Fate of Photoinduced Electron Transfer Reactions with Temperature- and pH-Induced Assembly/Disassembly of Star Block Copolymer Micelles.

The assembly/disassembly of star block copolymers induced by changes in temperature or pH of the medium is anticipated to have interesting implications for hosting/releasing drugs and tuning chemical reactions. This study investigates the possibility of employing the dually sensitive self-assembly of an ethylene oxide-propylene oxide star block copolymer, Tetronic T904, to influence photoinduced electron transfer (ET) reactions, on switching from the assembled state (micelle) when temperature is above the critical micelle temperature (CMT) and pH of the medium is above the pKa of T904 to the dissociated (unimer) state when either the temperature is below the CMT or the polymer is protonated. Steady-state and time-resolved fluorescence techniques have been used to characterize the microenvironments of the reactants in T904 solutions under different temperature and pH conditions and to determine ET rate constants. Interestingly, the bimolecular ET rate constants in both assembled and disassembled states of T904 depict a bell-shaped correlation with the driving force of the reaction, in accordance with Marcus inversion behavior instead of the usual Rehm-Weller behavior seen in conventional solvents. The assembly/disassembly of T904 stimulated by temperature or pH affects the micropolarity in the reactant environment, the magnitude of ET rate constants, and the position of inversion on the exergonicity scale.

Fluorescence Coupling to Internal Modes of 1D Photonic Crystals Characterized by Back Focal Plane Imaging.

The coupling of fluorescence with surface electromagnetic modes, such as surface plasmons on thin metal films or Bloch surface waves (BSW) on truncated one-dimensional photonic crystals (1DPC), are presently utilized for many fluorescence-based applications. In addition to the surface wave, 1DPCs also support other electromagnetic modes that are confined within the 1DPC structure. These internal modes (IMs) have not received much attention for fluorescence coupling due to lack of spatial overlap of their electric fields with the surface bound fluorophores. However, our recent studies have indicated that the fluorescence coupling with IMs occurs quite efficiently. This observed internal mode-coupled emission (IMCE) is (similar to BSW-coupled emission) indeed wavelength dependent, directional and S-polarized. In this paper, we have carried out back-focal plane (BFP) imaging to reveal that the IMs of 1DPCs can couple with surface bound excited dye molecules, with or without a BSW mode presence. Depending on the emission wavelength, the coupling is observed with BSW and IMs or only IMs of the 1DPC structure. The experimental results are well matching with numerical simulations. The occurrence of IMCE regardless of the availability of BSWs removes the dependence on just the surface mode for obtaining coupled emission from 1DPCs. The observation of IMCE is expected to widen the scope of 1DPCs for surface-based fluorescence sensing and assays.

Directional Emission from Metal-Dielectric-Metal Structures: Effect of Mixed Metal Layers, Dye Location and Dielectric Thickness.

Metal-dielectric-metal (MDM) structures provide directional emission close to the surface normal, which offers opportunities for new design formats in fluorescence based applications. The directional emission arises due to near-field coupling of fluorophores with the optical modes present in the MDM substrate. Reflectivity simulations and dispersion diagrams provide a basic understanding of the mode profiles and the factors that affect the coupling efficiency and the spatial distribution of the coupled emission. This work reveals that the composition of the metal layers, the location of the dye in the MDM substrate and the dielectric thickness are important parameters that can be chosen to tune the color of the emission wavelength, the angle of observation, the angular divergence of the emission and the polarization of the emitted light. These features are valuable for displays and optical signage.

A study on the photophysics of 9-amino-10-cyanoanthracene: probing its dual absorption and emission behavior.

The photophysics of a donor-acceptor substituted chromophore, 9-amino-10-cyanoanthracene (ACAN), has been investigated in polar and nonpolar solvents to understand its intriguing dual absorption and emission behavior. Steady-state and time-resolved fluorescence studies clearly indicate that the short wavelength emission band of ACAN arises from the higher excited singlet state, S2, while the longer wavelength emission band arises from the intramolecular charge transfer (ICT) state, S1. Interestingly, both these states can be populated by direct excitation from the ground state. Temperature dependent studies reveal a pronounced activation controlled nonradiative decay channel for the ICT state of ACAN. It is proposed that this activation controlled nonradiative de-excitation arises because of a large relative displacement and a cross-over of the potential energy (PE) surfaces of ACAN in the ground and the ICT states, as a result of different twist angles of the amino group in these two states. Qualitative PE diagrams have accordingly been presented to correlate and rationalize the observed results. The present study also brings to light the interesting excited state prototropic behavior of ACAN and the consequent modulation of the ICT emission that has not been reported in the literature so far.

Stimuli-responsive supramolecular micellar assemblies of cetylpyridinium chloride with cucurbit[5/7]urils.

This article demonstrates, for the first time, construction of novel cucurbituril (CB)-adorned supramolecular micellar assemblies of a cationic surfactant, cetylpyridinium chloride (CPC), through noncovalent host-guest interactions. The distinct cation receptor features and cavity dimensions of the CB5 and CB7 homologues assert that the macrocyclic hosts remain complexed with the CPC monomers and take part in the micelle formation, a unique observation in contrast to that of the classical host, ß-cyclodextrin. The cooperative contributions of the CB macrocycles in the micelle formation have been documented by the photochemical, surface tension, conductivity, DOSY NMR, and SANS measurements. The contrasting downward and upward shifts in the cmc of the CPC surfactant, respectively, with CB5 and CB7 hosts provide a unique opportunity for the controlled tuning of the micellization region for CPC from 0.57 to 1.6 mM, by using a combination of the macrocyclic hosts. The article also establishes the reversible response of these soft supramolecular micellar structures to thermal-stimuli, which projects their utility for on-demand smart drug-delivery vehicles.

Tuning Fluorescence Direction with Plasmonic Metal-Dielectric- Metal Substrates.

Controlling the emission properties of fluorophores is essential for improving the performance of fluorescence-based techniques in modern biochemical research, medical diagnosis, and sensing. Fluorescence emission is isotropic in nature, which makes it difficult to capture more than a small fraction of the total emission. Metal- dielectric-metal (MDM) substrates, discussed in this Letter, convert isotropic fluorescence into beaming emission normal to the substrate. This improves fluorescence collection efficiency and also opens up new avenues for a wide range of fluorescence-based applications. We suggest that MDM substrates can be readily adapted for multiple uses, such as in microarray formats, for directional fluorescence studies of multiple probes or for molecule-specific sensing with a high degree of spatial control over the fluorescence emission. SECTION: Physical Processes in Nanomaterials and Nanostructures.

Silver-Gold Nanocomposite Substrates for Metal-Enhanced Fluorescence: Ensemble and Single-Molecule Spectroscopic Studies.

In recent years, there has been a growing interest in the studies involving the interactions of fluorophores with plasmonic nanostructures or nanoparticles. These interactions lead to several favorable effects such as increase in the fluorescence intensities, increased photostabilities, and reduced excited-state lifetimes that can be exploited to improve the capabilities of present fluorescence methodologies. In this regard, we report the use of newly developed silver-gold nanocomposite (Ag-Au-NC) structures as substrates for metal-enhanced fluorescence (MEF). The Ag-Au-NC substrates have been prepared by a one-step galvanic replacement reaction from thin silver films coated on glass slides. This approach is simple and suitable for the fabrication of MEF substrates with large area. We have observed about 15-fold enhancement in the fluorescence intensity of ATTO655 from ensemble fluorescence measurements using these substrates. The fluorescence enhancement on the Ag-Au-NC substrates is also accompanied by a reduction in the fluorescence lifetime of ATTO655, which is consistent with the fluorophore-plasmon coupling mechanism. Single-molecule fluorescence measurements have been performed to gain more insight into the metal-fluorophore interactions and to unravel the heterogeneity in the interaction of individual fluorophores with the fabricated substrates. The single-molecule studies are in good agreement with the ensemble measurements and show maximum enhancements of ~50-fold for molecules located in proximity to the "hotspots" on the substrates. In essence, the Ag-Au-NC substrates have a very good potential for various MEF applications.

Differential distribution of pigment-protein complexes in the Thylakoid membranes of Synechocystis 6803.

Thylakoids in Synechocystis 6803, though apparently uniform in appearance in ultrastructure, were found to consist of segments which were functionally dissimilar and had distinct proteomes. These thylakoid segments can be isolated from Synechocystis 6803 by successive ultracentrifugation of cell free extracts at 40,000×g (40 k segments), 90,000×g (90 k segments) and 150,000×g (150 k segments). Electron microscopy showed differences in their appearance. 40 k segments looked feathery and fluffy, whereas the 90 k and 150 k thylakoid membrane segments appeared tiny and less fluffy. The absorption spectra showed heterogeneous distribution of pigment-protein complexes in the three types of segments. The photochemical activities of Photosystem I (PSI) and Photosystem II (PSII) showed unequal distributions in 40 k, 90 k and 150 k segments which were substantiated with low temperature fluorescence measurements. The ratio of PSII/PSI fluorescence emission at 77 K (λ(ex) = 435 nm) was highest in 150 k segments indicating higher PSII per unit PSI in these segments. The chlorophyll fluorescence lifetimes in the membranes, determined with a time-correlated single-photon counting technique, could be resolved in three components: τ(1) (=) <40 ps, τ(2) (=) 425-900 ps and τ(3) (=) 2.4-3.2 ns. The percentage contribution of the fastest component (τ(1)) decreased in the order 40 k > 90 k > 150 k segments whereas that of the other two components showed a reversed trend. These studies indicated differential distribution of pigment-protein complexes in the three membrane segments suggesting heterogeneity in the thylakoids of Synechocystis 6803.

Early detection of insulin fibrillation: a fluorescence lifetime assay to probe the pre-fibrillar regime.

Using human insulin, we report for the first time, the evaluation of fluorescence lifetime of an extrinsically added fluorogenic dye, Thioflavin T (ThT), as a more sensitive and convenient method for the assessment of fibrillation in the pre-fibrillar regime.

Silver Nanostructures for Fluorescence Correlation Spectroscopy: Reduced Volumes and Increased Signal Intensities.

Fluorescence correlation spectroscopy (FCS) is a widely used technique to investigate the interactions and dynamics of molecules, below micromolar concentrations. Silver nanostructure (AgNS) substrates can extend the applicability of FCS to higher concentrations, which is useful for many biologically relevant reactions. Additionally, these substrates can improve detection efficiency by increasing fluorescence signal intensities. The ease of preparation of the AgNS substrates in comparison to previously investigated materials prepared by top-down nanofabrication is expected to make them readily available and suitable for various FCS applications.

Fluorescence enhancement using silver-gold nanocomposite substrates.

Metal-enhanced fluorescence (MEF) is a newly emerging phenomenon in which the near-field interactions of fluorophores with the plasmons in metallic nanostructures can lead to substantial fluorescence enhancements. In the present study, we have investigated the use of silver-gold nanocomposite (Ag-Au-NC) structures, prepared by the galvanic replacement reaction of silver with gold, as plasmonic substrates for MEF. We have observed significant enhancement in the fluorescence intensities and decrease in the fluorescence lifetimes of two commonly used dyes, ATTO655 and Cy5, using the fabricated Ag-Au-NC substrates. Interestingly, the fluorescence enhancement depends on the amount of residual silver present in the substrates after the galvanic replacement reaction. Our results show that the galvanic replacement reaction is a very facile and powerful route to prepare Ag-Au-NC substrates that can be suitable for various MEF based applications.

Recognition-mediated light-up of thiazole orange with cucurbit[8]uril: exchange and release by chemical stimuli.

This article reports a convenient supramolecular strategy to construct fluorescent photoswitchable molecular assemblies between a macrocyclic host, cucurbit[8]uril (CB8), and a fluorogenic dye, thiazole orange (TO). The interaction mechanism and the stable stoichiometric host-guest arrangements have been claimed on the basis of the optical absorption, steady-state and time-resolved fluorescence lifetime and anisotropy measurements, and also the geometry optimization studies. The CB8 recognized TO in its 2:2 stoichiometry exhibited spectacular fluorescence enhancement of the order of 1700 fold, which is the largest directly determined value so far reported for a dye in an organic macrocyclic system. This prospective 2CB8:2TO assembly responded to selected chemical stimuli such as metal ions, adamantylamine, and tryptophan, providing different dissociation mechanisms and demonstrating a controlled exchange and release action desired with such noncovalently linked assemblies. Positively, considering the aqueous solubility and biocompatibility of the host-guest constituents, this methodology can evolve into a general approach to deliver and operate intracellularly functional molecular components under chemical/thermal/optical trigger control, especially for therapeutic applications.

Surfactant-induced aggregation patterns of thiazole orange: a photophysical study.

The aggregation behavior of the DNA marker dye thiazole orange (TO), has been investigated in two types of surfactant assemblies, namely, premicelles/micelles of sodium dodecyl sulfate (SDS) and pre reverse micelles/reverse micelles of sodium bis(2-ethylhexyl) sulfosuccinate (AOT). In the case of an SDS/water system, absorption spectral changes of TO signify the formation of H-aggregates and H-dimers of the dye at premicellar concentrations, which subsequently convert to the monomeric form beyond the critical micellar concentration (cmc). Interestingly, the observed changes in the absorption and emission characteristics due to the surfactant-induced formation of H-aggregates/dimers of TO are found to be useful to estimate the surfactant concentration parameters for premicellar aggregation of SDS. In the case of an AOT/n-heptane system, similarly, H-aggregates/dimers are observed at low AOT concentrations, below the cmc. However, in this case, the H-dimers persist even beyond the cmc. This is attributed to the strong tendency of TO for self-aggregation and its favorable electrostatic interactions with the AOT head groups. With increasing water content in the AOT reverse micelles, the hydration of the dye leads to the conversion of H-dimers to the monomeric form. The steady-state fluorescence results are nicely corroborated with those from time-resolved fluorescence studies and demonstrate the interesting behavior of the surfactant-induced aggregation of TO dye.

Surface functionalized silver nanoparticle conjugates: demonstration of uptake and release of a phototherapeutic porphyrin dye.

A facile supramolecular approach to prepare surface functionalized silver nanoparticle conjugates has been established and their enhanced molecular recognition features have been explored to demonstrate the uptake and stimulus responsive release of a phototherapeutic porphyrin dye, the TMPyP.

Contrasting guest binding interaction of cucurbit[7-8]urils with neutral red dye: controlled exchange of multiple guests.

Interactions among macrocyclic hosts and dyes/drugs have been explored extensively for their direct usage in controlled uptake and release of large number of potential drug molecules. In this paper we report the non-covalent interaction of cucurbit[8]uril macrocycle (CB8) with a biologically important dye, neutral red, by absorption and fluorescence spectroscopy. A comparative analysis with the complexation behaviour of the dye with CB7, the lower homologue of CB8, indicates contrasting guest binding behaviour with significant changes in the photophysical characteristics of the dye. While CB7 interaction leads to a 1 ratio 1 stoichiometry resulting in approximately 6 fold enhancement in the fluorescence emission of the dye, CB8 displays signatures for a 1 ratio 2 host-guest stoichiometry with drastic reduction in the fluorescence emission. Apart from the evaluation of approximately 2 unit shift in the protolytic equilibrium on complexation (pK(a) shift), the measurements with tryptophan established a selective guest exchange to favour a co-localized dimer inside the CB8 cavity. In a protein medium (BSA), the 1 ratio 2 complex was converted to a 1 ratio 1 ratio 1 CB8-NRH(+)-BSA complex. The finding that NRH(+) can be transferred from CB8 to BSA, even though the binding constant for NRH(+)-CB8 is much higher than NRH(+)-BSA, is projected for a controlled slow release of NRH(+) towards BSA. Since the release and activity of drugs can be controlled by regulating the protolytic equilibrium, the macromolecular encapsulation and release of NRH(+) demonstrated here provide information relevant to host-guest based drug delivery systems and its applications.

Inhibiting intramolecular electron transfer in flavin adenine dinucleotide by host-guest interaction: a fluorescence study.

Modulation in the photophysical properties and intramolecular electron transfer behavior of the flavin adenine dinucleotide (FAD) molecule has been investigated in the presence of the macrocyclic hosts, alpha-, beta- and gamma-cyclodextrins (CDs), using absorption and steady-state and time-resolved fluorescence measurements. The results demonstrate that only the beta-CD host has a suitable cavity dimension to form a weak inclusion complex with FAD by encapsulating the adenine moiety, which is the preferred binding site in the large FAD molecule. Interestingly, in spite of the weak binding interaction, a significant enhancement in the fluorescence intensity of FAD is observed on complexation with beta-CD, and this has been attributed mainly to the modulation in the conformational dynamics of FAD in the presence of beta-CD. In aqueous solutions, a good fraction of FAD molecules exist in a "closed" conformation with the adenine and isoalloxazine rings stacked on each other, thus leading to very efficient fluorescence quenching due to the ultrafast intramolecular electron transfer from adenine to the isoalloxazine moiety. Complex formation with beta-CD inhibits this intramolecular electron transfer by changing the "closed" conformation of FAD to the "open" form, wherein the adenine and isoalloxazine moieties are widely separated, thus prohibiting the fluorescence quenching process. Further evidence for the conformational changes has been obtained by the observation of a long lifetime component in the fluorescence decay of FAD in the presence of beta-CD, which corresponds to the decay of the unquenched "open" form of FAD. Fluorescence up-conversion studies also indicate the absence of any ultrafast component in the fluorescence decay arising from the complexed FAD, thus supporting the formation of the "open" form in the presence of beta-CD, with no intramolecular electron transfer.

Cooperative metal ion binding to a cucurbit[7]uril-thioflavin T complex: demonstration of a stimulus-responsive fluorescent supramolecular capsule.

We report an intriguing noncovalent interaction of thioflavin T (ThT), a fibril diagnostic dye, with the versatile macrocyclic host molecule cucurbit[7]uril (CB7) in the presence of metal cations. ThT forms both 1:1 (CB7.ThT) and 2:1 [(CB7)(2).ThT] complexes with CB7 host, leading to specific structural arrangements. Addition of competitive guests like metal cations to the 1:1 stoichiometric complex displays expected competitive binding interactions with CB7, leading to decreased fluorescence intensity from ThT. However, addition of metal ions to the 2:1 complex leads to unusual enhancement in the fluorescence emission ( approximately 270-fold in the presence of Ca(2+) and approximately 160-fold in the presence of Na(+)). These contrasting observations on the fluorescence enhancement with change in the stoichiometric equilibrium have been investigated explicitly for a feasible binding model. Detailed photophysical characterization with supporting data from NMR and anisotropy measurements has led to the revelation of a novel stimulus-responsive cooperative metal ion binding to the stoichiometrically selected (CB7)(2).ThT complex, demonstrating a highly fluorescent supramolecular nanocapsule. The first example of a noncovalently packed fluorescent complex became feasible due to the structural arrangement of the host-guest complex in the 2:1 stoichiometry with two CB7 portals providing strong negative charge density for the metal ions to group and seal the complex, thus protecting the incorporated dye. To further strengthen the usefulness of the supramolecular capsule established here, rupture of the capsular complex has been demonstrated with a strong competitive guest, 1-amantadine hydrochloride, which helped in disrupting the capsule to release the dye. It is proposed here that by judicious design of the chromophore (guest) structure, such capsular assemblies can be explored for the binding and release of drug molecules, for fluorescence on-off systems, and as building blocks for molecular architectures displaying unique properties.

Control of the supramolecular excimer formation of thioflavin T within a cucurbit[8]uril host: a fluorescence on/off mechanism.

On or off? A new excimer band at lambda = 570 nm was visualized during the noncovalent host-guest interaction between thioflavin T (ThT) and cucurbit[8]uril (CB8). Controlled dissociation of this assembly in the presence of Ca(2+) was demonstrated as an on/off fluorescence switch (see picture).

Modulation of excited-state proton-transfer reactions of 7-hydroxy-4-methylcoumarin in ionic and nonionic reverse micelles.

The prototropic behavior of the dye, 7-hydroxy-4-methylcoumarin (7H4MC), has been studied in cationic benzyldimethylhexadecylammonium chloride (BDHC) and nonionic poly(oxyethylene)(tetramethylbutyl)phenyl ether (TritonX-100, TX-100) reverse micelles using ground-state absorption and steady-state and time-resolved fluorescence measurements. The results have been compared with the previous results in the anionic sodium 1,4-bis(2-ethylhexyl)sulfosuccinate (AOT) reverse micelles. Although the probe dye, 7H4MC, is indicated to reside in the interfacial region in all of the reverse micelles studied, significant differences have been observed in the evolution of the different prototropic species. In BDHC reverse micelles, the anionic form is favored over the tautomeric form at the higher w0 values, which is contrary to the observation in AOT reverse micelles where both of these forms are almost equally produced. The higher propensity for the formation of the anionic form in BDHC reverse micelles has been explained on the basis of the additional electrostatic stabilization of the anionic species in the cationic BDHC reverse micelles compared to that in the anionic AOT reverse micelles. On the other hand, in TX-100 reverse micelles, the anionic form is not very evident, but interestingly, the tautomer form begins to appear beyond w0=2. The appearance of the tautomeric species apparently coincides with the formation of the water pool in the TX-100 reverse micelles. However, due to the more restricted nature of the water molecules within this reverse micelle (mostly dispersed around the oxyethylene chains), deprotonation of the 7H4MC dye and the consequential stabilization of the anionic form are considerably reduced. The results clearly reveal that the aqueous environment in the vicinity of the probe is quite different for the reverse micelles considered, and these differences largely modulate the prototropic processes of the excited dye.

Host-guest interaction of 1,4-dihydroxy-9,10-anthraquinone (quinizarin) with cyclodextrins.

The interaction of 1,4-dihydroxy-9,10-anthraquinone, (quinizarin; QZ), with alpha-, beta- and gamma-cyclodextrin (CD) hosts, has been investigated using absorption, and steady-state and time-resolved fluorescence measurements, in order to understand the effects of cavity size of CDs on the binding of QZ molecule and the changes in the photophysical properties of QZ in the microenvironment of the hosts. The results demonstrate that QZ forms inclusion complexes with all the CDs. The low binding constants as well as the thermodynamic parameters indicate that the mode of interaction between QZ and CDs is mainly hydrophobic in nature. The relative magnitudes of the binding constants and the differential enhancements in the fluorescence intensity of QZ upon complexation with the CDs can be explained by considering the relative dimensions of the host cavity and the guest molecule, as well as the orientation of the guest molecule inside the CD cavity. It is proposed that the unsubstituted benzene ring of QZ is encapsulated within alpha- and beta-CD cavities whereas the dihydroxy-substituted aromatic ring is encapsulated within the gamma-CD cavity. This is further supported by the complexation studies of the QZ.CD systems with Al(III) ions. It is observed that the complexation of QZ with the metal ion is enhanced in the QZ.alpha-CD and QZ.beta-CD systems whereas it is significantly reduced in the QZ.gamma-CD system, due to shielding of the chelating groups of the dye inside the CD cavity in the latter case.