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A combination of fluorescence spectroscopy and quantum chemical calculations is used to study the extent of fluorescence enhancement of an epicocconone in different surfactant assemblies, which are found to be very different from each other. The analysis reveals that the depth-dependent microviscosity highly affects the fluorescence quantum that further proves the involvement of fluorophore as a dual emitting marker for fluorescence microscopy.

The effect of microenvironment of a Nafion membrane on the excited-state proton transfer (ESPT) of 2-(2'-pyridyl)benzimidazole (2PBI) was investigated by steady-state fluorescence spectroscopy. The results suggest that mechanism of ESPT depends on the water content of the membrane, but is slowed down by cation exchange due to dehydration of the membrane, the less acidic environment around 2PBI molecules, and slowing down of the proton-transfer process.

The aggregation/deaggregation of chlorin p(super 6) with the surfactants CTAB, SDS, and TX 100 are studied using absorption, florescence, and light scattering techniques. Surfactants with ionic as well as nonionic headgroups are found to be capable of inducing aggregation of chlorin p6 at submicellar concentrations.

Excited-state proton transfer (ESPT) of 2-(2'-pyridyl)benzimidazole (2PBI) in reverse micelles is studied by steady-state and time-resolved fluorescence spectroscopy. The requirement of ESPT to occur in microheterogenous media is that the medium should be a ternary system comprised of water and a hydrophobic phase separated by a negatively charged interface.

Picosecond time-resolved Stokes shift is used in studying solvation of 4-aminophthalimide (4-AP) in water-in-oil microemulsion of Triton X-100 in mixed solvents with 30% benzene and 70% n-hexane. Triton is a neutral surfactant which does not use sodium ions in the solvation of microemulsions. Results show the steady-state spectrum of 4-AP in microemulsion is very different from that in TX reverse micelle or in water. This suggests that 4-AP molecules stay in the water pool and the micropolarity of the water pool is different from ordinary water.

A study was conducted to analyze solvation dynamics in coumarin 480 in dimyristoylphosphatidylcholine vesicles in water using picosecond time-resolved Stokes shift. The response time of the experimental setup was about 50 ps while the wavelength of excitation for the time-resolved studies was 300 nm. Experimental results indicated that coumarin 480 is a sensitive fluorescent probe the microenvironment of the vesicles.

The specific binding of two model drugs for photodynamic therapy, namely chlorin [p.sub.6] and purpurin 18 in the vicinity of Sudlow's Site I of HSA is examined by monitoring the intrinsic fluorescence of single tryptophanyl residue and by competitive binding with warfarin. Time-resolved fluorescence results of competitive binding with warfarin have confirmed that they bind to the warfarin binding site.

The binding of serum albumin and lipoprotein with chlorin p6 and purpurin 18, two structurally related chlorins, is studied to understand the role for these proteins as endogenous carriers for these drugs. Low-density lipoproteins (LDLs) are found to be the preferred carrier for both the compounds, particularly at a slightly acidic pH, resembling the situation in tumor cells.

The effect of micellar environment on the excited state p5roton transfer (ESPT) of 2-(2'-pyridyl )benzimidazole (2PBI) is investigated by steady state and time resolved fluorescence spectroscopy. Three factors namely diffusion controlled protonation of the normal form of 2PBI, slow and possibly incomplete solvation of the transition state, leading to a slowing down of the proton transfer process and a similar slow dynamics of the tautomeric excited state is explored.

Effectsof annealing have been studied on the chemical structure,water uptake, and acidity of the cast Nafion thin film of thickness∼ 6 μm using a fluorescent probe, 2-(3′-pyridyl)benzimidazole(3PBI), and attenuated total reflactance infrared (ATR-IR) spectroscopy.Nonannealed films and thick Nafion 117 membrane have been used asreference materials, in order to develop a complete understandingof the effect. Annealing has been found to cause a decrease in acidityof otherwise highly acidic ionomer, as sensed by the fluorescent probeand loss of water, as reflected in the ATR-IR spectrum. This observationis surprising and cannot be explained in the light of previous reportsof physical changes. Our ATR-IR study has revealed changes in thechemical structure of the hydrophilic part of the ionomer, leadingto the formation of sulfonic acid anhydrides. This phenomenon canrationalize the decreased acidity reported in our fluorescence study.Interestingly, acid treatment of the annealed film restores the acidityof the unannealed films. This cannot be rationalized simply by a greaterproton uptake from the solution, as the film has to be electroneutral.It appears that the anhydrides formed during the annealing processundergo acid hydrolysis, leading to an increase in the number of SO3–groups and, consequently, an increasein the number of H3O+ions in the water channels.Besides, the films can be hydrated to an extent that is much greaterthan Nafion membranes, but the water uptake of acid treated annealedfilm is slightly less than that of nonannealed films. Hence, we concludethat, along with annealing, acid treatment, a procedure that is generallynot performed on the cast films, is an important pretreatment procedureto improve the acidity and hence the transport properties of the castfilm. [ABSTRACT FROM AUTHOR]

The conductivity of polyaniline (PANI)is ascribed to its emeraldinesalt (PANI-ES), which is formed by protonation of its emeraldine base(PANI-EB) by acids. Generally, mineral acids are used for this purpose,but the use of dopants and additives to maintain the required acidityprovides an alternative method to the preparation of PANI-ES. Thepresent work attempts to achieve the protonation by the use of a weakorganic acid, namely, 6-cyano-2-naphthol (6CN2), which is generallyused as a superphotoacid, as its excited state pKais significantly smaller than its ground state pKa. The question here is if the protonation ofthe aniline moieties in PANI takes place and if it does, whether ittakes place by dissociation of the ground state or the excited stateof 6CN2. Room temperature conductance measurements were carried outto see the effect of doping. The formation of PANI-ES from PANI-EBhas been monitored by UV–vis spectrophotometry. When a polarcounterion is inserted into the polymer matrix, it changes the environmentof the nearby chains by introducing defects, reorganization of chargesas a result of interaction with the polymer. Morphological investigationwas done using optical microscopy, field emission gun scanning electronmicroscopy (FEGSEM), and field emission gun transmission electronmicroscopy FEGTEM. The influence of 6CN2 on the crystallinity of thepolymer was determined by X-ray diffraction (XRD). [ABSTRACT FROM AUTHOR]

2-(4′-Pyridyl)benzimidazole (4PBI) has been usedto investigate the microheterogeneity of water nanochannels of Nafionmembranes at two different hydration levels. Native as well as cation-exchangedNafion membranes are found to protonate one of the two monoprotonatedforms of 4PBI selectively. In native membranes and in those in whichH3O+ions are replaced by (CH3)4N+ions, the pyridyl nitrogen is protonated preferentially.In Na+-exchanged membranes, however, the benzimidazolenitrogen is protonated selectively. Unlike other fluorescent probesused in earlier studies, 4PBI can differentiate between the two differentcation-exchanged membranes at lower as well as higher hydration levels. [ABSTRACT FROM AUTHOR]

(2,2′-Bipyridyl)-3-3′-diol (BP(OH)2) undergoesexcited state intramolecular double proton transfer (ESIDPT). Thephotophysics of this molecule has been investigated, in the highlyacidic water channels inside nafion membrane. The dianionic enolateform of the dye, which is formed only in alkaline conditions in neataqueous solutions, is found to be present in its excited state innative nafion membrane at low water content. This surprising phenomenonhas been explained in the light of loss of protons from the fluorophoreto the medium, due to increased electrostatic interactions with thependant sulfonate groups of nafion, at the lower water content. Thisobservation fortifies the model of microscopic interactions that wehave proposed in recent past. Further, the ground state of the diketoform, observed only in neat aqueous solutions so far, is found tobe present in Na+exchanged membranes at lower levels ofhydration. The steady-state spectral features of the dye is differentin (CH3)4N+exchanged membranes thanthat of Na+exchanged membranes, unlike in our earlierstudies with coumarin 102 and 2-(2′-pyridyl)benzimidazole,here we observed formation of an unusual ground state in Na+exchanged membranes, while no such feature was observed in (CH3)4N+exchanged membranes. [ABSTRACT FROM AUTHOR]

A molecular level understanding of the mobility of cations in the Nafion membrane has been attempted, using the excited state proton transfer (ESPT) process in the fluorescent probe Coumarin 102. ESPT is hindered significantly upon decreasing the water content. Using TRANES (time-resolved area normalized emission spectroscopy), the evolution of the ESPT state is clearly observed over hundreds of picoseconds in lower water content, implying that ESPT is hindered even in the nanovolume probed by the dye. Most remarkably, in the partially dried membrane, the predominant fluorescent species is the zwitterionic form, generated by excited state deprotonation of the cationic form. This implies that the molecule loses a proton from its nitrogen center in the excited state, as usual, but cannot recapture it readily at the oxygen center, at low water contents. This phenomenon is rationalized in light of an increased electrostatic attraction that is experienced by cations upon drying. [ABSTRACT FROM AUTHOR]

We have assessed the potential of salophen, a tetradentate Schiff base, in fluorescence sensing of Al3+ ions. While performing this investigation, we have noticed conflicting literature reports on the fluorescence spectral maximum and quantum yield of salophen. So, the compound has been purified by repeated crystallization. Fluorescence studies have been performed on samples in which the absorption and excitation spectra are completely superimposable. The purified compound exhibits a feeble fluorescence at 545 nm, associated with an ultrafast fluorescence decay. This is rationalized by excited state proton transfer and torsional motions within the molecule, which provide efficient nonradiative channels of deactivation of its excited state. The fluorescence quantum yield increases upon complexation of salophen with Zn2+ as well as Al3+. The increase is significantly more upon complexation with Al3+. However, fluorescence maxima are similar for the two complexes. This indicates that fluorescence intensity may not be a good parameter for Al3+ sensing by salophen, in the presence of a large excess of Zn2+. This problem can be circumvented if fluorescence lifetime is used as the sensing parameter, as the lifetime of the Al3+ complex is in the nanosecond time regime while that of the Zn2+ complex is in tens of picoseconds. The significant difference in the fluorescence quantum yield and lifetime between the two complexes is explained as follows: the Al3+ complex is monomeric, but the Zn2+ complex is dimeric. Quantum chemical calculations indicate a higher density of states near the locally excited state for the dimeric complex. This may lead to more efficient nonradiative pathways. [ABSTRACT FROM AUTHOR]

Dimethyl-2,5-bis(4-methoxyphenylamino)terephthalate (DBMPT) is a water-insoluble fluorogenic molecule, which has been rendered water-soluble in physiological conditions, by the addition of triblock copolymers (TBPs), P123 (PEO 19 PPO 69 PEO 19 ), and F127 (PEO 100 PPO 65 PEO 100 ). DBMPT-TBP mixed aggregates, formed in the process, exhibit significant aggregation-induced enhancement of emission, with nanosecond fluorescence lifetimes. Dynamics involved in suppression of nonradiative pathways and consequent enhancement of fluorescence are followed by femtosecond transient absorption and time-resolved fluorescence spectroscopic techniques. Interestingly, shapes of the aggregates formed with the two TBPs are found to be very different, even though they differ only in the length of hydrophilic blocks. DBMPT-P123 aggregates are micrometer-sized and spherical, while DBMPT-F127 aggregates form nanorods. Evolution of their morphologies, as a function of TBP concentration, is monitored using cryo-TEM, FESEM, and fluorescence lifetime imaging microscopy. Fluorescence lifetime distribution provides useful insight into microheterogeneity in these mixed aggregates. Excellent cell permeability is observed for DBMPT-F127 nanorods, in contrast to DBMPT-P123 microspheres. These fluorescent nanorods exhibit the ability to mark lipid droplets within the cell and hence bear the promise for application in intracellular imaging.

Dynamics of preferential solvation of 5-aminoquinoline (5AQ) has been studied in hexane-alcohol solvent mixtures. Significant spectral red shift and fluorescence quenching are observed, even for miniscule amounts of alcohols. A nonlinear variation of these parameters, with alcohol mole fraction, indicates dipolar enrichment around 5AQ in its excited state. A nanosecond rise at the red end of emission spectra is ascribed to translational diffusion of alcohol molecules and consequent preferential solvation of 5AQ. Electrostatic stabilization and hydrogen bonding between the dipolar excited state of the fluorophore and polar alcohol molecules are major driving factors here. Time evolution of emission spectra, from those in a predominantly hexane-like environment to those in a predominantly alcohol-like environment, takes place in nanosecond time scale and depends on the alcohol concentration in the mixture. This in agreement with calculations using an existing model. Hence, 5AQ is found to be a suitable fluorescent probe for the study of dynamics in complex systems.

The role of the amino group in the excited-state dynamics of 3-aminoquinoline (3AQ) has been investigated by comparison with its synthetic derivative 3-(piperidin-1-yl)quinoline (3PQ). The absence of amino hydrogen atoms in 3PQ eliminates, to a large extent, the complexity of the excited-state processes observed in 3AQ. The polarity of the medium is found to be the most important determinant in the nonradiative rate constants of 3PQ, unlike in 3AQ where hydrogen bonding plays the most significant role. The nonradiative rate constants decrease with increase in micropolarity. This trend is opposite to what is usually observed with dipolar states. Temperature dependence of the fluorescence spectra and lifetime has been studied to understand this unexpected observation. An unusual redshift in the emission of 3AQ and 3PQ is observed in nonpolar media at low temperatures. This is surprising, as a process involving a barrier is expected to be hindered at low temperatures and be manifested in a blueshift of the spectra, due to the predominance of the locally excited (LE) state. Moreover, the variation of emission maxima of 3AQ with temperature is sigmoidal in nature, indicating the involvement of two distinct states. The counterintuitive observation of the predominance of the state with comparatively lower emission energy, at low temperatures, establishes the following: the photophysics in 3AQ is dominated by a LE state at room temperature in nonpolar media. This state is associated with rapid flip-flop of the amino group, which provides an efficient nonradiative channel of deactivation. At low temperatures, this flip-flop is hindered and the molecule can undergo intramolecular charge transfer (ICT), whereby the lower energy state is populated. In the case of 3PQ, the ICT state is the only one present, owing to the tertiary amino group.

We have assessed the potential of salophen, a tetradentate Schiff base, in fluorescence sensing of Al 3+ ions. While performing this investigation, we have noticed conflicting literature reports on the fluorescence spectral maximum and quantum yield of salophen. So, the compound has been purified by repeated crystallization. Fluorescence studies have been performed on samples in which the absorption and excitation spectra are completely superimposable. The purified compound exhibits a feeble fluorescence at 545 nm, associated with an ultrafast fluorescence decay. This is rationalized by excited state proton transfer and torsional motions within the molecule, which provide efficient nonradiative channels of deactivation of its excited state. The fluorescence quantum yield increases upon complexation of salophen with Zn 2+ as well as Al 3+ . The increase is significantly more upon complexation with Al 3+ . However, fluorescence maxima are similar for the two complexes. This indicates that fluorescence intensity may not be a good parameter for Al 3+ sensing by salophen, in the presence of a large excess of Zn 2+ . This problem can be circumvented if fluorescence lifetime is used as the sensing parameter, as the lifetime of the Al 3+ complex is in the nanosecond time regime while that of the Zn 2+ complex is in tens of picoseconds. The significant difference in the fluorescence quantum yield and lifetime between the two complexes is explained as follows: the Al 3+ complex is monomeric, but the Zn 2+ complex is dimeric. Quantum chemical calculations indicate a higher density of states near the locally excited state for the dimeric complex. This may lead to more efficient nonradiative pathways.

The natural product epicocconone, owing to its unique fluorescence properties, has been developed into a range of products used in biotechnology, especially proteomics. However, its weak green fluorescence in its native state, while advantageous for proteomics applications, is a disadvantage in other applications that require two-color readouts. Here we report the photophysical characterization of two brightly fluorescent analogues of epicocconone. These analogues, with naphthyl or pyridyl groups replacing the heptatriene chain, resulted in bright fluorescence in both the native state and the long Stokes shifted enamine. Time-resolved fluorescence studies and DFT calculations were carried out to understand the excited state processes involved in fluorescence. Results showed the p-chloro group on the pyridyl is responsible for the high fluorescence of the native fluorophore. The application of one of these compounds for staining electrophoresis gels is exemplified.

Engineering the properties of fluorescent probes through modifications of the fluorophore structure has become a subject of interest in recent times. By doing this, the photophysical and photochemical properties of the modified fluorophore can be understood and this can guide the design and synthesis of better fluorophores for use in biotechnology. In this work, the electronic spectra and fluorescence decay kinetics of four analogues of the fluorescent natural product epicocconone were investigated. Epicocconone is unique in that the native state is weakly green fluorescent, whereas the enamine formed reversibly with proteins is highly emissive in the red. It was found that the ultrafast dynamics of the analogues depends profoundly on the H-bonding effect of solvents and solvent viscosity though solvent polarity also plays a role. Comparing the steady state and time-resolved data, the weak fluorescence of epicocconone in its native state is most likely due to the photoisomerization of the hydrocarbon side chain, while the keto enol moiety also has a role to play in determining the fluorescence quantum yield. This understanding is expected to aid the design of better protein stains from the same family.

2-(4'-Pyridyl)benzimidazole (4PBI) can exist in several states of protonation, having three basic nitrogen atoms. The equilibria involving these states, in ground as well as in excited states, are found to be affected significantly by cyclodextrins (CDs). The formation of inclusion complexes of this compound with all three varieties of cyclodextrins is observed to be more favorable at pH 9 than at pH 4, due to the predominance of the neutral form of dye at pH 9. The binding affinity of 4PBI to CDs is found to be governed by two factors: (i) the size of the host and (ii) the mode of insertion of 4PBI. We find that, for the host with a smaller cavity (α-CD), insertion of the dye with a pyridyl face is favored, whereas, for γ-CD, the preference is shifted toward the benzimidazole face of the dye. For β-CD, the binding affinity of the dye is maximum due to perfect cavity matching with the guest. A combination of steric factor and hydrogen bonding interaction is found to be responsible for modulation of the protonation-deprotonation equilibria of the guest molecule in the inclusion complex. Surprisingly, a protonated form is found to be promoted upon inclusion in cyclodextrins, under certain conditions. This is an unusual behavior and has been rationalized by prototropism involving the hydroxyl protons of cyclodextrin molecules.

An attempt is made to draw a line of comparison between the extent of rigidity of the hydration layers bound to the interfacial region of lamellar structures of Aerosol OT (AOT, sodium bis(2-ethylhexyl) sulfosuccinate) in water, in the presence and absence of an organic solvent using POM, SAXS, cryo-TEM, and time-resolved fluorescence spectroscopy. These systems are ternary mixtures of AOT, water, and n-heptane containing lamellar structures in an aqueous layer at higher w(0) values (w(0) = 300 and 150) and a binary solution of 20 and 50% AOT in neat water (w/w). The solvation shells residing at the vicinity of these lamellar structures are monitored using two different coumarin probes (C153 and C500). It is intended to envisage a comparative solvation dynamics study of the restricted aqueous region confined in lamellar structures formed in ternary mixture and binary solution. Though steady state measurements show a similar microenvironment probed by the fluorophores in lamellar structures formed in the two different aqueous phases, temporal evolution of the solvent correlation function C(t) unveils the existence of lamellar structures with different degrees of confinement of water layers in these two systems. A slower relaxation of the restricted aqueous region in lamellar structures of binary solution signifies the presence of more rigid interfacially bound water layers at the lamellar interface than in the ternary mixture having a similar weight percentage of AOT in water. The present investigation concludes that the lamellar structures formed under two different conditions provide a similar hydrophobic environment with different extents of localized water populations at the lamellar interface as manifested by the solvent relaxation time in agreement with SAXS and cryo-TEM images.

In this present investigation, attempt is made to use [2,2'-bipyridyl]-3,3'-diol (BP(OH)(2)) as a marker to study albumin-SDS interactions and to obtain structural information about these aggregates. It is also intended to contemplate the effect of these aggregates on the excited-state proton-transfer dynamics of BP(OH)(2). Steady-state and time-resolved fluorescence spectroscopic techniques are employed to elucidate the nature of interaction of two homologous carrier proteins, human serum albumin (HSA) and bovine serum albumin (BSA), with negatively charged surfactant sodium dodecyl sulfate (SDS). Both spectral and temporal behavior of BP(OH)(2) in these albumin-SDS aggregates strongly affirm an initial competitive binding of SDS in high-energy binding sites of albumin. Unlike normal SDS micelles, the absence of formation of the monocation of BP(OH)(2) at the negatively charged interface of SDS is rationalized by screening of the micellar interface in the presence of denatured protein which wraps around these surfactant aggregates. An enhanced extent of excited-state proton transfer is manifested by a corresponding increase in fluorescence quantum yield of BP(OH)(2) in these aggregates. Temporal evolution of BP(OH)(2) at different emission wavelengths fortifies the formation of normal micelles post saturation. All our observations are found to corroborate with the necklace and bead model proposed for protein-surfactant aggregates.

2,2'-(Pyridyl)benzimidazole is used as a probe to explore proton transfer through nafion membranes. The probe marks the availability of water in native as well as cation-exchanged membrane. Using steady state and time-resolved fluorescence studies, it has been shown that the rotation of the pyridyl and benzimidazole rings with respect to each other, which is ultrafast in higher water contents, is hindered as the water content in the membranes is decreased. In cation-exchanged membranes, it is observed that the formation of the ESPT (excited state proton transfer) state is reduced to a large extent. Thus, it may be inferred that the proton transport is observed to be hindered even in molecular dimensions of one water molecule thereby bolstering the contention that it may not be essential for water channels to break for proton conductivity to decrease., (© 2012 American Chemical Society)

The binding of [2,2'-bipyridyl]-3,3'-diol (BP(OH)(2)) with ionic and neutral surfactants like cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfate (SDS), and Triton X-100 (TX-100) has been studied by steady-state and time-resolved fluorescence spectroscopy. The absorption as well as emission spectra of BP(OH)(2) are highly sensitive toward the variation of surfactant concentration and hydrophobicity of the environment. The fluorescent state of the diketo form gains stability in surfactant assemblies, leading to a red-shifted emission spectra. A sharp increase in the fluorescence quantum yield near critical micellar concentration (CMC) is encountered followed by saturation. This indicates a complete encapsulation of BP(OH)(2) in the micelles. The maximum fluorescence quantum yield in anionic SDS is rationalized by the formation of cationic fluorophore at the Stern layer. The increase in quantum yield in neutral TX-100 is attributed to higher microviscosity experienced by the fluorophore in the palisade layer. A direct support in favor of this argument in TX-100 is provided by the viscosity dependence exhibited by the probe in different concentrations of sucrose solutions. CTAB exhibiting only hydrophobic effect shows least increase in qunatum yield of BP(OH)(2) among all the surfactants. Time-resolved fluorescence study of BP(OH)(2) in micelles is used as a tool to monitor the extent of micellization in the lipophilic cavity. An increase in fluorescence quantum yield as well as lifetime of BP(OH)(2) upon micellization indicates an enhanced extent of ESIDPT in hydrophobic medium.