Fluorescence Experiments Research Articles

Fluorescence Study of Aggregation Behaviour of Cationic Surfactant Carbethopendecinium Bromide and its Comparison with Cetyltrimethylammonium Bromide

Abstract The aggregation behaviour of the cationic surfactant carbethopendecinium bromide (Septonex) and cetyltrimethylammonium bromide (CTAB) in aqueous solution was investigated by steady-state and time-resolved fluorescence experiments. Values of critical micelle concentration were obtained from the pyrene emission polarity index. The micellar aggregation numbers for four different concentrations of both surfactants were determined by steady-state and time-resolved fluorescence quenching methods, using pyrene as a probe and cetylpyridinium chloride as a quencher. The results obtained for Septonex were compared with the aggregation parameters of CTAB. Generally, Septonex micelles were formed by a smaller number of surfactant monomers than in the case of CTAB and the critical micelle concentration of Septonex is lower than that of CTAB. Also, the concentration dependence of the micelle aggregation number was found.

5-Methoxyquinoline Photobasicity Is Mediated by Water Oxidation.

A mechanism explaining the photobasicity of 5-methoxyquinoline (5-MeOQ) is proposed on the basis of nonadiabatic molecular dynamics simulations using time-dependent hybrid density functional theory (TDDFT) and fewest switches surface hopping (SH) and analysis of existing ultrafast spectroscopy experiments. According to the TDDFT-SH simulations, the rate-determining step is hole transfer from photoexcited 5-MeOQ to adjacent water molecules within ∼5 ps followed by rapid electron-coupled proton transfer and deactivation to the ground state. This fast redox-catalyzed proton transfer mechanism is consistent with simple thermodynamic arguments, correlated wavefunction calculations, and recent isotope substitution time-resolved fluorescence experiments. Although energetically feasible, direct protonation of 5-MeOQ in the S1 state appears to be too slow to contribute significantly to 5-MeOQ photobasicity in aqueous solution.

Heterobimetallic nickel(II) and palladium(II) complexes derived from S-benzyl-N- (ferrocenyl)methylenedithiocarbazate: Trypanocidal activity and interaction with Trypanosoma cruzi Old Yellow Enzyme (TcOYE)

Reactions of Ni(II) and Pd(II) precursors with S-benzyl-N-(ferrocenyl)methylenedithiocarbazate (HFedtc) led to the formation of heterobimetallic complexes of the type [MII(Fedtc)2] (M = Ni and Pd). The characterization of the compounds involved the determination of melting point, FTIR, UV–Vis, 1H NMR, elemental analysis and electrochemical experiments. Furthermore, the crystalline structures of HFedtc and [NiII(Fedtc)2] were determined by single crystal X-ray diffraction. The compounds were evaluated against the intracellular form of Trypanosoma cruzi (Tulahuen Lac-Z strain) and the cytotoxicity assays were assessed using LLC-MK2 cells. The results showed that the coordination of HFedtc to Ni(II) or Pd(II) decreases the in vitro trypanocidal activity while the cytotoxicity against LLC-MK2 cells does not change significantly. [PdII(Fedtc)2] showed the greater potential between the two complexes studied, showing an SI value of 8.9. However, this value is not better than that of the free ligand with an SI of 40, a similar value to that of the standard drug benznidazole (SI = 48). Additionally, molecular docking simulations were performed with Trypanosoma cruzi Old Yellow Enzyme (TcOYE), which predicted that HFedtc binds to the protein, almost parallel to the flavin mononucleotide (FMN) prosthetic group, while the [NiII(Fedtc)2] complex was docked into the enzyme binding site in a significantly different manner. In order to confirm the hypothetical interaction, in vitro experiments of fluorescence quenching and enzymatic activity were performed which indicated that, although HFedtc was not processed by the enzyme, it was able to act as a competitive inhibitor, blocking the hydride transfer from the FMN prosthetic group of the enzyme to the menadione substrate.

Effect of the Substitution Position on the Electronic and Solvatochromic Properties of Isocyanoaminonaphthalene (ICAN) Fluorophores.

The properties of 1,4-isocyanoaminonaphthalene (1,4-ICAN) and 2,6-isocyanoaminonaphthalene (2,6-ICAN) isomers are discussed in comparison with those of 1,5-isocyanoaminonaphthalene (1,5-ICAN), which exhibits a large positive solvatochromic shift similar to that of Prodan. In these isocyanoaminonaphthalene derivatives, the isocyano and the amine group serve as the donor and acceptor moieties, respectively. It was found that the positions of the donor and the acceptor groups in these naphthalene derivatives greatly influence the Stokes and solvatochromic shifts, which decrease in the following order: 1,5-ICAN > 2,6-ICAN > 1,4-ICAN. According to high-level quantum chemical calculations, this order is well correlated with the charge transfer character of these compounds upon excitation. Furthermore, unlike 1,5-ICAN, the 1,4-ICAN and 2,6-ICAN isomers showed relatively high quantum yields in water, that were determined to be 0.62 and 0.21, respectively. In addition, time-resolved fluorescence experiments revealed that both the radiative and non-radiative decay rates for these three ICAN isomers varied unusually with the solvent polarity parameter ET(30). The explanations of the influence of the solvent polarity on the resulting steady-state and time-resolved fluorescence emission spectra are also discussed.

Role of GRP78 inhibiting artesunate-induced ferroptosis in KRAS mutant pancreatic cancer cells.

Objective: To investigate the exact role of GRP78 in artesunate-induced ferroptosis in KRAS mutant pancreatic cancer cells.Methods: Artesunate-induced KRAS mutant human pancreatic cancer cells (AsPC-1 and PaTU8988) ferroptosis was confirmed by fluorescent staining experiments and CCK8. Western blot and short-hairpin RNA-based knockdown assays were conducted to detect GRP78 activity and its role in artesunate-induced ferroptosis.Results: Artesunate induced AsPC-1 and PaTU8988 cell death in ferroptosis manner, rather than necrosis or apoptosis. In addition, artesunate increased the mRNA and protein levels of GRP78 in a concentration-dependent manner in AsPC-1 and PaTU8988 cells. Knockdown GRP78 enhanced artesunate-induced ferroptosis of pancreatic cancer cells in vitro and in vivo.Conclusion: Combining artesunate with GRP78 inhibition may be a novel maneuver for effective killing of KRAS mutant pancreatic ductal adenocarcinoma cells.

Synthesis and Preclinical Evaluation of the Fibrin-Binding Cyclic Peptide 18F-iCREKA: Comparison with Its Contrasted Linear Peptide.

Purpose Cys-Arg-Glu-Lys-Ala (CREKA) is a pentapeptide which can target fibrin-fibronectin complexes. Our previous study has built a probe called iCREKA which was based on CREKA and has proved the feasibility and specificity of iCREKA by the fluorescence experiment. The purpose of this study is to achieve the 18F-labeled iCREKA and make preclinical evaluation of the 18F-iCREKA with comparison of its contrasted linear peptide (LP). Methods CREKA, LP, and iCREKA were labeled by the Al18F labeling method, respectively. These 18F-labeled peptides were evaluated by the radiochemistry, binding affinity, in vitro stability, in vivo stability, micro-PET imaging, and biodistribution tests. Results 18F-NOTA-iCREKA was stable both in vitro and in vivo. However, 18F-NOTA-CREKA and 18F-NOTA-LP were both unstable. The FITC or 18F-labeled iCREKA could be abundantly discovered only in matrix metalloproteinases- (MMPs-) 2/9 highly expressed U87MG cells, while the FITC or 18F-labeled LP could also be abundantly discovered in MMP-2/9 lowly expressed Caov3 cells. Biodistribution and micropositron emission tomography (PET) imaging revealed that the U87MG xenografts showed a higher uptake of 18F-NOTA-iCREKA than 18F-NOTA-LP while the Caov3 xenografts showed very low uptake of both 18F-NOTA-iCREKA and 18F-NOTA-LP. The tumor-to-muscle (T/M) ratio of 18F-NOTA-iCREKA (9.93 ± 0.42) was obviously higher than 18F-NOTA-LP (2.69 ± 0.35) in U87MG xenografts. Conclusions The novel CREKA-based probe 18F-NOTA-iCREKA could get a high uptake in U87MG cells and high T/M ratio in U87MG mice. It was more stable and specific than the 18F-NOTA-LP.

Bisnaphthalimide-based fluorescent sensor for detecting alcohol and application in evaluating the liquors

A fluorescent sensor (bisNAP) was designed to output the light-up signals in alcohols, which was used to detect alcoholic strength of distillate spirit. bisNAP was constructed according to the Först resonance energy transfer (FRET) process with two naphthalimide chromophores. A 4-phenylnapthalimide (NAPP), a bright emitter in wide range of polarity, was chose as the energy donor and a 4-imine naphthalimide (NAPI) was chose as the energy acceptor. The emission band of NAPP clearly overlapped with the absorption band of NAPI that met the maximum of energy transfer efficiency. The polarity-related experiments showed that the emission of bisNAP was enhanced in the high polar solvents, such as alcohols and DMSO. The fluorescent decay experiments were used to explore the nature of above inverse phenomenon. The SEM images were obtained from different solvents and the powder, which could further understand the interaction of fluorescent molecules surrounding by the solvent molecular cages with different polarity. Consideration of its high brightness in alcohols, the bisNAP was used in rapid detecting the alcoholic strength of the spirit products. Furthermore, this work provided an efficient strategy for designing the “turn on” sensors working in the high polar mediums in several fields.

Investigation of inhibitory potential of quercetin to the pyruvate dehydrogenase kinase 3: Towards implications in anticancer therapy

Pyruvate dehydrogenase kinase 3 (PDK3) is a mitochondrial protein, has recently been considered as a potential pharmacological target for varying types of cancer. Here, we report the binding mechanism of quercetin to the PDK3 by using molecular docking, simulation, fluorescence spectroscopy and isothermal titration calorimetric assays. Molecular docking along with simulation provided an in-depth analysis of protein-ligand interactions. We have observed that quercetin interacts to the important residues of active site cavity of PDK3 and shows a well-ordered conformational fitting. The stability of quercetin-PDK3 complex is maintained by several non-covalent interactions throughout the simulation. To complement in silico findings with the experiments, we have successfully expressed and purified human PDK3. Both fluorescence and isothermal titration calorimetric experiments showed excellent binding affinity of quercetin to the PDK3. Kinase inhibition assay further revealed a significant inhibitory potential of quercetin to the PDK3 with the IC50 values in μM range. Quercetin is non-toxic to HEK293, and significantly inhibits the proliferation of cancer (HepG2 and A549) cell lines. All these observations clearly indicate that quercetin may be further evaluated as promising therapeutic molecule for PDK3 with required modifications and in vivo validation.

Entering the spotlight: Chitosan oligosaccharides as novel activators of CaCCs/TMEM16A

Calcium-activated chloride channels (CaCCs)/TMEM16A control diverse fundamental physiological functions, and abnormal function of TMEM16A will lead to various diseases including asthma, hypertension, gastrointestinal hypomotility and cancers. Therefore, TMEM16A as drug targets for related diseases has been increasingly concerned by researchers. In this work, COS were reported as novel natural activators of TMEM16A. It was demonstrated that COS can activate TMEM16A in a concentration dependent manner, with an EC50 of 74.5 μg/mL. Then, fluorescence experiments and inside-out patch clamp experiments were combined to confirm that COS can directly activate TMEM16A. Further, we compared the activation effects of COS monomers DP2 to DP6, with DP3 the best activator. Molecular simulation was performed to find that the binding sites between DP3 and TMEM16A are E143 and E146 in TMEM16A, and it was speculated that COS and TMEM16A may be combined by electrostatic interaction. Finally, we verified that guinea pig ileum contraction was promoted by COS and the monomers through activating TMEM16A. Collectively, COS are novel efficient natural activators of TMEM16A, with potential to be developed to treatment diseases caused by down-regulation of TMEM16A including gastrointestinal hypomotility.

Comparative studies of interaction of β-lactoglobulin with three polyphenols

To investigate the interaction mechanism between bovine protein β-lactoglobulin (β-LG) and theaflavin (TA), chlorogenic acid (CA) and delphinidin-3-O-glucoside (D3G), multi-spectrometry analytical methods and molecular modeling were applied. Fluorescence experiments proved that polyphenols strongly quenched the intrinsic fluorescence of β-LG mainly through static quenching and the main interaction force was hydrophobic interaction. Moreover, Fourier transform infrared (FTIR) and circular dichroism (CD) indicated that polyphenols changed β-LG secondary and tertiary structure. Enzyme-linked immunosorbent assay and molecular modeling study manifested that complex of β-LG with polyphenols could significantly reduce the IgE-binding capacity of β-LG due to the polyphenol binding site directly obscures the IgE linear epitopes. In conclusion, polyphenols had impact on the structure and potential functionality of β-LG, which would be valuable in dairy processing industry and food nutrition security.

Antibacterial activity of prenylated benzopyrans from Peperomia obtusifolia (Piperaceae)

Peperomia obtusifolia is a herbaceous perennial plant native to the Americas reported as a traditional medicine to treat snake bites and as a skin cleanser. The bioassay-guided fractionation of crude extracts from aerial parts of P. obtusifolia against a panel of clinically important fungi and bacteria, showed that hexane and dichloromethane extracts demonstrated selective bacterial inhibition, allowing the isolation of the known compounds peperobtusin A (1), and 3,4-dihydro-5-hydroxy-2,7-dimethyl-8-(3”-methyl-2”-butenyl)-2-(4’-methyl-1’,3’-pentadienyl)-2H-1-benzopyran-6-carboxylic acid (2) from dichloromethane extract. Compound 2 was active against Gram-positive bacteria including community acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) isolates and an Enterococcus faecalis vancomycin-resistant strain, with minimal inhibitory concentration (MIC) values of 4 μg/mL (10.8 μM) and 8 μg/mL (21.6 μM) respectively. The interaction of compound 2 with the bacterial membrane was demonstrated by means of Zeta potential experiments on S. aureus, then confirming the membrane damage by fluorescent microscopy experiments.

Excited State Decay Dynamics in 3‐Formyl‐4‐hydroxy Benzoic Acid: Understanding the Global Picture of an ESIPT‐Driven Multiple‐Emissive Species

AbstractRelaxation dynamics of excited state intramolecular proton transfer (ESIPT) probe 3‐formyl‐4‐hydroxy benzoic acid (FHBA) was studied by steady state, nanosecond time‐resolved fluorescence (TRF) and femtosecond transient absorption (TA) experiments in combination with adequate level of quantum chemical calculations. Global analysis of the transient data with target model reveals that enol conformers of both the neutral FHBA (I) and proton dissociated mono‐anionic species (III) undergo ESIPT in picosecond time scale (τESIPT ≈1.2 ps) towards the formation of corresponding vibrationally hot keto structure. Rapid intramolecular vibrational cooling (τ≈20 ps) follows singlet deactivation of the corresponding conformers with fluorescence lifetime (τfluo) of ∼5.1 and 2.0 ns, respectively as confirmed from stimulated emission (SE) and/or time resolved fluorescence experiments. Selective excitation of the di‐anionic structure (II) at 400 nm follows relatively simple excited state non‐ESIPT decay dynamics having fluorescence emission at 500 nm with τfluo ≈ 1.2 ns. Complete optimization of individual structures and construction of ESIPT profile was done by HF(CIS) as well as (TD)DFT calculation using 6–311++G(d,p) basis set and found to be consistent with the experimental observations.

Observations of the low-lying Ω = 1/2 states of nickel deuteride, NiD

While the spectrum of NiH has been extensively studied, only the Ω = 3/2 and 5/2 components of the strongly interacting 2Δ, 2Π and 2Σ+ molecular states arising from the 3d9 ground-state configuration of Ni+ have been reported for the deuterated form, NiD. We report on laser excitation and resolved fluorescence experiments that definitively locate rotational levels of v = 0–2 in the 2Σ+ state, and of v = 0 and v = 1 in the low-lying 2Π1/2 state of 58NiD.

The red bands of TaH: Identification of the ground and low-lying electronic states

Doppler-limited laser excitation spectra were recorded for 12 bands of the molecule tantalum hydride (TaH) in the red region, covering the wavelengths 605–665 nm. These excitation bands originate from three different lower electronic states. Dispersed fluorescence experiments revealed three additional low-lying electronic states of the molecule. The identities and energies of these six states are found to be in good agreement with the multiconfiguration self-consistent field calculations of Koseki et al. (2004). The ground state is X3Φ2, with the [0.1]0+ state lying only 76 ± 6 cm−1 above it. The upper states display energy shifts caused by numerous local perturbations with other dark states.

A hemicyanine and cucurbit[n]uril inclusion complex: competitive guest binding of cucurbit[7]uril and cucurbit[8]uril

ABSTRACTThe interaction between the hemicyanine indole derivative H and the cucubit[n]urils Q[7] and Q[8] has been studied using 1H NMR and UV spectroscopy as well as by fluorescence experiments. Competitive studies on the inclusion of H by Q[7] and Q[8] have also been conducted, and reveal that on changing the size of the Q[n] cavity, the binding behaviour can be very different.

The optical properties of 9-amino-9H-xanthene derivatives in different pH and their application for biomarkers in lysosome and mitochondria

Two xanthene compounds attached with 4-pyridinylimino (2a) and 2-quinolinylimino (2b) were prepared. Their deprotonated-protonated equilibrium between pyridine/quinoline/ xanthene units and the corresponding pyridinum/quinolinium/xanthylium units were evaluated in different pH by absorption and emission spectra. Probes 2a-b gave the colorimetric changes and emission responses at 550–750 nm, and their pKas (2a: pKa1 = 4.53, pKa2 = 5.95; 2b: pKa1 = 2.33, pKa2 = 6.18) were calculated by emission spectra. To verify the protonation process of probes 2a-b, the other two xanthylene derivatives attached with pyridinium (3a) and quinolinium (3b) were designed and synthesized; they exhibited only the equilibrium between xanthene and xanthylium as expected. Probe 3a showed a ratiometric change at 525–750 nm, and probe 3b displayed an OFF–ON emission at 500–700 nm with the decrease of the solution acidity. The calculated pKas were 6.18 and 1.49, respectively. Moreover, the fluorescent imaging experiments indicated that probes 2a–b were lysosome biomarkers while probes 3a–b were mitochondrial biomarkers for HeLa cells.

Spectroscopic methodologies and molecular docking studies on the interaction of antimalarial drug piperaquine and its metabolites with human serum albumin.

Artemisinin-based combination therapy is widely used for the treatment of uncomplicated Plasmodium falciparum malaria, and piperaquine (PQ) is one of the important partner drugs. During the biotransformation of PQ, M1 (N-oxidation product), M2 (N-oxidation product), M3 (carboxylic acid product), M4 (N-dealkylation product), and M5 (N-oxidated product of M4) are formed by cytochrome P450 pathways. Despite decades of clinical use, the interactions between PQ and its main metabolites (PQs) with human serum albumin (HSA) have not been reported. In the present study, the binding of PQs with HSA under physiological conditions was investigated systematically through fluorescence, circular dichroism (CD) spectroscopy, and molecular docking methods. The experimental results show that the intrinsic fluorescence quenching of HSA was induced by those compounds resulting from the formation of stable HSA-compound complexes. The main forces involved in the interactions between PQ, M1, and M2 which bind to HSA were hydrogen s and van der Waals forces, while the interactions of M3, M4, and M5 were driven by hydrophobic forces. The main binding sites of the compounds to HSA were also examined by classical fluorescent marker experiments and molecular docking studies. Binding constants (Kb) revealed that the affinities of the PQ, M1, M2, M3, and M4 to HSA were stronger than that of M5. Additionally, the binding rates of PQs with HSA were determined by ultrafiltration methods. Consistent with the binding constant results, the binding rate of M5 was lower than the binding rates of PQ, M1, M2, M3, and M4. Furthermore, PQs binding to HSA led to conformational and structural alterations of HSA, as revealed by multi-spectroscopic studies. In order to investigate one possible mechanism by which PQs inhibit the growth of malaria-causing Plasmodium parasites, 1H NMR spectroscopy was performed to investigate the interaction of the PQs with heme. This study is beneficial to enhance our understanding of the ecotoxicology and environmental behaviors of PQ and its metabolites.

Zinc(II) complexes of indole thiosemicarbazones: DNA/protein binding, molecular docking and in vitro cytotoxicity studies

Four Zn(II) complexes (1–4) featuring indole thiosemicarbazones were synthesized and well-characterized by elemental analyses and various spectroscopic (UV–Vis, FT-IR, 1H NMR, 13C NMR and mass) techniques. The single crystal X-ray crystallographic study revealed a tetrahedral geometry for complexes 1 and 3. The interactions of the Zn(II) complexes 1–4 with calf thymus DNA (CT DNA) were examined by UV–Vis spectroscopy and viscosity measurements, which implied that the complexes bound to CT DNA via intercalation, and complex 4 showed a higher binding affinity than the other complexes. The protein binding interaction of the complexes was monitored by fluorescence and absorption techniques, which showed that the complexes could bind effectively with bovine serum albumin (BSA) and the type of quenching mechanism was found to be static. Synchronous fluorescence experiments showed the changes in the conformations of the protein micro regions. Molecular docking studies were performed in order to get a better picture of the binding of the complexes with the molecular targets DNA hexamer and BSA. The in vitro cytotoxicity of the complexes against two human cancer (A549 and MCF7) cell lines, two human non-tumorigenic (MCF-10A and HEK-293) cell lines and one non-cancerous mouse fibroblasts (L929) cell line was evaluated using an MTT assay. Complex 4, which has an N-terminal cyclohexyl group, showed moderate activity [IC50 = 37.9 (A549) and 60.3 μM (MCF7)] that was comparable with the familiar anticancer drug cisplatin. Also, fortunately, the activity of complex 4 was found to be specific to cancer cells. The apoptosis cell death mechanism for complex 4 was assessed by the Hoechst staining method.

Silicon Wafer Functionalization with a Luminescent Tb(III) Coordination Complex: Synthesis, Characterization, and Application to the Optical Detection of NO in the Gas Phase.

A new luminescent Tb-DOTAGA (1,4,7,10-tetraazacyclododecane-1-glutaric-4,7,10- triacetic acid) complex (TbL) was synthesized and covalently immobilized on a silicon wafer. The grafting process was monitored by means of IR and XPS spectroscopies and the optical properties of the functionalized silicon wafer (TbL@Si) were investigated by fluorescence experiments. A homemade setup was then implemented in order to follow TbL@Si optical properties in the presence of gaseous nitric oxide (NO). The prima facie results indicated that in the presence of NO, the wafer fluorescence was partially quenched. This quenching was reversible as soon as NO was pumped outside the fluorescence cell, which could be interesting for the further development of lanthanide labelled silicon wafers as gas phase sensors.

Surface solvation and hindered isomerization at the water/silica interface explored with second harmonic generation.

Resonantly enhanced second harmonic generation (SHG) spectra of Coumarin 152 (C152) adsorbed at the water-silica interface show that C152 experiences a local dielectric environment slightly more polar than that of bulk water. This result stands in contrast to recently reported time-resolved fluorescence experiments and simulations that suggest an alkane-like permittivity for interfacial water at strongly associating, hydrophilic solid surfaces. Taken together, these results imply that while the static electric field across the aqueous-silica interface may be large, restricted water dynamics lead to apparent nonpolar solvation behavior similar to that experienced by solutes in confinement. Resonance-enhanced SHG spectra and time-resolved fluorescence of C152 adsorbed to aqueous-hydrophobic silica surfaces show that when water's ability to hydrogen bond with the silica surface is eliminated, a solute's interfacial solvation and corresponding ability to photoisomerize converge to an intermediate limit similar to that experienced in bulk acetone or methanol. While water structure and dynamics at solid-liquid interfaces have received considerable attention, results presented below show how strong solvent-substrate interactions can create conflicting pictures of solute reactivity across buried interfaces.