Deboleena Sarkar

Binding Interaction of Cationic Phenazinium Dyes with Calf Thymus DNA: A Comparative Study

Absorption, steady-state fluorescence, steady-state fluorescence anisotropy, and intrinsic and induced circular dichroism (CD) have been exploited to explore the binding of calf thymus DNA (ctDNA) with three cationic phenazinium dyes, viz., phenosafranin (PSF), safranin-T (ST), and safranin-O (SO). The absorption and fluorescence spectra of all the three dyes reflect significant modifications upon interaction with the DNA. A comparative study of the dyes with respect to modification of fluorescence and fluorescence anisotropy upon binding, effect of urea, iodide-induced fluorescence quenching, and CD measurements reveal that the dyes bind to the ctDNA principally in an intercalative fashion. The effect of ionic strength indicates that electrostatic attraction between the cationic dyes and ctDNA is also an important component of the dye-DNA interaction. Intrinsic and induced CD studies help to assess the structural effects of dyes binding to DNA and confirm the intercalative mode of binding as suggested by fluorescence and other studies. Finally it is proposed that dyes with bulkier substitutions are intercalated into the DNA to a lesser extent.

Deciphering the perturbation of serum albumins by a ketocyanine dye: a spectroscopic approach.

Steady state and time resolved fluorometric and circular dichroism (CD) techniques have been exploited to explore the binding interaction of a ketocyanine dye, namely, 2-[3-(N-methyl-N-phenylamino)-2-propenylidene] indanone (MPAPI) with transport proteins, bovine serum albumin (BSA) and human serum albumin (HSA). The emission spectrum of buffered solution of the dye is found to be perturbed remarkably upon binding with the proteins. An explicit study with respect to modification of fluorescence and fluorescence anisotropy upon binding, effect of denaturant, fluorescence lifetime and CD measurements reveal that the dye binds with both BSA and HSA; the binding being stronger with the latter. Denaturation and CD studies reveal that stability of the proteins increases upon binding with the dye. The probable binding sites of the dye in the proteinous environments have been assessed from fluorescence resonance energy transfer (FRET) study. The probe is argued to be located in the inter domain cleft region of HSA (near Trp-214). From the similarity in the fluorescence behavior of the dye in BSA and HSA it is inferred that in BSA environment the probe is located near Trp-212 rather than Trp-132.

Photophysics and Rotational Dynamics of a β-Carboline Analogue in Nonionic Micelles: Effect of Variation of Length of the Headgroup and the Tail of the Surfactant

Effect of variation of length of nonionic surfactants in terms of the headgroup as well as the tail part on the photophysical and rotational dynamical properties of a beta-carboline analogue, 3-acetyl-4-oxo-6,7-dihydro-12H-indolo-[2,3-a]quinolizine (AODIQ) has been investigated. Steady-state and time-resolved fluorescence and fluorescence anisotropy have been exploited for the purpose. The experiments revealed modification of the photophysics of AODIQ by the conjugate effect of polarity and rigidity of the micellar environments with varying poly(ethylene oxide) chain length in the case of Triton X series and the alkyl chain length in the case of Tween series surfactants. Fluorometric studies suggest that the fluorophore resides at the micelle-water interface in all these systems. The enhancements in the steady-state anisotropy in all the micellar media compared to those in pure aqueous solution reflect that the fluorophore is located in motionally restricted regions introduced by the nonionic micelles. Contrary to the single exponential nature of the fluorescence anisotropy decay of AODIQ in aqueous medium, they were found to be biexponential in the micellar environments. The rotational relaxation of AODIQ in the micellar environments has been discussed in light of the two-step and wobbling in a cone model. The model helps to evaluate different rotational parameters and to ascertain the location of the fluorophore in the micellar media. The significant feature is that the motional restriction decreases with an increase in the poly(ethylene oxide) chain length while it increases with an increase in the alkyl chain length. The difference in the extent of water penetration due to variation in the thickness of the palisade layer and therefore a variation in the micellar polarity with a variation of the length of poly(ethylene oxide) and alkyl chain has been argued to be responsible.

Probing the binding interaction of a phenazinium dye with serum transport proteins: a combined fluorometric and circular dichroism study

In the present investigation, an attempt has been made to study the interaction of phenosafranin (PSF), a cationic phenazinium dye with the transport proteins, bovine serum albumin (BSA) and human serum albumin (HSA), employing steady‐state and time‐resolved fluorometric and circular dichroism (CD) techniques. The photophysical properties of the dye are altered on binding with the serum proteins. An explicit study with respect to the modification of the fluorescence and fluorescence anisotropy upon binding, effect of denaturant, fluorescence lifetime and CD measurements reveal that the dye binds to both BSA and HSA with almost the same affinity. Far‐UV CD spectra indicate a decrease in the percentage of α‐helicity only for BSA upon binding with the probe. Near‐UV CD responses indicate an alteration in the tertiary structure of both the transport proteins because of binding.

Binding of a cationic phenazinium dye in anionic liposomal membrane: a spectacular modification in the photophysics

Interaction of a cationic phenazinium dye, phenosafranin (PSF), with the anionic liposomal vesicle/bilayer of dimyristoyl-L-alpha-phosphatidylglycerol (DMPG) has been demonstrated using steady state and time resolved fluorescence and fluorescence anisotropy techniques. The charge transfer emission spectrum of PSF shows a dramatic modification in terms of fluorescence yield together with an appreciable hypsochromic shift in the lipid environment. The blue shift indicates a lowering in polarity inside the vesicle as compared to that in bulk water. The fluorescence and fluorescence quenching studies and micropolarity determination reveal that the cationic fluorophore has a profound binding interaction with the anionic DMPG membrane. Anisotropy study indicates the imposition of a motional restriction on the probe inside the bilayer. The electrostatic interaction between the cationic dye and the anionic lipid membrane has been argued to be the reason behind all these observations. The results could be useful in analyzing membrane organization and heterogeneity in natural membranes exploiting PSF or alike compounds as fluorescent probes.

Spectroscopic Characterization of Phenazinium Dye Aggregates in Water and Acetonitrile Media: Effect of Methyl Substitution on the Aggregation Phenomenon

Absorption, fluorescence, and fluorescence excitation spectral studies of two planar, cationic phenazinium dyes, namely, phenosafranin (PSF) and safranin-T (ST), have been performed in protic and aprotic polar solvents. The studies reveal the formation of both J- and H-aggregates in concentrated solutions. The planarity of the phenazinium skeleton and the presence of a positive charge are attributed to be the driving force for this aggregation behavior. The aggregates are established to be dimers only. The positive inductive effect of the methyl substituents in safranin-T augments the aggregation process. The experiments reveal that for both dyes, the polar protic solvent favors the aggregation process more than the aprotic solvent.

Differential Interaction of β-Cyclodextrin with Lipids of Varying Surface Charges: A Spectral Deciphering Using a Cationic Phenazinium Dye

Interaction of phenosafranin (PSF), a biologically potent cationic dye molecule, has been studied with zwitterionic and anionic lipid membranes of dimyristoyl-L-alpha-phosphatidylcholine (DMPC) and dimyristoyl-L-alpha-phosphatidylglycerol (DMPG), respectively. The effect of cyclic oligosaccharide, beta-cyclodextrin (beta-CD), on the stability of these probe-bound lipid bilayers has also been investigated exploiting steady state and time-resolved fluorescence, steady state fluorescence anisotropy, and dynamic light scattering techniques. An interpretation of membrane destabilization upon interaction of cyclodextrin with the lipids was drawn exploiting PSF as an extrinsic fluorescent probe. The fluorophore showed discernible interactions with DMPC and DMPG vesicles. Experimental results reveal that the extent of interaction of PSF with DMPG is greater compared to that with DMPC. Addition of beta-CD into the PSF-bound lipids showed a differential effect for the two lipids of varying surface charge characteristics. In the case of DMPC, addition of beta-CD resulted in a preferential interaction of the probe with CD. However, addition of beta-CD to PSF-bound DMPG resulted in the selective interaction of DMPG with the added CD leading to the release of the probe into the bulk aqueous medium.

Application of anionic micelle for dramatic enhancement in the quenching-based metal ion fluorosensing

We introduce a simple and efficient strategy to enhance the efficiency of a quenching-based fluorosensor for metal ions by several orders of magnitude by using commercially available anionic surfactants varying hydrophobic chain length. Anionic surfactants with a proper choice of hydrophobic chain length at their optimum concentrations are efficient to boost up the efficiency of copper ion sensor dramatically. This simple and convenient strategy is, in general, applicable to quenching-based fluorosensors, new or established, in aqueous solution. It is powerful enough to transform a virtually non-sensor fluorophore to a sensor with a commendable efficiency with the help of proper surfactant. Thus, in this communication, light has been thrown on the application of surfactants toward increasing fluorosensing efficiency of a quenching based sensor.

Photophysics and rotational relaxation dynamics of a β-carboline based fluorophore in cationic alkyltrimethylammonium bromide micelles

Photophysics and rotational relaxation dynamics of a beta-carboline analog, 3-acetyl-4-oxo-6,7-dihydro-12H-indolo-[2,3-a] quinolizine (AODIQ) have been investigated in cationic alkyltrimethylammonium bromide (nTAB) micelles using steady-state and time-resolved fluorometric techniques. The study reveals modification of its photophysics by the conjugate effect of polarity and rigidity of the micellar environments with varying alkyl chain lengths of the surfactants. Furthermore, it suggests that the fluorophore resides at the micelle-water interfacial domain. Contrary to the single exponential nature of the fluorescence anisotropy decay of AODIQ in aqueous medium, the decay is found to be biexponential in all the micellar environments studied. The enhancements in the steady-state anisotropy and rotational relaxation time in the micellar media compared to that in pure aqueous solution reflect that the fluorophore resides in a motionally restricted environment introduced by the cationic micelles. The rotational correlation time increases marginally with an increase in the surfactant chain length. The rotational relaxation of AODIQ in the micellar environments has been discussed in the light of the two-step and wobbling in a cone model. The model helps in evaluating different rotational parameters and in ascertaining the location of the fluorophore in the micellar media. This technique provides valuable information regarding the rotational relaxations of the fluorophore within an organized assembly. When the lifetime measurements and orientational relaxation measurements are combined, significant inferences can be made regarding the partitioning of the probe in different regions of the micelles.

Photophysics and rotational relaxation dynamics of cationic phenazinium dyes in anionic reverse micelles: effect of methyl substitution

We present here, a detailed photophysical and rotational relaxation dynamical study of three structurally analogous cationic dyes, namely, phenosafranin (PSF), safranin-T (ST), and safranin-O (SO), carried out in well characterized, monodispersed biomimicking anionic reverse micellar nanocavities composed of sodium bis(2-ethylhexyl)sulfosuccinate (AOT)/heptane with increasing water contents. The dyes belong to the phenazinium family and they differ in terms of methyl substitution on the planar phenazinium skeleton. The objective of the present study is to investigate the modification in the photophysical and dynamical behavior of the dyes with the change in the size of the water pool of the reverse micelle and thereby to explore the role of methyl substitution. Steady state and time resolved emission and anisotropy studies have been exploited for the purpose. The dyes are found to exhibit a marked decrease in the fluorescence anisotropy with increasing water/surfactant mole ratio (w), i.e., the water pool size in the reverse micellar core, implying that overall motional restriction experienced by the molecules are decreased with increasing hydration. Some of the depth dependent fluorescence parameters such as fluorescence maximum, fluorescence anisotropy (r) have been monitored for exploring the microenvironment around the probes in the reverse micelles. Fluorescence studies suggest that at low w values, the probes do not penetrate into the reverse micellar core; rather it binds at the interfacial region. Estimates of the micropolarity at the binding sites of the probe molecule have been determined as a function of w. Finally, dynamic studies reveal that both the lifetime and rotational relaxation time decrease with an increase in w for all the three probes, the extent of the decrease being more for PSF than ST and SO. This indicates a stronger binding of the reverse micelle with ST and SO compared to that with PSF which is rationalized in terms of an increase in the hydrophobicity of the former two dyes because of the methyl substitution on the phenazinium moiety.