A Study of the Interaction Between Malachite Green and Lysozyme by Steady-State Fluorescence

The interaction of a N-methylated diaminotriphenylmethane dye, malachite green, with lysozyme was investigated by fluorescence spectroscopic techniques under physiological conditions. The binding parameters have been evaluated by fluorescence quenching methods. The results revealed that malachite green caused the fluorescence quenching of lysozyme through a static quenching procedure. The thermodynamic parameters like ΔH and ΔS were calculated to be −15.33 kJ mol⁻¹ and 19.47 J mol⁻¹ K⁻¹ according to van’t Hoff equation, respectively, which proves main interaction between malachite green and lysozyme is hydrophobic forces and hydrogen bond contact. The distance r between donor (lysozyme) and acceptor (malachite green) was obtained to be 3.82 nm according to Fӧrster’s theory. The results of synchronous fluorescence, UV/vis and three-dimensional fluorescence spectra showed that binding of malachite green with lysozyme can induce conformational changes in lysozyme. In addition, the effects of common ions on the constants of lysozyme-malachite green complex were also discussed.     

Fluorescence and Electrochemical Sensing of Pesticides Methomyl, Aldicarb and Prometryne by the Luminescent Europium-3-Carboxycoumarin Probe

This work describes the application of time resolved fluorescence in microtiterplates and electrochemical methods on glassy carbon electrode for investigating the interactions of europium-3-carboxycoumarin with pesticides aldicarb, methomyl and prometryne. Stern-volmer studies at different temperatures indicate that static quenching dominates for methomyl, aldicarb and prometryne. By using Lineweaver-Burk equation binding constants were determined at 303 K, 308 K and 313 K. A thermodynamic analysis showed that the reaction is spontaneous with ΔG being negative. The enthalpy ΔH and the entropy ΔS of reactions were all determined. A time-resolved (gated) luminescence-based method for determination of pesticides in microtiterplate format using the long-lived europium-3-carboxycoumarin has been developed. The limit of detection is 4.80, 5.06 and 8.01 μmol L⁻¹ for methomyl, prometryne and aldicarb, respectively. This is the lowest limit of detection achieved so far for luminescent lanthanide-based probes for pesticides. The interaction of the probe with the pesticides has been investigated using cyclic voltammetry (CV), differential pulse polarography (DPP), square wave voltammetry (SWV) and linear sweep voltammetry (LSV) on a glassy carbon electrode in I = 0.1 mol L⁻¹ p-toluenesulfonate at 25 °C. The diffusion coefficients of the reduced species are calculated. The main properties of the electrode reaction occurring in a finite diffusion space are the quasireversible maximum and the splitting of the net SWV peak for Eu(III) ions in the ternary complex formed . It was observed that the increase of the cathodic peak currents using LSV is linear with the increase of pesticides concentration in the range 5 × 10⁻⁷ to 1 × 10⁻⁵ mol L⁻¹. The detection limit (DL) were about 1.01, 2.23 and 1.89 μmolL⁻¹ for aldicarb, methomyl and prometryne, respectively. In order to assess the analytical applicability of the method, the influence of various potentially interfering species was examined. Influence of interfering species on the recovery of 10 μmol L⁻¹ pesticides has been investigated.     

A Coumarin-Based Fluorescent Chemosensor for Zn<Superscript>2+</Superscript> in Aqueous Ethanol Media

A coumarin-based fluorescent chemosensor 1 for Zn²⁺ was designed and synthesized. Compound 1 exhibits lower background fluorescence due to intramolecular photoinduced electron transfer. However, upon mixing with Zn²⁺ in 30% (v/v) aqueous ethanol, a “turn-on” fluorescence emission is observed. The fluorescence emission increases linearly with Zn²⁺ concentration in the range 0.5–10 μmol L⁻¹ with a detection limit of 0.29 μmol L⁻¹. No remarkable emission enhancement was, however, observed for other metal ions. The proposed chemosensor was applied to the determination of Zn²⁺ in water samples with satisfactory results.     

Study on the Interaction Mechanism of Lysozyme and Bromophenol Blue by Fluorescence Spectroscopy

The interaction of lysozyme with bromophenol blue (BPB) in acetate buffer (pH 6.0) was studied by fluorescence quenching method for the first time. It was found that BPB could conspicuously quench the fluorescence of lysozyme by the static quenching process, possibly due to the binding on the active site near Trp62. The binding parameters including the binding constant and the number of binding site were calculated. The thermodynamic parameters ΔH°, ΔS° and ΔG° at different temperatures were obtained. The formation of lysozyme–BPB complex depended on the cooperation of the hydrophobic and electrostatic forces. And the binding average distance between lysozyme and BPB was determined. The effect of common metal ions on the binding constant of lysozyme–BPB was also examined.     

Interaction of Thyroxine with 7 Hydroxycoumarin: A Fluorescence Quenching Study

The interaction between thyroxine hormone and 7 hydroxycoumarin (7HC) was investigated using fluorescence quenching method. The experimental results showed that thyroxine could quench the fluorescence of 7HC by forming the 7HC–thyroxine complex with static quenching. The apparent binding constants (K) between 7HC and thyroxine were determined to be 1.51 × 10⁴ (297 K) and 9.06 × 10³ (310 K). The binding sites (n) 0.98 ± 0.1. The thermodynamic parameters showed that the interaction between 7HC and thyroxine was driven mainly by hydrogen bonding interactions and van der Waals force. Calibration for thyroxine, based on quenching titration data, was linear in the concentration range 2.0 × 10⁻⁸ to 3.0 × 10⁻⁷ mol/l. The relative standard deviation was 2.58% for 2.0 × 10⁻⁷ mol/l thyroxine (n = 4) and the 3σ limit of detection was 3.42 × 10⁻⁸ mol/l in cationic surfactant CTAB medium.     

Biophysical Studies on the Interactions of a Classic Mitochondrial Uncoupler with Bovine Serum Albumin by Spectroscopic,Isothermal Titration Calorimetric and Molecular Modeling Methods

The interaction between a classic uncoupler (2,4-dinitrophenol, DNP) and bovine serum albumin (BSA) was investigated by fluorescence spectroscopy under the physiological conditions. The fluorescence quenching constants were calculated by the Stern-Volmer equation, and based upon the temperature dependence of quenching constants, it was proved that DNP caused a static quenching of the intrinsic fluorescence of BSA. Owing to the static quenching mechanism, different associative binding constants at various temperatures were determined and thus the thermodynamic parameters, namely enthalpy (ΔH = −21.12 kJ mol⁻¹) and entropy changes (ΔS = 23.51 J mol⁻¹ K⁻¹) could be calculated based on the binding constants. Moreover, the enthalpy and entropy changes are consistent with the “Enthalpy-Entropy Compensation” equation obtained from our previous work. The negative enthalpy and positive entropy indicated that the electrostatic interactions played a major role in DNP-BSA binding process. Site marker competitive displacement experiments were carried out by using fluorescence and isothermal titration calorimetry (ITC) methods. These results showed that DNP bound with high affinity to Sudlow’s site I (subdomain IIA) of BSA. The distance (r = 3.78 nm) between donor (BSA) and acceptor (DNP) was obtained according to the mechanism of fluorescence resonance energy transfer (FRET). Furthermore, the results of synchronous fluorescence and circular dichroism (CD) spectroscopic studies indicated that the microenvironment and the secondary conformation of BSA were altered. The above results were supported by theoretical molecular modeling methods.     

Fluorescence Enhancement Effect for the Determination of Polychlorinated Biphenyls with Bovine Serum Albumin

A sensitive and selective method for the trace determination of 3, 3’, 4, 4’-tetrachlorobiphenyl (PCB77) by using bovine serum albumin (BSA) as a fluorescence probe was introduced. Under optimum conditions, the enhanced fluorescence intensity was proportional to the concentration of polychlorinated biphenyls in the range of 8.9 × 10⁻⁸–5.0 × 10⁻⁶ mol L⁻¹ for PCB77, and 5.0 × 10⁻⁷–5.0 × 10⁻⁶ mol L⁻¹ for 2, 2’, 5, 5’-tetrachlorbiphenyl (PCB52). The detection limits (S/N = 3) of PCB77 and PCB52 were 2.6 × 10⁻⁸ mol L⁻¹ and 2.9 × 10⁻⁷ mol L⁻¹, respectively. Furthermore, the fluorescence enhancement mechanism was discussed in detail. Results indicated that fluorescence enhancement of the system originated from the formation of BSA-PCBs complexes. In addition, PCBs were mainly bound to the tyrosine residues in BSA molecules.     

Binding of Engeletin with Bovine Serum Albumin: Insights from Spectroscopic Investigations

In this paper, several spectroscopic techniques were used to investigate the interaction of engeletin (ELN) with bovine serum albumin (BSA). The analysis of UV–Vis absorption and fluorescence spectra revealed that ELN and BSA formed a static complex ELN–BSA, and ELN quenched the fluorescence of BSA effectively. According to the thermodynamic parameters ΔS ⁰ = 47.27 J·mol⁻¹·K⁻¹ and ΔΗ ⁰ = −10.34 kJ·mol⁻¹, the hydrophobic and hydrogen bond interactions were suggested to be the major interaction forces between ELN and BSA. Raman spectroscopy indicated that the binding of ELN slightly changed the conformations and microenviroment of BSA and decreased the α–helix content of BSA.     

Sensitive Determination of Norfloxacin by the Fluorescence Probe of Terbium (III)- Sodium Dodecylbenzene Sulfonate and Its Luminescence Mechanism

The fluorescence system of the norfloxacin-Tb³⁺- sodium dodecylbenzene sulfonate (SDBS) was investigated in this paper. The experiments indicated that the fluorescence intensity of the Tb³⁺-SDBS was greatly enhanced by the norfloxacin. On the basis of the above findings, a sensitive fluorimetric method for determining the norfloxacin was established. The fluorescence intensity was measured by a 1-cm quartz cell with the excitation wavelength of 290 nm and the emission wavelength of 545 nm. The enhanced fluorescence intensity of the system (Δ F) showed a good linear relationship with the concentration of norfloxacin in the range of 5.0×10⁻⁹ mol L⁻¹–2.0×10⁻⁶ mol L⁻¹, its correlation coefficient was 0.9991 and the detection limit (S/N=3) was 1.2×10⁻⁹ mol L⁻¹. The presented method was used to determine the norfloxacin in real pharmaceutical samples. The luminescence mechanism was also discussed in detail. In the fluorescence system of the norfloxacin-Tb³⁺-SDBS, the SDBS not only acted as the surfactant, but also acted as the energy donor.     

Simultaneous Determination of Acetylsalicylic Acid and Caffeine in Pharmaceutical Formulation by First Derivative Synchronous Fluorimetric Method

A sensitive, rapid, and specific assay has been developed for the simultaneous determination of acetylsalicylic acid and caffeine in commercial tablets based on their natural fluorescence. The mixture of these drugs was resolved by first derivative synchronous fluorimetric technique using two scans. At Δλ=106 nm, using first derivative synchronous scanning, only acetylsalicylic acid yields a detectable signal at 316 nm (peak to zero method) which is unaffected by caffeine. At Δλ=30 nm, the signal of caffeine at 288 nm (peak to zero method) is not affected by acetylsalicylic acid. The range of application is between 0.021 and 41.62 μg ml⁻¹ (correlation coefficient, R=0.9995) for acetylsalicylic acid and between 0.4486 and 44.86 μg ml⁻¹ (correlation coefficient, R=0.99786) for caffeine. The recovery range of 98.40–102% for acetylsalicylic acid and 90–100.5% for caffeine from their synthetic mixture was reported. Overall recovery of both compounds about 97–99% for acetylsalicylic acid and 97–98% for caffeine was obtained from real sample analysis. The detection limits are 0.0013 μg ml⁻¹ and 0.0306 μg ml⁻¹ for acetylsalicylic acid and caffeine, respectively. The relative standard deviation (n=10) for 20 μg ml⁻¹ of acetylsalicylic acid is 2.75% and for 2.2 μg ml⁻¹of caffeine is 1.7%.     

Fluorescence Quenching Reaction of Polyvinylpyrrolidone-Eosin Y System for the Determination of Polyvinylpyrrolidone

In pH 1.8 ∼ 2.8 weak acid medium, polyvinylpyrrolidone (PVP) and Eosin Y reacted to form complex that could result in Eosin Y (EY) fluorescence quenching. The maximum quenching wavelength was at 542 nm. The fluorescence quenching (ΔF) was proportional to the concentration of polyvinylpyrrolidone in a certain range. The linear range, the correlation coefficient and the detection limit were 0.33 ∼ 2.0 μg•mL⁻¹, 0.9994 and 99.6 ng•mL⁻¹, respectively. The influences of the coexistence substances were tested and the results showed that the method had good selectivity. Therefore, a new method based on fluorescence quenching of eosin Y by PVP for the determination of trace PVP was developed. The method was sensitive, simple and rapid, which was applied to the determination of trace PVP in the beer with satisfactory results. The reaction mechanism was also discussed.     

Binding of Quercetin to Lysozyme as Probed by Spectroscopic Analysis and Molecular Simulation

The binding of quercetin to lysozyme (LYSO) in aqueous solution was investigated by fluorescence spectroscopy, UV-vis absorption spectroscopy and molecular simulation at pH 7.4. The fluorescence quenching of LYSO by addition of quercetin is due to static quenching, the binding constants, K _a , were 3.63 × 10⁴, 3.31 × 10⁴ and 2.85 × 10⁴ L·mol⁻¹ at 288, 298 and 308 K, respectively. The thermodynamic parameters, enthalpy change, ∆H, and entropy change, ∆S, were noted to be −7.56 kJ·mol⁻¹ and 61.07 J·mol⁻¹·K⁻¹. The results indicated that hydrophobic interaction may play a major role in the binding process. The distance r between the donor (LYSO) and acceptor (quercetin) was determined as 3.34 nm by the fluorescence resonance energy transfer. The synchronous fluorescence spectroscopy showed the polarity around the tryptophan residues increased and the hydrophobicity decreased. Furthermore, the study of molecular simulation indicated that quercetin could bind to the active site (a pocket made up of 24 amino-acid residues) of LYSO mainly via hydrophobic interactions and that there were hydrogen interactions between the residues (Gln 57, Ile 98) of LYSO and quercetin. The accessible surface area (ASA) calculation verified the important roles of tryptophan (Trp) residues during the binding process.     

An ionic liquid-type multiwall carbon nanotubes paste electrode for electrochemical investigation and determination of morphine

The direct electrochemistry of morphine on modified multiwall carbon nanotubes using carbon ionic liquid (i.e., 1-butyl-3-methylimidazolium hexafluoro phosphate, ([C4mim]–[PF6])) was studied. It was found that the electrode showed sensitive voltammetric response to morphine. The experimental results suggested that the modified electrode promoted electron transfer reaction for the oxidation of morphine. The electron transfer coefficient and charge transfer resistant (R _ct) of morphine at the modified electrode were calculated. Under the optimized conditions at pH 8.0, the peak current was linear to morphine concentrations over the concentration range of 0.45–450 μmol L⁻¹, using differential pulse voltammetry. The detection limit was 0.14 μmol L⁻¹. The proposed method was successfully applied to the determination of morphine in both ampoules and urine samples.     

Fluorescence Interaction and Determination of Calf Thymus DNA with Two Ethidium Derivatives

In this paper, we reported the syntheses and investigation of the modes of binding to DNA of the two new ethidium derivatives containing benzoyl and phenylacetyl groups of both amines at 3-and 8- positions. The interactions between calf thymus DNA (ct-DNA) and the two derivatives, 3,8-dibenzoylamino-5-ethyl-6-phenylphenantridinium cloride (E2) and 3,8-diphenylacetylamino-5-ethyl-6-phenylphenantridinium chloride (E3), were investigated by fluorescence quenching spectra and UV-vis absorption spectra. The Stern-Volmer quenching constants, binding constants, binding sites and the corresponding thermodynamic parameters ΔH, ΔS and ΔG were calculated at different temperatures. The results indicated the formation of E2 and E3-DNA complexes and van der Waals interactions as the predominant intermolecular forces in stabilizing for each complex. In addition, increasing nucleophilicity of the functional groups at 3- and 8- positions exhibited the respectable increment the DNA binding affinities of derivatives. The results of absorption, ionic strength and iodide ion quenching suggested that the interaction mode of E2 and E3 with ct-DNA was intercalative binding. The limit of detection (LOD) of ct-DNA were 7.49 × 10⁻⁸ (n = 4) and 4.18 × 10⁻⁸ mol/l (n = 7) in presence of E2 and E3, respectively.     

A Fluorescein-based Fluorogenic Probe for Fluoride Ion Based on the Fluoride-induced Cleavage of <Emphasis Type="Italic">tert</Emphasis>-butyldimethylsilyl Ether

A highly sensitive and selective fluorogenic probe for fluoride ion, fluorescein di-tert-butyldimethylsilyl ether (FTBS), was designed and synthesized. FTBS was a colorless, non-fluorescent compound and was synthesized via the one-step reaction of fluorescein with tert-butyldimethylsilyl chloride. Upon incubation with fluoride ion in DMF-water solution (7 : 3, V/V), the Si-O bond of FTBS was cleaved, causing a large increase in fluorescence intensity and thereby allowing a selective detection of fluoride ion. The fluorescence increase is linearly with fluoride concentration in the range 0.1–2.0 μmol L⁻¹ with a detection limit of 0.041 μmol L⁻¹ (3σ). The excellent selective signaling behavior of the proposed probe was found to originate from the high affinity of silicon toward fluoride ion. The method has been successfully applied to the fluoride determination in multi-trace elements injection and toothpaste samples, and the results are agreed well with those obtained by the fluoride-ion selective electrode method.     

Determination of Trace Mercury by Solid Substrate-Room Temperature Phosphorimetry Quenching Method Based on Catalytic Effect of Hg2+on Formation of the Ion Association Complex [Sn(XO)6]4+·[(Fin)4]

A new method for the determination of trace mercury by solid substrate-room temperature phosphorimetry (SS-RTP) quenching method has been established. In glycine-HCl buffer solution, xylenol orange (XO) can react with Sn⁴⁺ to form the complex [Sn(XO)₆]⁴⁺. [Sn(XO)₆]⁴⁺ can interact with Fin⁻ (fluorescein anion) to form the ion associate [Sn(XO)₆]⁴⁺·[(Fin)₄]⁻, which can emit strong and stable room temperature phosphorescence (RTP) on polyamide membrane (PAM). Hg²⁺ can catalyze H₂O₂ oxidizing the ion association complex [Sn(XO)₆]⁴⁺·[(Fin)₄]⁻, which causes the RTP to quench. The ΔIp value is directly proportional to the concentration of Hg²⁺ in the range of 0.016–1.6 fg spot⁻¹ (corresponding concentration: 0.040–4.0 pg ml⁻¹, 0.40 μl spot⁻¹), and the regression equation of working cure is ΔIp=10.03+83.15 m Hg²⁺ (fg spot⁻¹), (r=0.9987, n=6) and the detection limit (LD) is 3.6 ag spot⁻¹(corresponding concentration: 9.0×10^–15 g ml⁻¹, the sample volume: 0.4 μl). This simple, rapid, accurate method is of high selectivity and good repeatability, and it has been successfully applied to the determination of trace mercury in real samples. The reaction mechanism for catalyzing H₂O₂ oxidizing the ion association complex ([Sn(XO)₆]⁴⁺·[(Fin)₄]⁻) SS-RTP quenching method to determine trace mercury is also discussed.     

Low thermal expansion behavior and transport properties of Ni and Ge co-doped manganese nitride materials at cryogenic temperatures

A series of Ni and Ge co-doped manganese nitride materials were fabricated by mechanical ball milling followed by solid-state sintering. Their thermal expansion properties and electrical and thermal conductivities were investigated in the temperature range of 77–300 K. The results show that Ni and Ge co-doped manganese nitride materials have negative thermal expansion (NTE), and the operation-temperature window of NTE shifts toward the lower temperature region and the variation of linear thermal expansion (ΔL/L _(300K)) in the operation-temperature window of NTE decreases with increasing Ni content. The combination of these two factors results in a low coefficient of thermal expansion (CTE) at cryogenic temperatures. The average CTE of Mn₃(Cu_0.2Ni_0.4Ge_0.4)N drops to ‘zero’ in the temperature range of 190–77 K. The values of electrical and thermal conductivities of the Ni and Ge co-doped manganese nitride materials are in the ranges of 2–3×10³ (ohm cm)⁻¹ and 1.6–3.4 W (m K)⁻¹, respectively.     

Spectrofluorimetric Assessment of Doxycycline Hydrochloride in Pharmaceutical Tablets and Serum Sample Based on the Enhancement of the Luminescence Intensity of the Optical Sensor Sm<Superscript>3+</Superscript> Ion

A simple and sensitive spectrofluorimetric method for determination of trace amount of doxycycline hydrochloride (DC) in pharmaceutical tablets and serum samples was developed. In ammonia buffer solution of pH 8.9 the doxycycline hydrochloride can remarkably enhance the luminescence intensity of the Sm³⁺ ion in Sm³⁺- DC complex at λ_ex = 400 nm. The produced luminescence intensity of Sm³⁺- DC complex in DMSO is in proportion to the concentration of DC and used as optical sensor for its determination. The dynamic range for the determination of DC is 1 × 10⁻⁸ – 5 × 10⁻⁶ mol L⁻¹ and in case of quantum yield calculations is 7 × 10⁻⁹ – 5 × 10⁻⁶ mol L⁻¹ with detection limit of 6.5 × 10⁻¹⁰ mol L⁻¹. The enhancement mechanism of the luminescence intensity in the Sm³⁺- DC system has been also discussed. A comparison with other spectrofluorimetric methods for tetracycline derivatives in which Eu³⁺ ion is used instead of Sm³⁺ ion is also studied.     

Preparation and Characterization of CdHgTe Nanoparticles and Their Application on the Determination of Proteins

CdHgTe nanoparticles (NPs) with the emission in the near-infrared regions were prepared in aqueous solution, and were characterized by transmission electron microscopy, X-ray diffraction spectrometry, spectrofluorometry and ultraviolet-visible spectrometry. Based on the fluorescence quenching of CdHgTe NPs in the presence of proteins, a novel method for the determination of proteins with CdHgTe NPs as a near-infrared fluorescence probe was developed. Maximum fluorescence quenching was observed with the excitation and emission wavelengths of 500 and 693 nm, respectively. Under the optimal conditions, the calibration graphs were linear in the range of 0.04 × 10⁻⁶–5.6 × 10⁻⁶ g ml⁻¹ for lysozyme (Lyz) and 0.06 × 10⁻⁶–6.1 × 10⁻⁶ g ml⁻¹ for bovine hemoglobin (BHb), respectively. The limits of detection were 13 ng ml⁻¹ for Lyz and 27 ng ml⁻¹ for BHb, respectively. Four synthetic samples were determined and the results were satisfied.     

Fabrication of a nanoparticle TiO<Subscript>2</Subscript> photoelectrochemical cell utilizing a solid polymeric electrolyte of PAN–PC–LiClO<Subscript>4</Subscript>

A nanoparticle TiO₂ solid-state photoelectrochemical cell utilizing as a solid electrolyte of poly(acrylonitrile)–propylene–carbonate–lithium perchlorate (PAN–PC–LiClO₄) has been fabricated. The performance of the device has been tested in the dark and under illumination of 100-mW cm⁻² light. A nanoparticle TiO₂ film was deposited onto indium tin oxide-covered glass substrate by controlled hydrolysis technique assisted with spin-coating technique. The average grain size for the TiO₂ film is 76 nm. LiClO₄ salt was used as a redox couple. The room temperature conductivity of the electrolyte is 4.2 × 10⁻⁴ S cm⁻¹. A graphite electrode was prepared onto a glass slide by electron beam evaporation technique. The device shows the rectification property in the dark and shows the photovoltaic effect under illumination. The best J _sc and V _oc of the device were 2.82 μA cm⁻² and V _oc of 0.58 V, respectively, obtained at the conductivity of 4.2 × 10⁻⁴ S cm⁻¹ and intensity of 100 mW cm⁻². The J _sc was improved by about three times by introducing nanoparticle TiO₂ and by using a solid electrolyte of PAN–PC–LiClO₄ replacing PVC–PC–LiClO₄ in the device. The current transport mechanism of the cell is also presented in this paper.