method to improve upconversion quantum yields in Yb,Er-based nanoparticles via emission dye-sensitization. Using the commercially available dye ATTO 542, chosen for its high radiative rate and significant spectral overlap with the green emission of Er3+, we decorate the surfaces of sub-25 nm hexagonal-phase Na(Y/Gd/Lu)0.8F4:Yb0.18Er0.02 upconverting nanoparticles with varying dye concentrations. Upconversion photoluminescence and absorption spectroscopy provide experimental confirmation of energy transfer to and emission from the dye molecules. Upconversion quantum yield is observed to increase with dye sensitization, with the highest enhancement measured for the smallest particles investigated (10.9 nm in diameter); specifically, these dye-decorated particles are more than 2× brighter than are unmodified, organic-soluble nanoparticles and more than 10× brighter than are water-soluble nanoparticles. We also observe 3× lifetime reductions with dye adsorption, confirming the quantum yield enhancement to result from the high radiative rate of the dye. The approach detailed in this work is widely implementable, renders the nanoparticles water-soluble, and most significantly improves sub-15 nm nanoparticles, making our method especially attractive for biological imaging applications.

Development and Evaluation of Lipid Nanoparticles Containing Natural Botanical Oil for Sun Protection: Characterization and in vitro and in vivo Human Skin Permeation and Toxicity.

The use of sunscreen products is widely promoted by schools, government agencies, and health-related organizations to minimize sunburn and skin damage. In this study, we developed stable solid lipid nanoparticles (SLNs) containing the chemical UV filter octyl methoxycinnamate (OMC). In parallel, we produced similar stable SLNs in which 20% of the OMC content was replaced by the botanical urucum oil. When these SLNs were applied to the skin of human volunteers, no changes in fluorescence lifetimes or redox ratios of the endogenous skin fluorophores were seen, suggesting that the formulations did not induce toxic responses in the skin. Ex vivo (skin diffusion) tests showed no significant penetration. In vitro studies showed that when 20% of the OMC was replaced by urucum oil, there was no reduction in skin protection factor (SPF), suggesting that a decrease in the amount of chemical filter may be a viable alternative for an effective sunscreen, in combination with an antioxidant-rich vegetable oil, such as urucum. There is a strong trend towards increasing safety of sun protection products through reduction in the use of chemical UV filters. This work supports this approach by producing formulations with lower concentrations of OMC, while maintaining the SPF. Further investigations of SPF in vivo are needed to assess the suitability of these formulations for human use.

Fluorescence lifetime imaging ophthalmoscopy.

Imaging techniques based on retinal autofluorescence have found broad applications in ophthalmology because they are extremely sensitive and noninvasive. Conventional fundus autofluorescence imaging measures fluorescence intensity of endogenous retinal fluorophores. It mainly derives its signal from lipofuscin at the level of the retinal pigment epithelium. Fundus autofluorescence, however, can not only be characterized by the spatial distribution of the fluorescence intensity or emission spectrum, but also by a characteristic fluorescence lifetime function. The fluorescence lifetime is the average amount of time a fluorophore remains in the excited state following excitation. Fluorescence lifetime imaging ophthalmoscopy (FLIO) is an emerging imaging modality for in vivo measurement of lifetimes of endogenous retinal fluorophores. Recent reports in this field have contributed to our understanding of the pathophysiology of various macular and retinal diseases. Within this review, the basic concept of fluorescence lifetime imaging is provided. It includes technical background information and correlation with in vitro measurements of individual retinal metabolites. In a second part, clinical applications of fluorescence lifetime imaging and fluorescence lifetime features of selected retinal diseases such as Stargardt disease, age-related macular degeneration, choroideremia, central serous chorioretinopathy, macular holes, diabetic retinopathy, and retinal artery occlusion are discussed. Potential areas of use for fluorescence lifetime imaging ophthalmoscopy will be outlined at the end of this review.

Synergistic effects of polymyxin and ionic liquids on lipid vesicle membrane stability and aggregation.

Ionic liquids (ILs) have been investigated for potential antibacterial and antibiotic applications due to their ability to destabilize and permeabilize the lipid bilayers in cell membranes. Bacterial assays have shown that combining ILs with antibiotics can provide a synergistic enhancement of their antibacterial activities. We have characterized the mechanism by which the conventional ILs 1-butyl-3-methylimidazolium chloride (BMICl) and 1-butyl-3-methylimidazolium tetrafluoroborate (BMIBF4) enhance the lipid membrane permeabilization of the well-known antibiotic polymyxin B (PMB). We studied the sizes and membrane permeabilities of multilamellar and unilamellar lipid bilayer vesicles in the presence of ILs alone in aqueous solution, PMB alone, and ILs combined together with PMB. Light scattering-based experiments show that vesicle sizes dramatically increase when ILs are combined with PMB, which suggests that the materials combine to synergistically enhance lipid membrane disruption leading to vesicle aggregation. Lipid bilayer leakage experiments using tris (2,2'-bipyridyl) ruthenium (II) (Ru(bpy)32+) trapped in lipid vesicles, in which the trapped Ru(bpy)32+ fluorescence lifetime increases when it leaks out of the vesicle, show that combining BMIBF4 and PMB together permeabilize the membrane significantly more than with PMB or the IL alone. This demonstrates that ILs can assist in antibiotic permeabilization of lipid bilayers which could explain the increased antibiotic activities in the presence of ILs in solution.