Investigation on the interaction between triclosan and bovine serum albumin by spectroscopic methods

Abstract

Multi-spectroscopic and molecular docking methods were used to study the interaction between triclosan (TCS) and bovine serum albumin (BSA). The results indicated that the fluorescence quenching of BSA by TCS was due to the formation of TCS–BSA complex through static quenching. This result was also demonstrated by time-resolved fluorescence experiment. The binding constants and number of binding sites between TCS and BSA were 1.30?×?10⁵ M^?1 and 1.17 at 298?K, respectively. The thermodynamic parameters were studied in detail which suggested that hydrophobic forces and hydrogen bond played major roles in the TCS–BSA interaction. Moreover, the site marker competitive experiments and docking studies revealed that TCS could bind BSA into site I in subdomain IIA. All the results of UV–vis spectrophotometry, circular dichroism spectroscopy and synchronous fluorescence spectroscopy showed that interaction between TCS and BSA induced conformation changes of BSA.     

Investigations on the interaction of the phototoxic alkaloid coralyne with serum albumins

The interaction of the phototoxic alkaloid coralyne with bovine and human serum albumins (BSA, HSA) was investigated. Absorbance and fluorescence quenching experiments revealed the formation of strong complexes. Based on the binding parameters calculated from Stern-Volmer quenching method, coralyne has higher affinity to BSA (∼10⁵ M⁻¹) compared to HSA (∼10⁴ M⁻¹). Forster resonance energy transfer studies showed that the specific binding distances between Trp (donor) of the proteins and coralyne (acceptor) were 2.95 and 3.10 nm, respectively. The bindings were favored by negative enthalpy and a stronger favorable entropy contribution. The heat capacity values for binding to BSA and HSA were similar, indicating the involvement of similar molecular forces in the complexation. Competitive binding experiments using site markers demonstrated that coralyne binds to site I (subdomain IIA) of both proteins. The secondary structure of the proteins was altered, suggesting a small but definitive partial unfolding on complexation.     

Binding of triclosan to human serum albumin: insight into the molecular toxicity of emerging contaminant

Purpose

The interaction between triclosan (TCS) and human serum albumin (HSA) was investigated in order to obtain the binding mechanism, binding constant, the type of binding force, the binding distance between the donor and acceptor, and the effect of TCS on the conformation change of HSA.

Methods

A HSA solution was added to the quartz cell and then titrated by successive addition of TCS. The fluorescence quenching spectra and synchronous spectra were recorded with the excitation and emission slits of the passage of band set at 10 and 20 nm. Three-dimensional fluorescence spectra of HSA were recorded before and after the addition of TCS. The capillary electrophoresis was conducted with the pressure injection mode at 0.5 psi for 5 s, separation under 25 kV, and detection at 214 nm.

Results

Fluorescence data indicated the fluorescence quenching of HSA by TCS was static quenching, and the quenching constants (K _a ) were 1.14?×?10⁵, 8.75?×?10⁴, 6.67?×?10⁴, and 5.00?×?10⁴ at 293, 298, 303, and 309 K, respectively. The thermodynamic parameters, enthalpy change (??H) and entropy change (??S) for the interaction were calculated to be ?37.9 kJ mol^?1 and 32.6 J?mol^?1 K^?1. The binding distance between TCS and tryptophan residues of HSA was obtained to be 1.81 nm according to F??rster nonradioactive energy transfer theory. The UV-Vis absorption spectroscopy, the synchronous fluorescence spectroscopy, three-dimensional fluorescence spectroscopy, and circular dichroism spectroscopy revealed the alterations of HSA secondary structure in the presence of TCS. Finally, the interaction between TCS and HSA was further confirmed by capillary electrophoresis.

Conclusions

TCS was bound to HSA to form the TCS-HSA complex, with the binding distance of 1.81 nm. Hydrophobic interaction and hydrogen bond were dominated in the binding. TCS could change the secondary conformation of HSA. This work provides an insight into noncovalent interaction between emerging pollutants and protein, helping to elucidate the toxic mechanism of such pollutants.     

Investigation of the binding and simultaneous quantifications of propanil and bromoxynil herbicide concentrations in human serum albumin

This study reported the use of UV–visible and fluorescence spectroscopy and partial-least-square (PLS) multivariate regression for accurate and simultaneous quantifications of two widely used herbicides, propanil, 3′,4′-dichloropropionanilide (PPL) and bromoxynil, 3,5-dibromo-4-hydroxybenzonitrile (BXL) in human serum albumin (HSA) at physiological conditions. The binding affinity and thermodynamic properties of PPL-HSA and BXL-HSA complexes were also investigated. Partial-least-square (PLS) regression was used to collate the variability in the absorption or emission spectra of PPL-HSA and BXL-HSA complexes with PPL and/or BXL concentrations in HSA samples. The binding constants of 7.66× 10⁸ M^?1 for PPL-HSA and 4.88× 10⁶ M^?1 for BXL-HSA complexes were calculated at physiological conditions (temperature, 310 K; pH 7.4). Thermodynamic parameter values: enthalpy (ΔH) (13.99 kJ mol^?1), entropy (ΔS) (0.078 kJ mol^?1 K^?1), and Gibbs free energy (ΔG) (?10.19 kJ mol^?1) were determined for PPL-HSA complexation at physiological conditions. However, differences in thermodynamic property values of: ΔH (?214.3 kJ mol^?1), ΔS (?0.563 kJ mol^?1 K^?1), and ΔG (?39.70 kJ mol^?1) were observed for BXL–HSA complexes. The binding constants and negative ΔG values indicated strong binding affinity and thermodynamically favorability of PPL–HSA and BXL–HSA complex formation. Results of the PLS regression calibration showed good linearity (R² ≥ 0.998289), high sensitivity, and impressive low limit-of-detections (LODs) of 1.38× 10^?8 M for PPL and 1.68× 10^?8 M for BXL that are comparable and/or lower than many previously reported LODs for herbicide and pesticide analyses. Most importantly, PLS regression is capable of simultaneous quantifications of PPL and BXL concentrations in HSA samples with good accuracy and low errors of 3.66%. UV–visible spectrophotometers and spectrofluorometers are fairly inexpensive, easy to use, and are readily available in almost every laboratory, making this protocol excellent and affordable for routine analysis of weed/pest control chemical residues in humans. The results of this study are significant and remarkable that will provide critical insight into the binding mechanism of herbicide toxicity in humans and non-target organisms, which are of special interest in the area of biomedical study, environmental risk assessment, and ecotoxicology.     

Pesticide binding and urea-induced controlled release applications with calixarene naphthalimide molecules by host–guest complexation

ABSTRACT

Three novel calix[4]arene molecule-based 1,8 naphthalimide fluoroionophore for the selective determination of kesoxim-methyl were synthesized and used in pesticide binding studies. The possible interaction between pesticides and fluorescent calix[4]arene molecules was monitored by UV/Vis absorption and fluorescence spectroscopy. When compared the studied pesticides, kesoxim-methyl was strongly quenched the fluorescence intensity of upper rim-modified calix[4]arene. UV and fluorescence titration experiments were also studied to determine both the quenching mechanism and stoichiometric ratio consisted in complex formation. Furthermore, pesticide release experiments were also performed with a fertilizing agent as urea by using fluorescence spectroscopy technique.     

Potential toxicity of amphenicol antibiotic: binding of chloramphenicol to human serum albumin

Antibiotics are widely used in daily life but their abuse has posed a potential threat to human health. To evaluate the toxicity of chloramphenicol (CAP) at the protein level, the interaction between CAP and human serum albumin (HSA) was investigated by fluorescence, Ultraviolet–visible (UV–Vis) absorption, Fourier transform infrared (FT-IR) spectroscopy and molecular docking methods. Fluorescence data revealed that the fluorescence quenching of HSA by CAP was the result of the formation of CAP–HSA complex, and the binding constant was determined to be 3.196?×?10⁴ L mol^?1 at 310 K. The thermodynamic determination indicated that the interaction was driven by enthalpy change and entropy change together, where the multiple hydrogen bonds (CAP and the residues Arg 222 and His 242 of HSA) and van der Waals forces were the dominant binding force. The site marker competition revealed that CAP bound into sub-domain IIA of HSA. The binding of CAP induced the drastic reduction in α-helix conformation and the significant enhancement in β-sheet conformation of HSA. Molecular docking study further confirmed the binding mode obtained by experimental study. This work provides a new quantitative evaluation method for antibiotics to cause the protein damage.     

Investigation of metal ions binding of humic substances using fluorescence emission and synchronous‐scan spectroscopy

Abstract

The binding site interactions of IHSS humic substances, Suwannee River Humic Acid, Suwannee River Fulvic Acid, Nordic Fulvic Acid, and Aldrich Humic Acid with various metals ions and a herbicide, methyl viologen were investigated using fluorescence emission and synchronous‐scan spectroscopy. The metal ions used were, Fe(III), Cr(III), Cr(VI), Pb(II), Cu(II) and Ni(II). Stern‐Volmer constants, K_SV for these quenchers were determined at pH 4 and 8 using an ionic strength of 0.1M. For all four humic substances, and at both pH studied, Fe(III) was found to be the most efficient quencher. Quenching efficiency was found to be 3–10 times higher at pH 8. The bimolecular quenching rate constants were found to exceed the maximum considered for diffusion controlled interactions, and indicate that the fluorophore and quencher are in close physical association. Synchronous‐scan spectra were found to change with pH and provided useful information on binding site interactions between humic substances and these quenchers.     

Study on the mechanism of action between dimethyl phthalate and herring sperm DNA at molecular level

Dimethyl phthalate (DMP), a typical phthalic acid ester, is widespread in the environment and causes extensive concern due to its adverse effects on human health. To understand the genotoxicity of DMP at molecular level, the toxic interaction of DMP with herring sperm (hs) deoxyribonucleic acid (DNA; hs-DNA) was investigated in vitro under simulated physiological conditions using multi-spectroscopic techniques and a molecular modeling method. The results of Ultraviolet-Visible absorption spectroscopy, fluorescence emission spectroscopy, and circular dichroism spectra indicated that DMP interacts with hs-DNA in a groove-binding mode that changes the double helical structure of DNA. The binding constant and the number of binding sites calculated from the fluorescence quenching data were 565.718 L mol^?1 and 0.7872, respectively. A molecular modeling study revealed that DMP tends to bind with DNA in the A-T-rich regions of minor groove and that hydrogen bonding and van der Waals forces play main roles in the interaction. This research can help to elucidate the mechanism of DMP toxicity in vivo.     

Thermodynamics of binding of cadmium to bovine serum albumin

The binding isotherm of Cd2+ ion to bovine serum albumin (BSA) has been investigated by microcalorimetry at 310.15 K and pH 7.0. The thermodynamic parameters of the binding reaction have been determined, and the stoichiometry of the complex is 2:1, indicating that there exist two identical binding sites of BSA with Cd2+ ion. The value of deltarHthetam is -28.4+/-1.7 kJ mol(-1), the free energy of binding deltarGthetam is -25.2 kJ mol(-1), and the entropy of binding deltarSthetam is -10.3 J mol(-1) K(-1). The negative deltarHthetam and deltarSthetam values are observed for the binding reaction of Cd2+ ion and BSA, suggesting that the binding reaction is mainly enthalpy-driven and the entropy is unfavorable for it.     

The complexation of Cu(II) by anthropogenic fulvic acids extracted from composted urban and livestock wastes

Abstract

The fulvic acid (fua) fractions of two samples of composted solid wastes [urban (urfua) and livestock (lsfua) wastes], commercialized to be used in agriculture as organic correctives or fertilizers, were analyzed for their affinity towards Cu(II) at pH=6. Molecular fluorescence spectroscopy (synchronous mode) was used to monitor the quenching caused by the complexation upon addition of Cu(II) to fua. Spectral data were preprocessed by a chemometric self‐modeling mixture analysis method (SIMPLISMA) to detect the number of different types of fluorescent binding sites that exist in each fua, their spectra and the corresponding quenching profiles [fluorescence intensity as function of the total Cu(II) concentration]. From the analysis of the quenching profiles, the amount of binding sites (Cl) and the corresponding conditional stability constants (K') were calculated. Both fua samples have approximately Cl = 0.21 mmol/g and the logarithms of K’ are 4.21(3) and 4.51(8), respectively for urfua and lsfua. The differences detected between these fua samples and those extracted from natural soils can be attributed mainly to the relatively small humification extent of the present anthropogenic fua samples.     

Potential use of Caulerpa fastigiata biomass for removal of lead: kinetics, isotherms, thermodynamic, and characterization studies

The present study attempts to analyze the biosorption trend of biosorbent Caulerpa fastigiata (macroalgae) biomass for removal of toxic heavy metal ion Pb (II) from solution as a function of initial metal ion concentration, pH, temperature, sorbent dosage, and biomass particle size. The sorption data fitted with various isotherm models and Freundlich model was the best one with correlation coefficient of 0.999. Kinetic study results revealed that the sorption data on Pb (II) with correlation coefficient of 0.999 can best be represented by pseudo-second-order. The biosorption capacity (q _e ) of Pb (II) is 16.11?±?0.32 mg g^?1 on C. fastigiata biomass. Thermodynamic studies showed that the process is exothermic (ΔH° negative). Free energy change (ΔG°) with negative sign reflected the feasibility and spontaneous nature of the process. The SEM studies showed Pb (II) biosorption on selective grains of the biosorbent. The FTIR spectra indicated bands corresponding to –OH, COO^?, –CH, C?=?C, C?=?S, and –C–C– groups were involved in the biosorption process. The XRD pattern of the C. fastigiata was found to be mostly amorphous in nature.     

Interaction of diuron to human serum albumin: Insights from spectroscopic and molecular docking studies

This investigation was undertaken to determine the interaction of diuron with human serum albumin (HSA) was studied by monitoring the spectral behavior of diuron-HSA system. The fluorescence of HSA at 340 nm excited at 230 nm was obviously quenched by diuron due to dynamic collision and the quenching constant was of the order of 10⁴ L mol^?1 at 310 K. However, no fluorescence quenching was observed when excited at 280 nm. Thermodynamic investigations revealed that the combination between diuron and HSA was entropy driven by predominantly hydrophobic interactions. The binding of diuron induced the drastic reduction in α-helix conformation and the significant enhancement in β-turn conformation of HSA. In addition, both sites marker competition study and molecular modeling simulation evidenced the binding of diuron to HSA primarily took place in subdomain IIIA (Sudlow's site II).     

Fluorescence-based evidence for adsorptive binding of pyrene to effluent dissolved organic matter

Using fluorescence intensity measurements, pyrene interactions with different types of effluent dissolved organic matter (EDOM) originated from treated municipal wastewater are examined. Multiple observations show that fluorescence intensity of pyrene–EDOM solutions is non-linearly related to pyrene concentration, with distinct concave-up dependence. Testing the effect of pyrene concentration on fluorescence intensity of pyrene–EDOM solutions provides a tool to examine whether binding of an organic compound to EDOM follows linear or non-linear isotherm. Possible coupling between static and dynamic quenching effects was addressed while analyzing fluorescence data. Limited number of EDOM binding sites results in a non-linear binding isotherm such that the concept of pyrene “partitioning” between aqueous phase and “bulky” EDOM organic phase is hardly relevant. Maximal EDOM capacity for pyrene binding is estimated approximately as 0.1% w w⁻¹. Examination of the differences between the total fluorescence intensity of pyrene–EDOM solution and the fluorescence intensities of separated constituents (pyrene and EDOM) was used to illustrate the accumulation of pyrene–EDOM complexes and saturation of some EDOM binding sites. Strong interactions between pyrene and EDOM binding sites may result in pyrene distribution coefficients differing at least by a factor 3.5–7 at varying pyrene concentrations.     

A study on the effect of deoxynivalenol on serotonin receptor binding in fig brain membranes

Abstract

Central serotoninergic (5‐hydroxytryptamine, 5HT) pathways are believed to be involved in the mechanisms of anorexia and/or emesis evoked by the trichothecene mycotoxin deoxynivalenol (DON). Using an in vitro membrane receptor binding assay, the competitive potency of DON was investigated against several radioactive ligands that have a high affinity for selective 5HT‐receptor subgroups. Receptor site densities and displacement profiles in twelve selected regions of pig brain were investigated. Overall, DON possessed only minimal efficacy to competently block any of the 5HT‐ligands tested. IC₅₀ values (50% inhibitory concentration) of at least 5 mM DON were required to inhibit binding, and in certain regions concentrations of 100 mM were ineffective. In comparison, several standard 5HT‐antagonists showed 10³‐10⁵ times greater capability than DON to displace binding of these ligands. Because these results indicated DON possesses only weak affinity for the 5HT‐receptor subtypes investigated here, this suggested that in vivo, unless relatively high concentrations of the toxin are present, its pharmacological effects may be mediated by mechanisms other than a functional interaction with serotoninergic receptors at the central level.     

Quantitative Evaluation of Electron Injection Efficiency in Dye-Sensitized TiO<Subscript>2</Subscript> Films

The efficiency of electron injection (Φ_inj) in dye-sensitized nanocrystalline films was studied through transient absorption (TA) and time-resolved microwave conductivity (TRMC) measurements. Here, I show the absolute value of Φ_inj for several dye-sensitized nanocrystalline films and discuss the relationship between Φ_inj and the free energy change (?ΔG _inj) for the injection process. Some systems exhibited lower Φ_inj values even when ?ΔG _inj was sufficiently large to promote electron injection. Recent experimental findings are used to propose possible explanations for this phenomenon. Quantitative evaluation of Φ_inj using TA and TRMC will give us new insights for developing high-performance solar cell devices.     

Quantification of the interactions between Ca,Hg and extracellular polymeric substances (EPS) of sludge

The interactions between metals (Ca²⁺ and Hg²⁺) and extracellular polymeric substances (EPS) extracted from the aerobic and anaerobic sludge in wastewater treatment reactors were investigated using a combination of zeta potential measurement and 3-dimensional excitation–emission matrix (EEM) fluorescence spectroscopy with parallel factor (PARAFAC) analysis. Results show that Ca²⁺ had no substantial effects on the EEM fluorescence spectra of the EPS, but their zeta potentials increased with the increasing Ca²⁺ dosage. However, Hg²⁺ had a significant effect on the EEM fluorescence spectra of the EPS, while their zeta potentials seemed not to be affected by the dose of Hg²⁺. The interactions between Hg²⁺ and EPS were elucidated using the fluorescence quenching with PARAFAC analysis, while the interactions between Ca²⁺ and EPS were evaluated by the zeta potential technique. The binding constants for Hg²⁺ and EPS were two orders of magnitude higher than those for Ca²⁺ and EPS, suggesting that the binding mechanisms between Ca²⁺ and EPS were different from those between Hg²⁺ and EPS. The results might be useful for understanding the roles of EPS in bacterial self-protection against heavy metals and the aggregate formation mechanisms through ionic bridging interactions.     

Investigating interactions of phenanthrene with dissolved organic matter: limitations of Stern-Volmer plot

Although linear binding isotherms of hydrophobic organic chemicals (HOCs) with dissolved organic matter (DOM) are widely reported, several studies showed nonlinear HOC-DOM interactions. This study pointed out that fluorescence static quenching modeling (FSQM), which often uses a Stern-Volmer type plot to process the data from fluorescence quenching experiments, is conceptually different from the classic Stern-Volmer equation. We also emphasized that although linear Stern-Volmer plots are generally observed in literature, it does not necessarily indicate a linear HOC-DOM interaction. According to both mathematical simulation and laboratory sorption experiments in this study, nonlinear interactions could be concealed by the use of Stern-Volmer plot. Moreover, this study tested the two assumptions for applying FSQM to process binding data. Our results showed that binding coefficient (K(DOC)) for phenanthrene is neither independent of free solute concentration, nor DOM concentration, which is a critical limitation for using FSQM in a form of Stern-Volmer equation to examine HOC-DOM interactions. Therefore, the true characteristics of HOC-DOM interactions need to be examined using different ways of experimental design and data processing.     

Quantifying interactions between propranolol and dissolved organic matter (DOM) from different sources using fluorescence spectroscopy

Beta blockers are widely used pharmaceuticals that have been detected in the environment. Interactions between beta blockers and dissolved organic matter (DOM) may mutually alter their environmental behaviors. To assess this potential, propranolol (PRO) was used as a model beta blocker to quantify the complexation with DOM from different sources using the fluorescence quenching titration method. The sources of studied DOM samples were identified by excitation–emission matrix spectroscopy (EEMs) combined with fluorescence regional integration analysis. The results show that PRO intrinsic fluorescence was statically quenched by DOM addition. The resulting binding constants (log K _oc) ranged from 3.90 to 5.20, with the surface-water-filtered DOM samples claiming the lower log K _oc and HA having the highest log K _oc. Log K _oc is negatively correlated with the fluorescence index, biological index, and the percent fluorescence response (P _i,n) of protein-like region (P _I,n) and the P _i,n of microbial byproduct-like region (P _II,n) of DOM EEMs, while it is correlated positively with humification index and the P _i,n of UVC humic-like region (P _III,n). These results indicate that DOM samples from allochthonous materials rich in aromatic and humic-like components would strongly bind PRO in aquatic systems, and autochthonous DOM containing high protein-like components would bind PRO more weakly.     

Characterization of binding site heterogeneity for copper within dissolved organic matter fractions using two-dimensional correlation fluorescence spectroscopy

The heterogeneity of copper binding characteristics for dissolved organic matter (DOM) fractions was investigated based on the fluorescence quenching of the synchronous fluorescence spectra upon the addition of copper and two-dimensional correlation spectroscopy (2D-COS). Hydrophobic acid (HoA) and hydrophilic (Hi) fractions of two different DOM (algal and leaf litter DOM) were used for this study. For both DOM, fluorescence quenching occurred at a wider range of wavelengths for the HoA fractions compared to the Hi fractions. The combined information of the synchronous and asynchronous maps derived from 2D-COS provided a clear picture of the heterogeneous distribution of the copper binding sites within each DOM fraction, which was not readily recognized by a simple comparison of the changes in the synchronous fluorescence spectra upon the addition of copper. For the algal DOM, higher stability constants were exhibited for the HoA versus the Hi fractions. The logarithms of the stability constants ranged from 4.8 to 6.1 and from 4.5 to 5.0 for the HoA and the Hi fractions of the algal DOM, respectively, depending on the associated wavelength and the fitted models. In contrast, no distinctive difference in the binding characteristics was found between the two fractions of the leaf litter DOM. This suggests that influences of the structural and chemical properties of DOM on copper binding may differ for DOM from different sources. The relative difference of the calculated stability constants within the DOM fractions were consistent with the sequential orders interpreted from the asynchronous 2D-COS. It is expected that 2D-COS will be widely applied to other DOM studies requiring detailed information on the heterogeneous nature and subsequent effects under a range of environmental conditions.     

Microwave synthesis and antifungal evaluations of some chalcones and their derived diaryl-cyclohexenones

Microwave irradiation (MWI) of acetophenones and substituted benzaldehydes in water resulted in a “green-chemistry” procedure for the preparation of chalcones (1-14), through base catalyzed Claisen-Schmidt condensation reaction, in good yields. Further 3,5-diaryl-6-carbethoxy-2-cyclohexen-1-ones (1a-14a) were prepared through base catalyzed cyclocondensation of above chalcones with ethylacetoacetate using MWI as the energy source and silica as support. Out of fourteen cyclohexenones, ten (1a, 4a, 5a, 6a, 7a, 9a, 10a, 11a, 12a and 13a) are reported for the first time in literature. The synthesized compounds were characterized using various spectroscopic techniques, viz. (¹H NMR and IR) and screened for their antifungal activity in vitro against Sclerotium rolfsii and Rhizoctonia solani by poisoned food technique. The compounds tested were found to be active against R. solani whereas against S. rolfsii, moderate activity was observed, as evident from LC₅₀ values. The most potent compounds against R. solani were 1-(4-Fluoro-phenyl)-3-phenyl-propenone (13) and 1,3-Diphenyl-propenone (14) having LC₅₀ values of 2.36 and 2.49 mgL^{? 1} respectively (LC₅₀ of Hexaconazole = 1.12 mgL^{? 1}) and against S. rolfsii 3-(4-Fluoro-phenyl)-5-(3-nitro-phenyl)-6-carbethoxy-2-cyclohexen-1-one (12a) was most active having LC₅₀ value of 285 mgL^{? 1}compared to Hexaconazole (LC₅₀ = 1.27 mgL^{? 1}).