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The modulation of the prototropic equilibrium of a cancer cell photosensitizer, norharmane (NHM), within a niosome microheterogeneous environment has been investigated. The contrasting effects of temperature and extrinsically added salt on the photophysics of niosome-bound drug have been meticulously explored from steady-state and time-resolved spectroscopic techniques. The cation ⇌ neutral prototropic equilibrium of NHM is found to be preferentially favored toward the neutral species with increasing salt concentration, and the results are rationalized on the basis of water penetration to the hydration layer of niosome. The effects are typically reversed with temperature. The differential rotational relaxation behavior of NHM under various conditions has also been addressed from fluorescence anisotropy decay. Further, the study delineates the application of β-cyclodextrin (βCD) as a potential host system, leading to drug sequestration from the niosome-encapsulated state. To this end, a detailed investigation of the thermodynamics of the niosome:βCD interaction has been undertaken by isothermal titration calorimetry (ITC) to unravel the notable dependence of the thermodynamic parameters on temperature. Consequently, a critical analysis of the variation of the enthalpy change (ΔH) of the process with temperature leads to the unique observation of a positive heat capacity change (ΔCp) marking the hallmark of hydrophobic hydration.

Herein, we present an endeavor toward exploring the lacuna underlying the host:guest chemistry of inclusion complex formation between bile salt(s) and β-cyclodextrin(s) (βCDs). An extensive thermodynamic investigation based on isothermal titration calorimetry (ITC) demonstrates a dominant contribution from exothermic enthalpy change (ΔH < 0) accompanying the phenomenon of inclusion complex formation, along with a relatively smaller contribution to total free energy change from the entropic component. However, the negative heat capacity change (ΔCp < 0) displays the hallmark for a pivotal role of hydrophobic effect underlying the interaction. Contrary to the classical hydrophobic effect, such apparently paradoxical thermodynamic signature has been adequately described under the notion of "nonclassical hydrophobic effect". On the basis of our results, the displacement of disordered water from hydrophobic binding sites has been argued to mark the enthalpic signature and the key role of such interaction forces is further corroborated from enthalpy-entropy compensation behavior showing indication for almost complete compensation. To this end, we have quantified the interaction of two bile salt molecules (namely, sodium deoxycholate and sodium glycocholate) with a series of varying chemical substituents on the host counterpart, namely, βCD, (2-hydroxypropyl)-βCD, and methyl βCD.

Herein, the binding interaction of a potential chemotherapeutic antibacterial drug norfloxacin (NOF) with a serum transport protein, human serum albumin (HSA), is investigated. The prototropic transformation of the drug (NOF) is found to be remarkably modified following interaction with the protein as manifested through significant modulations of the photophysics of the drug. The predominant zwitterionic form of NOF in aqueous buffer phase undergoes transformation to the cationic form within the protein-encapsulated state. This implies the possible role of electrostatic interaction force in NOF-HSA binding. This postulate is further substantiated from the effect of ionic strength on the interaction process. To this end, the detailed study of the thermodynamics of the drug-protein interaction process from isothermal titration calorimetric (ITC) experiments is found to unfold the signature of electrostatic as well as hydrophobic interaction forces underlying the binding process. Thus, interplay of more than one interaction forces is argued to be responsible for the overall drug-protein binding. The ITC results reveal an important finding in terms of enthalpy-entropy compensation (EEC) characterizing the NOF-HSA binding. The effect of drug-binding on the native protein conformation has also been evaluated from circular dichroism (CD) spectroscopy which unveils partial rupture of the protein secondary structure. In conjunction to this, the functionality of the native protein (in terms of esterase-like activity) is found to be lowered as a result of binding with NOF. The AutoDock-based docking simulation unravels the probable binding location of NOF within the hydrophilic subdomain IA of HSA. The present program also focuses on exploring the dynamical aspects of the drug-protein interaction scenario. The rotational-relaxation dynamics of the protein-bound drug reveals the not-so-common "dip-and-rise" pattern.

The effect of variation of the size of headgroup as well as the length of hydrocarbon tail of nonionic surfactants on the photophysics and rotational-relaxation dynamics of a promising biological photosensitizer, norharmane, (NHM) has been investigated. The series of nonionic micelles employed for the study belongs to Triton X family (allowing the variation in poly(ethylene oxide) (PEO) chain length) and Tween family (providing access to vary the alkyl chain length of the surfactant tails). The spectral deciphering of the photophysics of the drug (NHM) within the series of the nonionic micelles reveals remarkable influence of binding of the drug with the micelles on the prototropic equilibrium which is notably favored toward the neutral species of the drug over the cationic counterpart. The strength of drug-micelle binding interaction as well as the extent of transformation of the cation ⇌ neutral prototropic equilibrium is found to be enormously governed by the variation of the headgroup size and the alkyl chain length of the surfactants. To this end, the equilibrium constant (Keq) and free energy change (ΔG) for cation ⇌ neutral prototropic transformation of the drug as a function of the micellar parameters have been meticulously explored from emission studies and comprehensively rationalized under the provision of the micellar hydration model, that is, the differential extents of water penetration to micellar units as a function of varying thickness of the palisade layer and hence a variation in the polarity of the micellar microenvironments. The significant enhancement in steady-state fluorescence anisotropy of NHM in micellar environments compared to that in bulk aqueous buffer phase substantiates the location of the drug in motionally constrained regions introduced by the nonionic micelles. Further, all these lines of arguments are effectively corroborated from time-resolved fluorescence experiments with particular emphasis on time-resolved anisotropy decay study of the drug within the micellar aggregates.

The present work demonstrates the photophysical characterization of the interaction of a promising cancer cell photosensitizer, harmane (HM), with biomimetic micellar nanocavities having varying surface charge characteristics. The polarity-sensitive prototropic transformation of HM is remarkably modified upon interaction with the macromolecular assemblies of micellar systems and is manifested through significant modulations on the absorption and emission profiles of HM. The ground- and excited-states prototropic equilibria of HM are found to be differentially modulated in various micellar assemblies. Out of various possibilities to assess the drug (HM)-micelle interaction mechanism, the postulate of varying extent of drug penetration into micellar units depending on the compactness of their headgroup arrangements is found to suitably rationalize and correlate different experimental findings, including the differences in binding constant (K) and free energy change (ΔG) of the interaction process. The micropolarity measurement has been exploited to evaluate the probable binding location of the drug which reveals that the cationic drug molecule does not penetrate deep into the micellar core region and the results are further substantiated from fluorescence quenching experiments. The work also pays proper attention to delineate the modulation in dynamical behaviors of the drug following interaction with the micellar systems. Wavelength-sensitive fluorescence parameters reveal the slower rate of solvent-relaxation around the excited probe within the micelle-encapsulated microheterogeneous environments. The enhancement of fluorescence anisotropy and rotational relaxation time of the drug in micellar environments from that in pure aqueous buffer suggests entrapment of the drug in motionally constrained regions introduced by the micelles.

The present study aims at exploring a detailed characterization of the binding interaction of a promising cancer cell photosensitizer, harmane (HM), with DNA extracted from herring sperm. The polarity-sensitive prototropic transformation of HM, a naturally occurring, fluorescent, drug-binding alkaloid, β-carboline, is remarkably modified upon interaction with DNA and is manifested through significant modulations on the absorption and emission profiles of HM. From the series of studies undertaken in the present program, for example, absorption; steady-state emission; the effect of chaotrope (urea); iodide ion-induced steady-state fluorescence quenching; circular dichroism (CD); and helix melting from absorption spectroscopy; the mode of binding of HM into the DNA helix has been substantiated to be principally intercalative. Concomitantly, a discernible dependence of the photophysics of the DNA-bound drug on the medium ionic strength indicates that electrostatic attraction should not be ignored in the interaction. Efforts have also been delivered to delineate the dynamical aspects of the interaction, such as modulation in time-resolved fluorescence decay and rotational relaxation dynamics of the drug within the DNA environment. In view of the prospective biological applications of HM, the issue of facile dissociation of intercalated HM from the DNA helix also comprises a crucial prerequisite for the functioning as an effective therapeutic agent. In this context, our results imply that the concept of detergent-sequestered dissociation of the drug from the drug-DNA complex can be a prospective strategy through an appropriate choice of the detergent molecule. The utility of the present work resides in exploring the potential applicability of the fluorescence property of HM for studying its interactions with a relevant biological target, for example, DNA. In addition, the methods and techniques used in the present work can also be exploited to study the interaction of HM with other biological, biomimicking assemblies and drug delivery vehicles, and so forth.

The present work describes the interaction of a promising cancer cell photosensitizer, harmane (HM), with a model transport protein, Bovine Serum Albumin (BSA). The studied molecule of interest (HM) belongs to the family of naturally occurring fluorescent drug-binding alkaloids, the β-carbolines. A combined use of steady-state and time-resolved fluorescence techniques is applied to follow and characterize the binding interaction. The polarity-dependent prototropic activity of HM is found to be responsible for the commendable sensitivity of the probe to the protein environments and is distinctly reflected on the emission profile. Steady-state fluorescence anisotropy study reveals the impartation of a considerable degree of motional restriction on the drug molecule as a result of binding to the protein. Contrary to the single-exponential nature of fluorescence anisotropy decay of HM in aqueous buffer, they are found to be biexponential in the protein environment. The rotational relaxation dynamics of HM within the protein has been interpreted on the lexicon of the Two-Step and Wobbling-in-Cone model. The probable binding location for the cationic drug is found to be the hydrophilic binding zone of BSA, i.e., domain I (characterized by a net negative charge). The AutoDock-based blind docking simulation has been explored for evaluating an unbiased result of the probable interaction site of HM in the protein. To unfold the effect of binding of the drug on the secondary structural content of the protein, circular dichroism (CD) spectroscopy has been exploited to see that binding of the drug accompanies some decrease in α-helical content of BSA, and the effect gradually saturates toward a higher drug/protein molar ratio., (© 2011 American Chemical Society)

The present work demonstrates the modulation of excited state intramolecular proton transfer (ESIPT) emission of 1-hydroxy-2-naphthaldehyde (HN12) upon its interaction with the liposomal vesicles/bilayer of dimyristoyl-l-α-phosphatidylcholine (DMPC) and dimyristoyl-l-α-phosphatidylglycerol (DMPG) and its subsequent implementation as an efficient molecular reporter for probing of microheterogeneous environments of lipid-bilayer system. Modifications on the emission profile of HN12 in terms of remarkable emission intensity enhancement coupled with a hypsochromic shift induced by the presence of DMPC and DMPG membranes have been interpreted to be vivid manifestations of the interactions between the two partners. Steady-state anisotropy, red-edge excitation shift (REES), and time-resolved fluorescence measurements have been fruitfully exploited to complement other experimental findings. Probable binding site of HN12 in the lipid-bilayers has been assessed on the basis of intertwining the results of fluorescence quenching with other experimental results and is further substantiated from docking studies.

A simple intramolecular charge transfer (ICT) compound, 5-(4-dimethylamino-phenyl)-penta-2,4-dienoic acid methyl ester (DPDAME), has been documented to be a potential molecular reporter for probing microheterogeneous environments of a model transport protein bovine serum albumin (BSA) using spectroscopic techniques. Meteoric modifications to the emission profile of DPDAME upon addition of BSA come out to be a result of its binding to hydrophobic subdomain IIA. The highly polarity-sensitive ICT emission of DPDAME is found to be a proficient extrinsic molecular reporter for efficient mapping of native, intermediate, unfolded, and refolded states of the protein. Experimental data coupled with a reinforcing support from theoretical simulation using CHARMM22 software confirm the binding site of the probe to be the subdomain IIA of BSA, while FRET study reveals a remarkably close approach of our extrinsic molecular reporter to Trp-212 (in domain IIA): the distance between DPDAME and Trp-212 is 1.437 nm. The caliber of DPDAME as an external fluorescence marker also extends to the depiction of protein-surfactant (BSA-SDS) interaction to commendable fruition. Additionally, the protective action of small amounts of SDS on urea-denatured protein is documented by polarity-sensitive ICT emission of the probe. The present study also reflects the enhancement of the stability of BSA with respect to chemically induced denaturation by urea as a result of binding to the probe DPDAME.