Binding of teicoplanin and vancomycin to bovine serum albumin in vitro: a multispectroscopic approach and molecular modeling

In this paper, the binding properties of teicoplanin and vancomycin to bovine serum albumin (BSA) were investigated using fluorescence quenching, synchronous fluorescence, Fourier transform infrared (FTIR), circular dichroism (CD) and UV–vis spectroscopic techniques and molecular docking under simulative physiological conditions. The results obtained from fluorescence quenching data revealed that the drug–BSA interaction altered the conformational structure of BSA. Meanwhile, the 3D fluorescence, CD, FTIR and UV–vis data demonstrated that the conformation of BSA was slightly altered in the presence of teicoplanin and vancomycin, with different reduced α‐helical contents. The binding distances for the drug–BSA system were provided by the efficiency of fluorescence resonance energy transfer (FRET). Furthermore, the thermodynamic analysis implied that hydrogen bond and van der Waals' forces were the main interaction for the drug–BSA systems, which agreed well with the results from the molecular modeling study. The results obtained herein will be of biological significance in future toxicological and pharmacological investigation. Copyright © 2013 John Wiley & Sons, Ltd.     

Molecular interactions of thymol with bovine serum albumin: Spectroscopic and molecular docking studies

Thymol is the main monoterpene phenol present in the essential oils which is used in the food industry as flavoring and preservative agent. In this study, the interaction of thymol with the concentration range of 1 to 6 μM and bovine serum albumin (BSA) at fixed concentration of 1 μM was investigated by fluorescence, UV‐vis, and molecular docking methods under physiological‐like condition. Fluorescence experiments were performed at 5 different temperatures, and the results showed that the fluorescence quenching of BSA by thymol was because of a static quenching mechanism. The obtained binding parameters, K, were in the order of 10⁴ M^?1, and the binding number, n, was approximately equal to unity indicating that there is 1 binding site for thymol on BSA. Calculated thermodynamic parameters for enthalpy (ΔH), entropy (ΔS), and Gibb's free energy (ΔG) showed that the reaction was spontaneous and hydrophobic interactions were the main forces in the binding of thymol to BSA. The results of UV‐vis spectroscopy and Arrhenius' theory showed the complex formation in the interaction of thymol and BSA. Negligible conformational changes in BSA by thymol were observed in fluorescence experiments, and the same results were also obtained from UV‐vis studies. Results of molecular docking indicated that the subdomain IA of BSA was the binding site for thymol.     

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.     

Interaction of piroxicam with bovine serum albumin investigated by spectroscopic,calorimetric and computational molecular methods

Abstract

The binding of drugs to serum proteins is governed by weak non-covalent forces. In this study, the nature and magnitude of the interactions between piroxicam (PRX) and bovine serum albumin (BSA) was assessed using spectroscopic, calorimetric and computational molecular methods. The fluorescence data revealed an atypical behavior during PRX and BSA interaction. The quenching process of tryptophan (Trp) by PRX is a dual one (approximately equal static and dynamic quenched components). The FRET results indicate that a non-radiative transfer of energy occurred. The association constant and the number of binding sites indicate moderate PRX and BSA binding. The competitive binding study indicates that PRX is bound to site I from the hydrophobic pocket of subdomain IIA of BSA. The synchronous spectra showed that the microenvironment around the BSA fluorophores and protein conformation do not change considerably. The Trp lifetimes revealed that PRX mainly quenches the fluorescence of Trp-213 situated in the hydrophobic domain. The CD and DSC investigation show that addition of PRX stabilizes the protein structure. ITC results revealed that BSA-PRX binding involves a combination of electrostatic, hydrophobic and hydrogen interactions. The analysis of the computational data is consistent with the experimental results. This thorough investigation of the PRX-BSA binding may provide support for other studies concerning moderate affinity drugs with serum protein.

Communicated by Ramaswamy H. Sarma     

Multispectroscopic exploration and molecular docking analysis on interaction of eriocitrin with bovine serum albumin

Eriocitrin is a flavanone glycoside, which exists in lemon or lime citrus fruits. It possesses antioxidant, anticancer, and anti‐allergy activities. In order to investigate the pharmacokinetics and pharmacological mechanisms of eriocitrin in vivo, the interaction between eriocitrin and bovine serum albumin (BSA) was studied under the simulated physiological conditions by multispectroscopic and molecular docking methods. The results well indicated that eriocitrin and BSA formed a new eriocitrin‐BSA complex because of intermolecular interactions, which was demonstrated by the results of ultraviolet‐visible (UV‐vis) absorption spectra. The intrinsic fluorescence of BSA was quenched by eriocitrin, and static quenching was the quenching mechanism. The number of binding sites (n) and binding constant (K_b) at 310 K were 1.22 and 2.84 × 10⁶ L mol^?1, respectively. The values of thermodynamic parameters revealed that the binding process was spontaneous, and the main forces were the hydrophobic interaction. The binding distance between eriocitrin and BSA was 3.43 nm. In addition, eriocitrin changed the conformation of BSA, which was proved by synchronous fluorescence and circular dichroism (CD) spectra. The results of site marker competitive experiments suggested that eriocitrin was more likely to be inserted into the subdomain IIA (site I), which was further certified by molecular docking studies.     

Combined spectroscopies and molecular docking approach to characterizing the binding interaction of enalapril with bovine serum albumin

The binding interaction between bovine serum albumin (BSA) and enalapril (ENPL) at the imitated physiological conditions (pH = 7.4) was investigated using UV–vis absorption spectroscopy (UV–vis), fluorescence emission spectroscopy (FES), synchronous fluorescence spectroscopy (SFS), Fourier transform infrared spectroscopy (FT‐IR), circular dichroism (CD) and molecular docking methods. It can be deduced from the experimental results from the steady‐state fluorescence spectroscopic titration that the intrinsic BSA fluorescence quenching mechanism induced by ENPL is static quenching, based on the decrease in the BSA quenching constants in the presence of ENPL with increase in temperature and BSA quenching rates >10¹⁰ L mol^?1 sec^?1. This result indicates that the ENPL–BSA complex is formed through an intermolecular interaction of ENPL with BSA. The main bonding forces for interaction of BSA and ENPL are van der Waal's forces and hydrogen bonding interaction based on negative values of Gibbs free energy change (ΔG ⁰), enthalpic change (ΔH ⁰) and entropic change (ΔS ⁰). The binding of ENPL with BSA is an enthalpy‐driven process due to |ΔH °| > |T ΔS °| in the binding process. The results of competitive binding experiments and molecular docking confirm that ENPL binds in BSA sub‐domain IIA (site I) and results in a slight change in BSA conformation, but BSA still retains its α‐helical secondary structure.     

Intermolecular interaction of fosinopril with bovine serum albumin (BSA): The multi‐spectroscopic and computational investigation

The intermolecular interaction of fosinopril, an angiotensin converting enzyme inhibitor with bovine serum albumin (BSA), has been investigated in physiological buffer (pH 7.4) by multi‐spectroscopic methods and molecular docking technique. The results obtained from fluorescence and UV absorption spectroscopy revealed that the fluorescence quenching mechanism of BSA induced by fosinopril was mediated by the combined dynamic and static quenching, and the static quenching was dominant in this system. The binding constant, K_b, value was found to lie between 2.69 × 10³ and 9.55 × 10³ M^?1 at experimental temperatures (293, 298, 303, and 308 K), implying the low or intermediate binding affinity between fosinopril and BSA. Competitive binding experiments with site markers (phenylbutazone and diazepam) suggested that fosinopril preferentially bound to the site I in sub‐domain IIA on BSA, as evidenced by molecular docking analysis. The negative sign for enthalpy change (ΔH⁰) and entropy change (ΔS⁰) indicated that van der Waals force and hydrogen bonds played important roles in the fosinopril‐BSA interaction, and 8‐anilino‐1‐naphthalenesulfonate binding assay experiments offered evidence of the involvements of hydrophobic interactions. Moreover, spectroscopic results (synchronous fluorescence, 3‐dimensional fluorescence, and Fourier transform infrared spectroscopy) indicated a slight conformational change in BSA upon fosinopril interaction.     

Probing the behavior of bovine serum albumin upon binding to atenolol: insights from spectroscopic and molecular docking approaches

Molecular interaction of atenolol, a selective β₁ receptor antagonist with the major carrier protein, bovine serum albumin (BSA), was investigated under imitated physiological conditions (pH 7.4) by means of fluorescence spectroscopy, UV absorption spectroscopy, Fourier transform infrared spectroscopy (FT-IR), and molecular modeling studies. The steady-state fluorescence spectra manifested that static type, due to formation of the atenolol-BSA complex, was the dominant mechanism for fluorescence quenching. The characteristic information about the binding interaction of atenolol with BSA in terms of binding constant (K_b) were determined by the UV–vis absorption titration, and were found to be in the order of 10³ M^?1 at different temperatures, indicating the existence of a weak binding in this system. Thermodynamic analysis revealed that the binding process was primarily mediated by van der Waals force and hydrogen bonds due to the negative sign for enthalpy change (ΔH⁰), entropy change (ΔS⁰). The molecular docking results elucidated that atenolol preferred binding on the site II of BSA according to the findings observed in competitive binding experiments. Moreover, via alterations in synchronous fluorescence, three-dimensional fluorescence and FT-IR spectral properties, it was concluded that atenolol could arouse slight configurational and micro-environmental changes of BSA.     

Combined experimental/computational aspects for the molecular interaction of empagliflozin with bovine serum albumin: Quantification and application to human plasma

Empagliflozin (EMP) is an oral antihyperglycemic agent for type 2 diabetic patients. The molecular binding of EMP to bovine serum albumin (BSA) was elucidated by a combined experimental/computational approach to fulfil the pharmacokinetics and pharmacodynamics gaps of the cited drug for further development. Fluorescence, synchronous, and three-dimensional fluorescence spectroscopy verified that EMP quenched BSA native fluorescence through a dual static/dynamic mechanism that was further supported by Fӧrster resonance energy transfer and ultraviolet absorption spectroscopy. Fourier transform infrared spectroscopy revealed the conformational variations in BSA secondary structure induced by EMP. Thermodynamic properties of the BSA–EMP complex were also investigated, and the hydrophobic interactions' role in the binding process was demonstrated by the computed enthalpy (ΔH = 6.558 kJ mol⁻¹) and entropy (ΔS = 69.333 J mol⁻¹ K⁻¹). Gibbs free energy (ΔG) values were negative at three distinct temperatures, illuminating the spontaneity of this interaction. In addition, molecular docking studies depicted the optimal fitting of EMP to BSA on Site I (sub-domain IIA) through three hydrogen bonds. Additionally, and based on the quenching effect of EMP on BSA fluorescence, this study suggests a simple validated spectrofluorometric method for the quantitation of the studied drug in bulk form and human plasma samples with reasonable recoveries (96.99–103.10%).     

Exploring the interaction of the photodynamic therapeutic agent thionine with bovine serum albumin: multispectroscopic and molecular docking studies

This study explores the binding interaction of thionine (TH) with bovine serum albumin (BSA) under physiological conditions (pH 7.40) using absorption, emission, synchronous emission, circular dichroism (CD) and three‐dimensional (3D) emission spectral studies. The results of emission titration experiments revealed that TH strongly quenches the intrinsic emission of BSA via a static quenching mechanism. The apparent binding constant (K) and number of binding sites (n) were calculated as 2.09 × 10⁵ dm³/mol and n~1, respectively. The negative free energy change value for the BSA–TH system suggested that the binding interaction was spontaneous and energetically favourable. The results from absorption, synchronous emission, CD and 3D emission spectral studies demonstrated that TH induces changes in the microenvironment and secondary structure in BSA. Site marker competitive binding experiments revealed that the binding site of TH was located in subdomain IIA (Sudlow site I) of BSA. The molecular docking study further substantiates Sudlow site I as the preferable binding site of TH in BSA. Copyright © 2014 John Wiley & Sons, Ltd.     

Binding of fexofenadine hydrochloride to bovine serum albumin: structural considerations by spectroscopic techniques and molecular docking

The binding interaction of peripheral H₁ receptor antagonist drug, fexofenadine hydrochloride to bovine serum albumin (BSA) is investigated by fluorescence spectroscopy in combination with UV-absorption spectroscopy under physiological conditions. The Stern–Volmer plots at different temperatures and the steady state fluorescence suggested a static type of interaction between fexofenadine and BSA. Binding constants were determined to provide a measure of the binding affinity between fexofenadine and BSA. It was found that BSA has one binding site for fexofenadine. On the basis of the competitive site marker experiments and thermodynamic results, it was considered that fexofenadine was primarily bound to the site I of BSA mainly by hydrogen bond and van der Waals force. Utilising Förster resonance energy transfer the distance, r between the donor, BSA and acceptor fexofenadine was obtained. Furthermore, the results of circular dichroism and synchronous fluorescence spectrum indicated that the secondary structure of BSA was changed in the presence of fexofenadine. Molecular docking was applied to further define the interaction of fexofenadine with BSA.     

Probing the binding interaction of AKR with human serum albumin by multiple fluorescence spectroscopy and molecular modeling

Human serum albumin (HSA) is the major transport protein affording endogenous and exogenous substances in plasma. It can affect the behavior and efficacy of chemicals in vivo through the binding interaction. AKR (3-O-α-l-arabinofuranosyl-kaempferol-7-O-α-l-rhamnopyranoside) is a flavonoid diglycoside with modulation of estrogen receptors (ERs). Herein, we investigated the binding interaction between AKR and HSA by multiple fluorescence spectroscopy and molecular modeling. As a result, AKR specifically binds in site I of HSA through hydrogen bonds, van der Waals force, and electrostatic interaction. The formation of AKR–HSA complex in binding process is spontaneously exothermic and leads to the static fluorescence quenching through affecting the microenvironment around the fluorophores. The complex also affects the backbone of HSA and makes AKR access to fluorophores. Molecular modeling gives the visualization of the interaction between AKR and HSA as well as ERs. The affinity of AKR with HSA is higher than the competitive site marker Warfarin. In addition, docking studies reveal the binding interaction of AKR with ERs through hydrogen bonds, van der Waals force, hydrophobic, and electrostatic interactions. And AKR is more favorable to ERβ. These results unravel the binding interaction of AKR with HSA and mechanism as an ERs modulator.     

Explication of bovine serum albumin binding with naphthyl hydroxamic acids using a multispectroscopic and molecular docking approach along with its antioxidant activity

In the present investigation, the protein‐binding properties of naphthyl‐based hydroxamic acids (HAs), N‐1‐naphthyllaurohydroxamic acid ( 1 ) and N‐1‐naphthyl‐p‐methylbenzohydroxamic acid ( 2 ) were studied using bovine serum albumin (BSA) and UV–visible spectroscopy, fluorescence spectroscopy, diffuse reflectance spectroscopy–Fourier transform infrared (DRS–FTIR), circular dichroism (CD), and cyclic voltammetry along with computational approaches, i.e. molecular docking. Alteration in the antioxidant activities of compound 1 and compound 2 during interaction with BSA was also studied. From the fluorescence studies, thermodynamic parameters such as Gibb's free energy (ΔG), entropy change (ΔS) and enthalpy change (ΔH) were calculated at five different temperatures (viz., 298, 303, 308, 313 or 318 K) for the HAs–BSA interaction. The results suggested that the binding process was enthalpy driven with dominating hydrogen bonds and van der Waals’ interactions for both compounds. Warfarin (WF) and ibuprofen (IB) were used for competitive site‐specific marker binding interaction and revealed that compound 1 and compound 2 were located in subdomain IIA (Sudlow's site I) on the BSA molecule. Conclusions based on above‐applied techniques signify that various non‐covalent forces were involved during the HAs–BSA interaction. Therefore the resulted HAs–BSA interaction manifested its effect in transportation, distribution and metabolism for the drug in the blood circulation system, therefore establishing HAs as a drug‐like molecule.     

Probing into the binding interaction between medroxyprogesterone acetate and bovine serum albumin (BSA): spectroscopic and molecular docking methods

To further understand the mechanism of action and pharmacokinetics of medroxyprogesterone acetate (MPA), the binding interaction of MPA with bovine serum albumin (BSA) under simulated physiological conditions (pH 7.4) was studied using fluorescence emission spectroscopy, synchronous fluorescence spectroscopy, circular dichroism and molecular docking methods. The experimental results reveal that the fluorescence of BSA quenches due to the formation of MPA–BSA complex. The number of binding sites (n) and the binding constant for MPA–BSA complex are ~1 and 4.6 × 10³ M^?1 at 310 K, respectively. However, it can be concluded that the binding process of MPA with BSA is spontaneous and the main interaction forces between MPA and BSA are van der Waals force and hydrogen bonding interaction due to the negative values of ΔG⁰, ΔH⁰ and ΔS⁰ in the binding process of MPA with BSA. MPA prefers binding on the hydrophobic cavity in subdomain IIIA (site II′′) of BSA resulting in a slight change in the conformation of BSA, but BSA retaining the α‐helix structure. Copyright © 2016 John Wiley & Sons, Ltd.     

Combined spectroscopies and molecular docking approach to characterizing the binding interaction between lisinopril and bovine serum albumin

To further understand the mode of action and pharmacokinetics of lisinopril, the binding interaction of lisinopril with bovine serum albumin (BSA) under imitated physiological conditions (pH 7.4) was investigated using fluorescence emission spectroscopy, synchronous fluorescence spectroscopy, Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD) and molecular docking methods. The results showed that the fluorescence quenching of BSA near 338 nm resulted from the formation of a lisinopril–BSA complex. The number of binding sites (n) for lisinopril binding on subdomain IIIA (site II) of BSA and the binding constant were ~ 1 and 2.04 × 10⁴ M^–1, respectively, at 310 K. The binding of lisinopril to BSA induced a slight change in the conformation of BSA, which retained its α‐helical structure. However, the binding of lisinopril with BSA was spontaneous and the main interaction forces involved were van der Waal's force and hydrogen bonding interaction as shown by the negative values of ΔG⁰, ΔH⁰ and ΔS⁰ for the binding of lisinopril with BSA. It was concluded from the molecular docking results that the flexibility of lisinopril also played an important role in increasing the stability of the lisinopril–BSA complex. Copyright © 2015 John Wiley & Sons, Ltd.     

Investigation of new indole derivatives of bovine serum albumin using spectroscopic and molecular docking techniques

We investigated the interaction of two derivatives of bis (indolyl) methane with bovine serum albumin (BSA) using spectroscopic and molecular docking calculations. Fluorescence quenching measurements revealed that the quenching mechanism was static. F?rster energy transfer measurements, synchronous fluorescence spectroscopy and docking studies demonstrated that both bis(indolyl)methanes bound to the Trp residues of BSA. The docking study confirmed that both bis(indolyl)methanes form hydrogen bonds and van der Waals interactions with BSA. Our molecular docking study indicated that the compounds are located within the binding pocket of subdomains IIB and IB of BSA. Fourier transform infrared spectroscopy demonstrated that both bis(indolyl)methane derivatives can interact with BSA and can affect the secondary structure of BSA.     

Synthesis of F16 conjugated with 5‐fluorouracil and biophysical investigation of its interaction with bovine serum albumin by a spectroscopic and molecular modeling approach

5‐Fluorouracil (5‐FU) has been widely used as a chemotherapy agent in the treatment of many types of solid tumors. Investigation of its antimetabolites led to the development of an entire class of fluorinated pyrimidines. However, the toxicity profile associated with 5‐FU is significant and includes diarrhea, mucositis, hand–foot syndrome and myelosuppression. In aiming at reducing of the side effects of 5‐FU, we have designed and synthesized delocalized lipophilic cations (DLCs) as a vehicle for the delivery of 5‐FU. DLCs accumulate selectively in the mitochondria of cancer cells because of the high mitochondrial transmembrane potential (ΔΨ_m). Many DLCs exhibited anti‐cancer efficacy and were explored as potential anti‐cancer drugs based on their selective accumulation in the mitochondria of cancer cells. F16, the DLC we used as a vehicle, is a small molecule that selectively inhibits tumor cell growth and dissipates mitochondrial membrane potential. The binding of the conjugate F16–5‐FU to bovine serum albumin (BSA) was investigated using spectroscopic and molecular modeling approaches. Fluorescence quenching constants were determined using the Stern–Volmer equation to provide a measure of the binding affinity between F16–5‐FU and BSA. The activation energy of the interaction between F16–5‐FU and BSA was calculated and the unusually high value was discussed in terms of the special structural block indicated by the molecular modeling approach. Molecular modeling showed that F16–5‐FU binds to human serum albumin in site II, which is consistent with the results of site‐competitive replacement experiments. It is suggested that hydrophobic and polar forces played important roles in the binding reaction, in accordance with the results of thermodynamic experiments. Copyright © 2012 John Wiley & Sons, Ltd.     

Interaction between fasudil hydrochloride and bovine serum albumin: spectroscopic study

The interaction between fasudil hydrochloride (FSD) and bovine serum albumin (BSA) was investigated using fluorescence and ultraviolet spectroscopy under imitated physiological conditions. The Stern–Volmer quenching model has been successfully applied and the results revealed that FSD could quench the intrinsic fluorescence of BSA effectively via static quenching. The binding constants and binding sites for the BSA–FSD system were evaluated. The corresponding thermodynamic parameters obtained at different temperatures indicated that hydrophobic force played a major role in the interaction of FSD and BSA. The distance between the donor (BSA) and the acceptor (FSD) was obtained according to fluorescence resonance energy transfer (FRET). Synchronous fluorescence spectroscopy and FT‐IR spectra showed that the conformation of BSA was changed in the presence of FSD. Copyright © 2015 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.