Copper nanoclusters as fluorescence‐quenching probes for the quantitative analysis of total iodine

Tannic acid‐coated copper nanoclusters (CuNCs@TA) were synthesized and used quantitatively to analyze iodine in kelp. Compared with other methods for iodine detection, the proposed method showed excellent performance. The iodine‐induced linear decrease in the fluorescence intensity of CuNCs@TA allowed the quantitative detection of iodine in the range 20–100 μM, and the limit of detection for iodine was 18 nM. The probe can be used for the determination of iodine in real samples with reliable and accurate results. Modified Stern–Volmer equation and thermodynamic calculation studies were used to discuss the quenching mechanism.     

Novel biosensor system model based on fluorescence quenching by a fluorescent streptavidin and carbazole‐labeled biotin

In the present study, a novel molecular biosensor system model was designed by using a couple of the fluorescent unnatural mutant streptavidin and the carbazole‐labeled biotin. BODIPY‐FL‐aminophenylalanine (BFLAF), a fluorescent unnatural amino acid was position‐specifically incorporated into Trp120 position of streptavidin by four‐base codon method. On the other hand, carbazole‐labeled biotin was synthesized as a quencher for the fluorescent Trp120BFLAF mutant streptavidin. The fluorescence of fluorescent Trp120BFLAF mutant streptavidin was decreased as we expected when carbazole‐labeled biotin was added into the mutant streptavidin solution. Furthermore, the fluorescence decrease of Trp120BFLAF mutant streptavidin with carbazole‐labeled biotin (100 nM) was recovered by the competitive addition of natural biotin. This result demonstrated that by measuring the fluorescence quenching and recovery, a couple of the fluorescent Trp120BFLAF mutant streptavidin and the carbazole‐labeled biotin were successfully applicable for quantification of free biotin as a molecular biosensor system. Copyright © 2016 John Wiley & Sons, Ltd.     

Determination of selenium via the fluorescence quenching effect of selenium on hemoglobin‐catalyzed peroxidative reaction

A new method for the determination of selenium based on its fluorescence quenching on the hemoglobin‐catalyzed reaction of H₂O₂ and l ‐tyrosine has been established. The effect of pH, foreign ions and the optimization of variables on the determination of selenium was examined. The calibration curve was found to be linear between the fluorescence quenching (F₀/F) and the concentration of selenium within the range of 0.16‐4.00 µg/mL. The detection limit was 1.96 ng/mL and the relative standard deviation was 3.14%. This method can be used for the determination of selenium in Se‐enriched garlic bulbs with satisfactory results. Copyright © 2014 John Wiley & Sons, Ltd.     

The transverse organisation of ubiquinones in mitochondrial membranes as determined by fluorescence quenching

The transverse organisation of ubiquinone in mitochondrial membranes was investigated by quenching a set of fluorescent fatty acids. We show that the fluorescent moiety of the probes is located at a graded series of depths in the mitochondrial membrane. The probes sense the characteristics of the lipid phase and do not significantly perturb mitochondrial function as measured by the respiratory control ratio and the ADP/O ratio. The anthroyloxy fatty acids are readily quenched by ubiquinone-10. A recently developed method in the analysis of quenching data was used to obtain the subvolume of the membrane within which the quenching interactions are confined. The results indicate that ubiquinone-10 is restricted to two sites in the transverse plane of the membrane: one near the surface and the other close to the bilayer centre. The implications of these findings for the two-pool model of ubiquinone organisation are discussed.Abbreviations n-AS n-(9-anthroyloxy) stearic acids (n=6,9,12) - n-AP n-(9-anthroyloxy) palmitic acids (n=2,16) - n-AF n-(9-anthroyloxy) fatty acids (n=2,6,9,12,16) - n nitroxide stearic acids (n=5,16) - UQ _n ubiquinone-n (n=4,6,10) - HBHM heavy beet heart mitochondria     

Effect of fluorescence quenching model on quinoline derivative with cobalt chloride metal ion using steady state and time resolved spectroscopic methods

Quinoline derivative, i.e. quinilone yellow with the scientific name [sodium 2-(2,3-dihydro-1,3-dioxo-1H-inden-2-yl)quinoline-6,8-disulphonate] (SQDS) is analysed for fluorescence resonance energy transfer (FRET). Fluorescence quenching mechanism is studied by employing steady state and transient state spectroscopic measurements. Cobalt chloride is used as quencher in the present study. Linearity was observed in Stern–Volmer plots for transient state as well as steady state. This was further attributed to a mechanism of collisional quenching. Efficiency in fluorescence quenching is observed as there is a correlation between quenching constants of both transient and steady state. A significant energy transfer is reported between metal ions and SQDS molecule, according to FRET theory. Characterization results are studied and analysed. Application in the field of non-linear optics are predicted for SQDS. With Kurtz and Perry powder technique, SHG (second harmonic generation) efficiency was measured using Q-switched mode locked Nd:YAG laser emitting 1064 nm the first time with this compound.     

A novel fluorescent assay for oxytetracycline hydrochloride based on fluorescence quenching of water‐soluble CdTe nanocrystals

A novel assay for oxytetracycline hydrochloride (OTC) based on fluorescence quenching was developed from the interaction between functionalized cadmium telluride quantum dots (CdTe QDs) and OTC. Optimum conditions for the detection of OTC were found after investigating all factors. Under optimum conditions, luminescence of CdTe nanocrystals (λ_ex = 365 nm, λ_em = 562 nm) was quenched by OTC in a concentration‐dependent manner best described by a modified Stern‐Volmer type equation. Good linearity was obtained with a regression coefficient of 0.9999 in the range of 1.34 ~ 13.4 x 10^‐5 mol/L and a limit of detection of 3.08 x 10^‐7 mol/L. In addition, the quenching mechanism was also established. The results imply that the close proximity of OTC‐CdTe was driven by electrostatic attraction and the resulting effective electron transfer from OTC to QDs could be responsible for fluorescence quenching of CdTe‐QDs. Copyright © 2012 John Wiley & Sons, Ltd.     

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.     

Solvatochromic study of 3‐N‐(N′‐methylacetamidino)benzanthrone and its interaction with dopamine by the fluorescence quenching mechanism

The change in photophysical properties of the organic molecule due to solvatochromic effect caused by different solvent environments at room temperature gives information about the dipole moments of 3‐N‐(N′‐methylacetamidino)benzanthrone (3‐MAB). The quantum yield, fluorescence lifetime of 3‐MAB was measured in different solvents to calculate radiative and non‐radiative rate constants. The results revealed that the excited state dipole moment (μ_e) is relatively larger compared to the ground state dipole moment (μ_g), indicating the excited state of the dye under study is more polar than the ground state and the same trend is noticed with theoretical calculations performed using the CAM‐B3LYP/6‐311+G(d,p) method. Further, the study on preferential solvation was carried out for 3‐MAB dye in ethyl acetate–methanol solvent mixture. The fluorescence quenching method has been employed for the detection of dopamine using 3‐MAB as fluorescent probe, using steady‐state and time resolved methods at room temperature. The method enables dopamine in the micro molar range to be detected. Also, an attempt to verify the quenching process by employing different models has been tried. Various rate parameters are measured using these models, our results indicates the quenching process is diffusion limited.     

Bimolecular fluorescence quenching reactions of the biologically active coumarin composite 2‐acetyl‐3H‐benzo[f]chromen‐3‐one in different solvents

A study on fluorescence quenching was carried out for the coumarin derivative 2‐acetyl‐3H‐benzo[f]chromen‐3‐one (2AHBC) with aniline at room temperature. Efficient fluorescence quenching was observed and Stern–Volmer (S–V) plots showed upward curves from linearity in all solvents of different polarities. For the solute 2AHBC, ground state complex formation does not hold in our study. The kinetic distance (r) value was found to be greater than the encounter distance (R) and indicated that the quenching reaction was held within the sphere of action. Diffusion‐limited reactions were found to be more prominent in high polarity solvents, namely dimethyl sulfoxide (DMSO), DMF, ACN, methanol, ethanol, propanol and DCM. The relationships between quenching constant (K_SV) and dielectric constants (ε) of the different solvents were studied.     

A sensitive fluorescence quenching method for the detection of tartrazine with acriflavine in soft drinks

In this work, a simple, rapid, sensitive, selective spectrofluorimetric method was applied to detect tartrazine. The fluorescence of acriflavine could be efficiently quenched by tartrazine. The method manifested real time response as well as presented satisfied linear relationship to tartrazine. The linear response range of tartrazine (R² = 0.9995) was from 0.056 to 5 μmol L^?1. The detection limit (3σ/k) was 0.017 μmol L^?1, indicating that this method could be applied to detect traces of tartrazine. The accuracy and precision of the method was further assured by recovery studies via a standard addition method, with percentage recoveries in the range of 96.0% to 103.0%. Moreover, a quenching mechanism was investigated systematically by the linear plots at varying temperatures based on the Stern–Volmer equation, fluorescence lifetime and UV–visible absorption spectra, all of which proved to be static quenching. This sensitive, selective assay possessed a great application prospect for the food industry owing to its simplicity and rapidity for the detection of tartrazine.     

Mercaptopropionic acid–capped CdTe quantum dots as fluorescence probe for the determination of salicylic acid in pharmaceutical products

Mercaptopropionic acid (MPA)–capped cadmium telluride (CdTe) quantum dot (QDs) fluorescent probes were synthesized in aqueous solution and used for the determination of salicylic acid. The interaction between the MPA–capped CdTe QDs and salicylic acid was studied using fluorescence spectroscopy and some parameters that could modify the fluorescence were investigated to optimize the measurements. Under optimum conditions, the quenched fluorescence intensity of MPA–capped CdTe QDs was linearly proportional to the concentration of salicylic acid in the range of 0.5–40 µg mL^–1 with a coefficient of determination of 0.998, and the limit of detection was 0.15 µg mL^–1. The method was successfully applied to the determination of salicylic acid in pharmaceutical products, and satisfactory results were obtained that were in agreement with both the high pressure liquid chromatography (HPLC) method and the claimed values. The recovery of the method was in the range 99 ± 3% to 105 ± 9%. The proposed method is simple, rapid, cost effective, highly sensitivity and eminently suitable for the quality control of pharmaceutical preparation. The possible mechanisms for the observed quenching reaction was also discussed. Copyright © 2015 John Wiley & Sons, Ltd.     

Spectrofluorimetric study on fluorescence quenching of tyrosine and l‐tryptophan by the aniracetam cognition enhancer drug: quenching mechanism using Stern–Volmer and double‐log plots

A novel approach on fluorescence quenching of tyrosine and l ‐tryptophan is presented for spectrofluorimetric determination of aniracetam in drug substances and products. The quenching mechanism was investigated using Stern–Volmer plots and ultraviolet spectra figures of quencher–fluorophore mixtures. Binding constant and stoichiometry were calculated using double‐log plots. The spectrofluorimetric method was optimized for the experimental conditions affecting fluorescence quenching including fluorophore concentration, diluent, and reaction time. Moreover, the pH‐rate profile of aniracetam was studied using simple kinetics and found to be stable within the pH range 5–8. Fluorescence quenching of tyrosine and l ‐tryptophan were observed on addition of aniracetam in aqueous medium at pH 5.5–6.5. Aniracetam quenched the fluorescence of tyrosine and l ‐tryptophan in the concentration range 1–20 μg/ml and 0.3–20 μg/ml, respectively, with binomial relationships between quenching values (ΔF) and aniracetam concentration. Limits of detection were found to be 0.10 μg/ml for tyrosine–aniracetam and 0.14 μg/ml for l ‐tryptophan–aniracetam. Method validation was performed as per ICH guidelines and demonstrated that the developed spectrofluorimetric method was accurate, precise, specific, and suitable for analysis of aniracetam in routine quality control laboratories. All experimental materials and solvents used are eco‐friendly, indicating that the cited spectrofluorimetric procedure is an excellent green method.     

Investigation on the interaction of cefpirome sulfate with lysozyme by fluorescence quenching spectroscopy and synchronous fluorescence spectroscopy

The reaction mechanism of cefpirome sulfate with lysozyme at different temperatures (298, 310 and 318 K) was investigated using fluorescence quenching and synchronous fluorescence spectroscopy under simulated physiological conditions. The results clearly demonstrated that cefpirome sulfate caused strong quenching of the fluorescence of lysozyme by a static quenching mechanism. The binding constants obtained using the above methods were of the same order of magnitude and very similar. Static electric forces played a key role in the interaction between cefpirome sulfate and lysozyme, and the number of binding sites in the interaction was close to 1. The values of Hill's coefficients were > 1, indicating that drugs or proteins showed a very weakly positive cooperativity in the system. In addition, the conclusions obtained from the two methods using the same equation were consistent. The results indicated that synchronous fluorescence spectrometry could be used to study the binding mechanism between drug and protein, and was a useful supplement to the fluorescence quenching method. In addition, the effect of cefpirome sulfate on the secondary structure of lysozyme was analyzed using circular dichroism spectroscopy. Copyright © 2015 John Wiley & Sons, Ltd.     

Nitrogen and sulfur co‐doped carbon nanodots toward bovine hemoglobin: A fluorescence quenching mechanism investigation

A deep understanding of the molecular interactions of carbon nanodots with biomacromolecules is essential for wider applications of carbon nanodots both in vitro and in vivo. Herein, nitrogen and sulfur co‐doped carbon dots (N,S‐CDs) with a quantum yield of 16% were synthesized by a 1‐step hydrothermal method. The N,S‐CDs exhibited a good dispersion, with a graphite‐like structure, along with the fluorescence lifetime of approximately 7.50 ns. Findings showed that the fluorescence of the N,S‐CDs was effectively quenched by bovine hemoglobin as a result of the static fluorescence quenching. The mentioned quenching mechanism was investigated by the Stern‐Volmer equation, temperature‐dependent quenching, and fluorescence lifetime measurements. The binding constants, number of binding sites, and the binding average distance between the energy donor N,S‐CDs and acceptor bovine hemoglobin were calculated as well. These findings will provide for valuable insights on the future bioapplications of N,S‐CDs.     

Entropy‐driven binding of octyl gallate in albumin: Failure in the application of temperature effect to distinguish dynamic and static fluorescence quenching

Fluorescence quenching is widely used to obtain association constants between proteins and ligands. This methodology is based on assumption that ground‐state complex between protein and ligand is responsible for quenching. Here, we call the attention about the risk of using the temperature criterion for decision of applying or not fluorescence quenching data to measure association constants. We demonstrated that hydrophobic effect can be the major force involved in the interaction and, as such, superposes the well‐established rationalization that host/guest complexation is weakened at higher temperatures due to loss of translational and rotational degrees of freedom. To do so, the complexation of bovine serum albumin with octyl gallate was studied by steady‐state, time‐resolved fluorescence spectroscopy and isothermal titration calorimetry. The results clearly demonstrated the complexation, even though the Stern‐Volmer constant increased at higher temperatures (1.6 × 10⁴ and 4.1 × 10⁵ mol^?1 L at 20°C and 40°C), which could suggest a simple dynamic process and not complexation. The entropy‐driven feature of the interaction was demonstrated by the unfavorable enthalpy (?H° = 104.4 kJmol^?1) but favorable entropy (?S° = 447.5 Jmol^?1 K^?1). The relevance of the ligand hydrophobicity was also evaluated by comparing ascorbic acid and its ester ascorbyl palmitate. Docking simulations showed a higher number of hydrophobic contacts and lower energy poses for the esters, confirming the experimental results. In conclusion, the well‐established rationalization that host/guest complexation is weakened at higher temperatures is not straightforward for protein‐ligand interactions. Hence, the temperature effect for a decision between static and dynamic quenching and its use to decide if a complexation at ground state is taking place between ligand and protein should not be used.     

Optical ascorbic acid sensor based on the fluorescence quenching of silver nanoparticles

A sensitive and selective fluorimetric sensor for the assay of ascorbic acid (AA) using silver nanoparticles as emission reagent was investigated. In this study, silver nanoparticles were prepared based on aqueous–gaseous phase reaction of silver nitrate solution and ammonia gas. The nanoparticles were water‐soluble, stable and had a narrow emission band. They were used as a fluorescence probe for the assay of ascorbic acid on its quenching effect on the emission of silver nanoparticles. The principal reason for quenching is likely to be a complexation between ascorbic acid and silver nanoparticles. The quenching mechanism was established by Stern–Volmer law. Under the optimum conditions, the quenched fluorescence intensity was linear with the concentration of ascorbic acid in the range of 4.1 × 10^?6 to 1.0 ×10^?4 m (r = 0.9985) with a detection limit of 1.0 × 10^?7 m . The RSD for repeatability of the sensor for the assay of ascorbic acid concentration of 3.0 × 10^?5 and 4.0 × 10^?6 m was found to be 1.5 and 1.3%, respectively. The proposed method was applied to the determination of ascorbic acid in vegetables and vitamin C tablets. Copyright © 2009 John Wiley & Sons, Ltd.     

NPQ(T): a chlorophyll fluorescence parameter for rapid estimation and imaging of non‐photochemical quenching of excitons in photosystem‐II‐associated antenna complexes

In photosynthesis, light energy is absorbed by light‐harvesting complexes and used to drive photochemistry. However, a fraction of absorbed light is lost to non‐photochemical quenching (NPQ) that reflects several important photosynthetic processes to dissipate excess energy. Currently, estimates of NPQ and its individual components (q_E, q_I, q_Z and q_T) are measured from pulse‐amplitude‐modulation (PAM) measurements of chlorophyll fluorescence yield and require measurements of the maximal yield of fluorescence in fully dark‐adapted material (F_m), when NPQ is assumed to be negligible. Unfortunately, this approach requires extensive dark acclimation, often precluding widespread or high‐throughput use, particularly under field conditions or in imaging applications, while introducing artefacts when F_m is measured in the presence of residual photodamaged centres. To address these limitations, we derived and characterized a new set of parameters, NPQ_(T), and its components that can be (1) measured in a few seconds, allowing for high‐throughput and field applications; (2) does not require full relaxation of quenching processes and thus can be applied to photoinhibited materials; (3) can distinguish between NPQ and chloroplast movements; and (4) can be used to image NPQ in plants with large leaf movements. We discuss the applications benefits and caveats of both approaches.     

Real‐time nucleic acid sequence–based amplification (NASBA) using an adenine‐induced quenching probe and an intercalator dye

Aims: We found that an adenine base caused fluorescence quenching of a fluorescein (FL)‐labelled probe in DNA:RNA hybrid sequences, and applied this finding to a nucleic acid sequence–based amplification (NASBA) method. Methods and Results: The present NASBA method employed a probe containing an FL‐modified thymine at its 3′ end and ethidium bromide (EtBr) on the basis of a combination of adenine‐induced quenching and fluorescence resonance energy transfer (FRET) between the FL donor and EtBr acceptor. This NASBA was used to detect Shiga toxin (STX) stx‐specific mRNA in STX‐producing Escherichia coli, demonstrating rapid quantification of the target gene with high sensitivity. Conclusion: Although the inherent quenching effect of adenine was inferior to that of guanine, FRET between the FL and EtBr moieties enhanced the adenine‐induced quenching, allowing rapid and sensitive real‐time NASBA detection. Significance and Impact of the Study: This study gives a novel real‐time diagnostic system based on NASBA for a sensitive mRNA (or viral RNA) detection.     

The relationship between non-photochemical quenching of chlorophyll fluorescence and the rate of photosystem 2 photochemistry in leaves

It has been suggested previously that non-photochemical quenching of chlorophyll fluorescence is associated with a decrease in the rate of photosystem 2 (PS 2) photochemistry. In this study analyses of fluorescence yield changes, induced by flashes in leaves exhibiting different amounts of non-photochemical quenching of fluorescence, are made to determine the effect of non-photochemical excitation energy quenching processes on the rate of PS 2 photochemistry. It is demonstrated that both the high-energy state and the more slowly relaxing components of non-photochemical quenching reduce the rate of PS 2 photochemistry. Flash dosage response curves for fluorescence yield show that non-photochemical quenching processes effectively decrease the relative effective absorption cross-section for PS 2 photochemistry. It is suggested that non-photochemical quenching processes exert an effect on the rate of PS 2 photochemistry by increasing the dissipation of excitation energy by non-radiative processes in the pigment matrices of PS 2, which consequently results in a decrease in the efficiency of delivery of excitation energy for PS 2 photochemistry.