Development of an optical sensor for chlortetracycline detection based on the fluorescence quenching of l‐tryptophan

A simple and selective spectrofluorimetric method for the detection of chlortetracycline (CTC) was studied. In pH 7.4 buffer medium l ‐tryptophan (l ‐Trp), applied as the fluorescence probe, interacted with CTC resulting in fluorescence quenching of the probe. CTC was detected with maximum excitation and emission wavelengths at λ_ex/λ_em = 275/350 nm. Notably, quenching of fluorescence intensities was positively proportional to the CTC concentration over the range of 0.65–30 μmol L^?1 and the limit of detection was 0.2 μmol L^?1. Effect of temperature shown in Stern?Volmer plots, absorption spectra and fluorescence lifetime determination, indicated that fluorescence quenching of l ‐Trp by CTC was mainly by static quenching. The proposed study used practical samples analysis satisfactorily.     

Luminescence of bacteriorhodopsin from Halobacterium halobium and its connection with the photochemical conversions of the chromophore

Red luminescence of purple membranes from Halobacterium halobium cells in suspension, dry film or freeze-dried preparations was studied and its emission, excitation and polarization spectra are reported. The emission spectra have three bands at 665–670, 720–730 and at 780–790 nm. The position (maximum at 580 nm) and shape of the excitation spectra are close to those of the absorption spectra. The spectra depend on experimental conditions, in particular on pH of the medium. Acidification increases the long wavelength part of the emission spectra and shifts the main excitation maximum 50–60 nm to the longer wavelength side. Low-temperature light-induced changes of the absorption, emission and excitation spectra are presented. Several absorbing and emitting species of bacteriorhodopsin are responsible for the observed spectral changes. The bacteriorhodopsin photoconversion rate constant was estimated to be about 1 · 10¹¹ s^?1 at ? 196°C from the quantum yields of the luminescence (1 · 10^?3) and photoreaction (1 · 10^?1). The temperature dependence of the luminescence quantum yield points to the existence of two or three quenching processes with different activation energies. High degree of luminescence polarization (about 45–47%) throughout the absorption and fluorescence spectra and its temperature independence show that there is no energy transfer between bacteriorhodopsin molecules and no chromophore rotation during the excitation lifetime. In carotenoid-containing membranes, energy migration from the bulk of carotenoids to bacteriorhodopsin was not found either. Bacteriorhodopsin phosphorescence was not observed in the 500–1100 nm region and the emission is believed to be fluorescence by nature.     

Interaction between felodipine and bovine serum albumin: fluorescence quenching study

The fluorescence quenching spectrum of bovine serum albumin (BSA) was investigated in the presence of felodipine (FLD) by spectroscopic methods including fluorescence spectroscopy and UV–Vis absorption spectroscopy. Stern–Volmer quenching was successfully applied and the corresponding thermodynamic parameters, namely enthalpy change (ΔH), free energy change (ΔG) and entropy change (ΔS) at different temperatures (304, 314 and 324 K) were calculated according to the Van't Hoff relation. This revealed that the hydrophobic interaction plays a major role in stabilizing the complex. The fluorescence spectrum of BSA was studied in presence of various concentrations of SDS surfactant. The distance (r) between donor (BSA) and acceptor (FLD) was obtained according to fluorescence resonance energy transfer (FRET). The synchronous fluorescence spectroscopy was used to investigate the effect of FLD on BSA molecule. The result shows that the conformation of BSA was changed in the presence of felodipine. Copyright © 2009 John Wiley & Sons, Ltd.     

In vitro study on binding interaction of quinapril with bovine serum albumin (BSA) using multi-spectroscopic and molecular docking methods

The binding interaction between quinapril (QNPL) and bovine serum albumin (BSA) in vitro has been investigated using UV absorption spectroscopy, steady-state fluorescence spectroscopic, synchronous fluorescence spectroscopy, 3D fluorescence spectroscopy, Fourier transform infrared spectroscopy, circular dichroism, and molecular docking methods for obtaining the binding information of QNPL with BSA. The experimental results confirm that the quenching mechanism of the intrinsic fluorescence of BSA induced by QNPL is static quenching based on the decrease in the quenching constants of BSA in the presence of QNPL with the increase in temperature and the quenching rates of BSA larger than 10¹⁰ L mol^?1 s^?1, indicating forming QNPL–BSA complex through the intermolecular binding interaction. The binding constant for the QNPL–BSA complex is in the order of 10⁵ M^?1, indicating there is stronger binding interaction of QNPL with BSA. The analysis of thermodynamic parameters together with molecular docking study reveal that the main binding forces in the binding process of QNPL with BSA are van der Waal’s forces and hydrogen bonding interaction. And, the binding interaction of BSA with QNPL is an enthalpy-driven process. Based on Förster resonance energy transfer, the binding distance between QNPL and BSA is calculated to be 2.76 nm. The results of the competitive binding experiments and molecular docking confirm that QNPL binds to sub-domain IIA (site I) of BSA. It is confirmed there is a slight change in the conformation of BSA after binding QNPL, but BSA still retains its secondary structure α-helicity.     

Spectral analysis on origination of the bands at 437 nm and 475.5 nm of chlorophyll fluorescence excitation spectrum in Arabidopsis chloroplasts

Chlorophyll fluorescence has been often used as an intrinsic optical molecular probe to study photosynthesis. In this study, the origin of bands at 437 and 475.5 nm in the chlorophyll fluorescence excitation spectrum for emission at 685 nm in Arabidopsis chloroplasts was investigated using various optical analysis methods. The results revealed that this fluorescence excitation spectrum was related to the absorption characteristics of pigment molecules in PSII complexes. Moreover, the excitation band centred at 475.5 nm had a blue shift, but the excitation band at 437 nm changed relatively less due to induction of non‐photochemical quenching (NPQ). Furthermore, fluorescence emission spectra showed that this blue shift occurred when excitation energy transfer from both chlorophyll b (Chl b) and carotenoids (Cars) to chlorophyll a (Chl a) was blocked. These results demonstrate that the excitation band at 437 nm was mainly contributed by Chl a, while the excitation band at 475.5 nm was mainly contributed by Chl b and Cars. The chlorophyll fluorescence excitation spectrum, therefore, could serve as a useful tool to describe specific characteristics of light absorption and energy transfer between light‐harvesting pigments. Copyright © 2015 John Wiley & Sons, Ltd.     

Blue-green fluorescence excited by UV laser on leaves of different species originates from cutin and is sensitive to leaf temperature

Under ultra-violet excitation, intact leaves generate a strong blue-green fluorescence emission with several bands. Their integrated energy is 6 to 11 times the energy released by chlorophyll a bands (Chappelle et al. 1984, Applied Optics 23, 134–138). This paper provides evidence that the blue-green fluorescence emission comes mainly from outer epidermal layers of the leaves and can be transferred on a quartz lamina by quickly dipping the leaves in organic solvents with subsequent solvent evaporation. Blue-green fluorescence displays a diffusion-controlled quenching of fluorescence intensity between 4°C (high fluorescence) and 37°C (low fluorescence). The blue-green fluorescence emissivity is not linked to short-term metabolic effects other than leaf temperature, but epidermis adaptations both to drought and to excessive radiation increase emissivity.     

Fluorescence spectrometric study on the interaction of tamibarotene with bovine serum albumin

The interaction between tamibarotene and bovine serum albumin (BSA) was studied using fluorescence quenching technique and ultraviolet–visible spectrophotometry. The results of experiments showed that tamibarotene could strongly quench the intrinsic fluorescence of BSA by a dynamic quenching mechanism. The apparent binding constant, number of binding site and corresponding thermodynamic parameters at different temperatures were calculated respectively, and the main interaction force between tamibarotene and BSA was proved to be hydrophobic force. Synchronous fluorescence spectra showed that tamibarotene changed the molecular conformation of BSA. When BSA concentration was 1.00 × 10^?6 mol L^?1, the quenched fluorescence ΔF had a good linear relationship with the concentration of tamibarotene in the range 1.00 × 10^?6 to 12.00 × 10^?6 mol L^?1 with the detection limit of 6.52 × 10^?7 mol L^?1. Copyright © 2010 John Wiley & Sons, Ltd.     

Interaction of cyproheptadine hydrochloride with human serum albumin using spectroscopy and molecular modeling methods

The interaction between cyproheptadine hydrochloride (CYP) and human serum albumin (HSA) was investigated by fluorescence spectroscopy, UV–vis absorption spectroscopy, Fourier transform infrared spectroscopy (FT‐IR) and molecular modeling at a physiological pH (7.40). Fluorescence of HSA was quenched remarkably by CYP and the quenching mechanism was considered as static quenching since it formed a complex. The association constants K_a and number of binding sites n were calculated at different temperatures. According to Förster's theory of non‐radiation energy transfer, the distance r between donor (human serum albumin) and acceptor (cyproheptadine hydrochloride) was obtained. The effect of common ions on the binding constant was also investigated. The effect of CYP on the conformation of HSA was analyzed using FT‐IR, synchronous fluorescence spectroscopy and 3D fluorescence spectra. The thermodynamic parameters ΔH and ΔS were calculated to be ?14.37 kJ mol^?1 and 38.03 J mol^?1 K^?1, respectively, which suggested that hydrophobic forces played a major role in stabilizing the HSA‐CYP complex. In addition, examination of molecular modeling indicated that CYP could bind to site I of HSA and that hydrophobic interaction was the major acting force, which was in agreement with binding mode studies. Copyright © 2012 John Wiley & Sons, Ltd.     

Probing the interaction of caffeic acid with ZnO nanoparticles

The binding of ZnO nanoparticles (NPs) and caffeic acid (CFA) was investigated using fluorescence quenching, UV/vis absorption spectrscopy, Fourier transform infrared (FTIR) spectroscopy, transmission electron microscopy (TEM) and dynamic light scattering (DLS) at different temperatures. The study results indicated fluorescence quenching between ZnO NPs and CFA rationalized in terms of a static quenching mechanism or the formation of non‐fluorescent CFA–ZnO. From fluorescence quenching spectral analysis, the binding constant (K_a), number of binding sites (n) and thermodynamic properties were determined. Values of the quenching (K_SV) and binding (K_a) constants decrease with increasing temperature and the number of binding sites n = 2. The thermodynamic parameters determined using Van't Hoff equation indicated that binding occurs spontaneously involving the hydrogen bond, and van der Waal's forces played a major role in the reaction of ZnO NPs with CFA. The FTIR, TEM and DLS measurements also indicated differences in the structure, morphology and size of CFA, ZnO NPs and their corresponding CFA–ZnO. Copyright © 2015 John Wiley & Sons, Ltd.     

The binding characteristics of the interaction between 3-(2-cyanoethyl) cytosine and human serum albumin

The binding characteristics of the interaction between 3-(2-cyanoethyl) cytosine (CECT) and human serum albumin (HSA) were investigated using fluorescence, UV absorption spectroscopic and molecular modeling techniques under simulative physiological conditions. The intrinsic fluorescence intensity of HSA was decreased with the addition of CECT. The fluorescence data handled by Stern–Volmer equation proved that the quenching mechanism of the interaction between CECT and HSA was a static quenching procedure. The binding constants evaluated utilizing the Lineweaver–Burk equation at 17, 27 and 37?°C, were 2.340?×?10⁴, 2.093?×?10⁴ and 1.899?×?10⁴?L?mol^?1, respectively. The thermodynamic parameters were calculated according to van’t Hoff equations. Negative enthalpy (ΔH) and positive entropy (ΔS) values indicated that both hydrogen bond and hydrophobic force played a major role in the binding process of CECT to HSA, which was consistent with the results of the molecular modeling study. In addition, the effect of other ions on the binding constant of CECT-HSA was examined.     

Study on the interaction between ginsenoside Rh2 and calf thymus DNA by spectroscopic techniques

The interaction between ginsenoside Rh2 (G‐Rh2) and calf thymus DNA (ctDNA) was investigated by spectroscopic methods including UV–vis absorption, fluorescence and circular dichroism (CD) spectroscopy, coupled with DNA melting techniques and viscosity measurements. Stern–Volmer plots at different temperatures proved that the quenching mechanism was a static quenching procedure. The thermodynamic parameters, enthalpy change (ΔH) and entropy change (ΔS) were calculated to be –22.83 KJ · mol^–1and 15.11 J · mol^–1 · K^–1by van ’t Hoff equation, suggesting that hydrophobic force might play a major role in the binding of G‐Rh2 to ctDNA. Moreover, the fluorescence quenching study with potassium iodide as quencher indicated that the K_SV (Stern–Volmer quenching constant) value for the bound G‐Rh2 with ctDNA was lower than the free G‐Rh2. The relative viscosity of ctDNA increased with the addition of G‐Rh2 and also the ctDNA melting temperature increased in the presence of G‐Rh2. Denatured DNA studies showed that quenching by single‐stranded DNA was less than that by double‐stranded DNA. The observed changes in CD spectra also demonstrated that the intensities of the positive and negative bands decreased with the addition of G‐Rh2. The experimental results suggest that G‐Rh2 molecules bind to ctDNA via an intercalative binding mode. Copyright © 2015 John Wiley & Sons, Ltd.     

Silicon nanoparticles synthesized using a microwave method and used as a label‐free fluorescent probe for detection of VB12

A simple and rapid detection strategy for vitamin B₁₂ (VB₁₂) was established based on label‐free silicon quantum dots (SiQDs); the detection mechanism was additionally investigated. SiQDs were synthesized using a one‐step microwave method, and their fluorescence was stronger than that synthesized using the hydrothermal method. SiQDs fluorescence was quenched using VB₁₂ due to the inner filter effect (IFE), which was demonstrated using ultraviolet (UV) absorption spectra, fluorescence lifetime, transmission electron microscopy and zeta potential analysis. Subsequently, quercetin (Que) and doxorubicin (Dox) with absorption peaks that overlapped the excitation or emission peaks of SiQDs respectively were used as control groups to investigate the quenching mechanism. Results showed that quenching efficiency was related to the level of overlap between the adsorption peak of the quencher and the excitation or emission peaks of SiQDs. A greater level of overlap caused a higher quenching efficiency. Therefore, the sensitive quenching of VB₁₂ for SiQDs was due to the synergistic effect of the synchronous overlap between the absorption peak of VB₁₂ with the excitation and emission peaks of SiQDs. Fluorescence quenching efficiency increased linearly in the 0.5 to 16.0 μmol·L^?1 VB₁₂ concentration range, and the detection limit was 158 nmol·L^?1. In addition, SiQDs were applied to determine VB₁₂ in tablets and human urine samples with satisfactory recoveries ranging from 97.7 to 101.1%.     

Study of the interaction between 8-azaguanine and bovine serum albumin using optical spectroscopy and molecular modeling methods

The interaction between 8-azaguanine (8-Azan) and bovine serum albumin (BSA) in Tris-HCl buffer solutions at pH 7.4 was investigated by means of fluorescence and ultraviolet-visible (UV-Vis) spectroscopy. At 298 K and 310 K, at a wavelength of excitation (λ _ex) of 282 nm, the fluorescence intensity decreased significantly with increasing concentrations of 8-Azan. Fluorescence static quenching was observed for BSA, which was attributed to the formation of a complex between 8-Azan and BSA during the binding reaction. This was illuminated further by the UV-Vis absorption spectra and the decomposition of the fluorescence spectra. The thermodynamic parameters ∆G, ∆H, ∆S were calculated. The results showed that the forces acting between 8-Azan and BSA were typical hydrophobic forces, and that the interaction process was spontaneous. The interaction distance r between 8-Azan and BSA, evaluated according to fluorescence resonance energy transfer theory, suggested that there is a high possibility of energy transfer from BSA to 8-Azan. Theoretical investigations based on homology modeling and molecular docking suggested that binding between 8-Azan and BSA is dominated by hydrophilic forces and hydrogen bonding. The theoretical investigations provided a good structural basis to explain the phenomenon of fluorescence quenching between 8-Azan and BSA.     

Spectroscopic exploration and thermodynamic characterization of desvenlafaxine interacting with fluorescent bovine serum albumin

The mechanism of the interaction between bovine serum albumin (BSA) and desvenlafaxine was studied using fluorescence, ultraviolet absorption, 3‐dimensional fluorescence spectroscopy, circular dichroism, synchronous fluorescence spectroscopy, cyclic voltametry, differential scanning calorimetry, and attenuated total reflection–Fourier transform infrared spectroscopic techniques under physiological condition at pH 7.4. Stern‐Volmer calculations authenticate the fluorescence of BSA that was quenched by desvenlafaxine in a collision quenching mode. The fluorescence quenching method was used to evaluate number of binding sites “n” and binding constant K _A that were measured, and various thermodynamic parameters were evaluated at different temperatures by using the van't Hoff equation and differential scanning calorimetry technique, which indicated a spontaneous and hydrophobic interaction between BSA and desvenlafaxine. According to the Förster theory we calculate the distance between the donor, BSA and acceptor, desvenlafaxine molecules. Furthermore, circular dichroism and attenuated total reflection–Fourier transform infrared spectroscopy indicate nominal changes in the secondary structure of the protein.     

Probing the structural interactions between methotrexate and dexamethasone with muscle cystatin: a biophysical study

Abstract

Drug protein interactions have gained considerable attention over the past many years. In the current communication the association of muscle cystatin (MC) with anti-rheumatic drugs methotrexate and dexamethasone was studied by thiol proteinase inhibitor assay, ultra violet (UV) absorption, fluorescence spectroscopy, and fluorescence transform infra-red spectroscopy (FTIR). A static pattern of quenching was noticed between muscle cystatin and methotrexate (MTX). Binding constant (K_a) of methotrexate to muscle cystatin was found to be 1?×?10^?7 M^?1 and the stoichiometry of binding was calculated to be one. Fluorescence measurement of the emission quenching reveals that the quenching process of cystatin by dexamethasone (DXN) was also static. The stoichiometry of binding and binding constant was also obtained. Additional evidence regarding MTX–MC and DXN–MC was obtained from UV spectroscopy and FTIR spectroscopic results. Such spectroscopic studies would help in modelling new candidate drugs for rheumatoid arthritis based on their cystatin binding profile.

Communicated by Ramaswamy H. Sarma     

Characterization of domain-specific interaction of synthesized dye with serum proteins by spectroscopic and docking approaches along with determination of in vitro cytotoxicity and antiviral activity

The interaction between a synthesized dye with proteins, bovine, and human serum albumin (BSA, HSA, respectively) under physiological conditions has been characterized in detail, by means of steady-state and time-resolved fluorescence, UV–vis absorption, and circular dichroism (CD) techniques. An extensive time-resolved fluorescence spectroscopic characterization of the quenching process has been undertaken in conjugation with temperature-dependent fluorescence quenching studies to divulge the actual quenching mechanism. From the thermodynamic observations, it is clear that the binding process is a spontaneous molecular interaction, in which van der Waals and hydrogen bonding interactions play the major roles. The UV–vis absorption and CD results confirm that the dye can induce conformational and micro-environmental changes of both the proteins. In addition, the dye binding provokes the functionality of the native proteins in terms of esterase-like activity. The average binding distance (r) between proteins and dye has been calculated using FRET. Cytotoxicity and antiviral effects of the dye have been found using Vero cell and HSV-1F virus by performing MTT assay. The AutoDock-based docking simulation reveals the probable binding location of dye within the sub-domain IIA of HSA and IB of BSA.     

Deciphering binding mechanism between bovine serum albumin and new pyrazoline compound K4

The binding mechanism of a new and possible drug candidate pyrazoline derivative compound K4 and bovine serum albumin (BSA) was investigated in buffer solution (pH 7.4) using ultraviolet–visible light absorption and steady‐state and synchronous fluorescence techniques. The fluorescence intensity of BSA was quenched in the presence of K4 . The quenching process between BSA and K4 was examined at four different temperatures. Decrease of the quenching constants calculated using the Stern–Volmer equation and at increasing temperature suggested that the interaction BSA– K4 was realized through a static quenching mechanism. Synchronous fluorescence measurements suggested that K4 bounded to BSA at the tryptophan region. Fourier transform infrared spectroscopy results showed that there was no significant change in polarity around the tryptophan residue The forces responsible for the BSA– K4 interaction were examined using thermodynamic parameters. In this study, the calculated negative value of ΔG, the negative value of ΔH and the positive value of ΔS pointed to the interaction being through spontaneous and electrostatic interactions that were dominant for our cases. This study provides a very useful in vitro model to researchers by mimicking in vivo conditions to estimate interactions between a possible drug candidate or a drug and body proteins.     

Mechanistic and conformational studies on the interaction of sulfamethazine with human immunoglobulin G by molecular modeling and multi‐spectroscopic approach in vitro

Binding interaction of sulfamethazine (SMZ) with human immunoglobulin G (HIgG) has been explored under physiological conditions. The interaction mechanism was firstly predicted through molecular modeling which showed that several hydrogen bonds participated in stabilizing the SMZ ? HIgG complex. Fluorescence spectroscopy, ultraviolet–visible (UV–vis) light absorption and circular dichroism (CD) spectroscopy were used to analyze the binding site, binding constants and effects of SMZ on HIgG stability and secondary structure. The binding parameters and thermodynamic parameters at different temperatures for the reaction have been calculated according to the Scatchard, Sips and Van 't Hoff equations, respectively. Experimental results showed that the quenching mechanism was a static quenching and there was one independent class of binding site on HIgG for SMZ during their interaction. The thermodynamic parameters of the reaction, namely standard enthalpy ΔH⁰ and entropy ΔS⁰, had been calculated to be ?19.12 kJ · mol^?1 and 20.22 J · mol^?1 · K^?1, respectively, which meant that the electrostatic interaction was the predominant intermolecular force in stabilizing the SMZ ? HIgG complex. Moreover, the conformational changes of HIgG in the presence of SMZ were confirmed by three‐dimensional fluorescence spectroscopy, UV–vis absorption spectroscopy and CD spectroscopy. Copyright © 2014 John Wiley & Sons, Ltd.     

Study on the interaction between anti‐tuberculosis drug ethambutol and bovine serum albumin: multispectroscopic and cyclic voltammetric approaches

The binding of bovine serum albumin (BSA) to ethambutol (EMB) was investigated using spectroscopic methods, viz., fluorescence, Fourier transform infrared (FTIR), ultraviolet (UV)/vis absorption and cyclic voltammetry techniques. Spectroscopic analysis of the emission quenching at different temperatures revealed that the quenching mechanism of serum albumin by EMB is static, which was also confirmed by lifetime measurements. The number of binding sites, n, and binding constant, K, were obtained at various temperatures. The distance, r, between EMB and the protein was evaluated according to the Förster energy transfer theory. Based on displacement experiments using site probes, viz., warfarin, ibuprofen and digitoxin, the site of binding of EMB in BSA was proposed to be Sudlow's site I. The effect of EMB on the conformation of BSA was analyzed by using synchronous fluorescence spectra (SFS) and 3D fluorescence spectra. The results of fluorescence, UV/vis absorption and FTIR spectra showed that the conformation of BSA was changed in the presence of EMB. The thermodynamic parameters including enthalpy change (ΔH⁰), entropy change (ΔS⁰) and free energy change (ΔG⁰) for BSA–EMB were calculated according to the van't Hoff equation and are discussed.