Rose Bengal‐Sensitized Photooxidation of the Dipeptides l‐Tryptophyl‐l‐Phenylalanine,l‐Tryptophyl‐l‐Tyrosine and l‐Tryptophyl‐l‐Tryptophan: Kinetics,Mechanism and Photoproducts¶

The Rose Bengal‐sensitized photooxidations of the dipeptides l ‐tryptophyl‐l ‐phenylalanine (Trp‐Phe), l ‐tryptophyl‐l ‐tyrosine (Trp‐Tyr) and l ‐tryptophyl‐l ‐tryptophan (Trp‐Trp) have been studied in pH 7 water solution using static photolysis and time‐resolved methods. Kinetic results indicate that the tryptophan (Trp) moiety interacts with singlet molecular oxygen (O₂(¹Δ_g)) both through chemical reaction and through physical quenching, and that the photooxidations can be compared with those of equimolecular mixtures of the corresponding free amino acids, with minimum, if any, influence of the peptide bond on the chemical reaction. This is not a common behavior in other di‐ and polypeptides of photooxidizable amino acids. The ratio between chemical (k_r) and overall (k_t) rate constants for the interaction O₂(¹Δ_g)‐dipeptide indicates that Trp‐Phe and Trp‐Trp are good candidates to suffer photodynamic action, with k_rlk_t values of 0.72 and 0.60, respectively (0.65 for free Trp). In the case of Trp‐Tyr, a lower k_rlk_t value (0.18) has been found, likely as a result of the high component of physical deactivation of O₂(¹Δ_g) by the tyrosine moiety. The analysis of the photooxidation products shows that the main target for O₂(¹Δ_g) attack is the Trp group and suggests a much lower accumulation of kynurenine‐type products, as compared with free Trp. This is possibly because of the occurrence of another accepted alternative pathway of oxidation that gives rise to 3a‐oxidized hydrogenated pyrrolo[2,3‐b]indoles.     

Photoreaction of Ketoprofen with Tryptophan and Tyrosine in Phosphate Buffer Solution

Reaction of excited ketoprofen (KP) with tryptophan (Trp) and tyrosine (Tyr) in a phosphate buffer solution was studied by the transient absorption spectroscopy. Both amino acids, which would interact with KP in bovine serum albumin [Monti, S. [2009] Phys. Chem. Chem. Phys., 11, 9104–9113], accelerated the proton transfer reaction to yield 3‐ethylbenzophenone ketyl biradical (EBPH) from KP carbanion, which was produced by photoexcitation of KP^? through decarboxylation. By means of the actinometry method with benzophenone, the reaction quantum yield was successfully estimated to be fairly large, and Trp, Tyr, DOPA and 4‐methylphenol were found to be a good proton donor for the carbanion. The formation rate constants of EBPH by the amino acids (k_r) were also determined to be (2.7 ± 0.1) × 10⁹ M^?1s^?1 for Trp and (7.8 ± 0.4) × 10⁸ M^?1s^?1 for Tyr, which were larger than those by basic amino acids and dipeptides reported. The reason for the highly efficient proton transfer reaction with Trp and Tyr would be explained by difference of the activation energy for the reaction. These results suggest that the proton transfer should be a key process for an initial photoreaction of KP with a protein, causing photosensitization in vivo.     

Chiral Sensing of Various Amino Acids Using Induced Circularly Polarized Luminescence from Europium(III) Complexes of Phenanthroline Dicarboxylic Acid Derivatives

Circularly polarized luminescence (CPL) was observed from [Eu(dppda)₂]^? (dppda=4,7‐diphenyl‐1,10‐phenanthroline‐2,9‐dicarboxylic acid) and [Eu(pzpda)₂]^? (pzpda=pyrazino[2,3‐f][1,10]phenanthroline‐7,10‐dicarboxylic acid) in aqueous solutions containing various amino acids. The selectivity of these complexes towards amino acids enabled them to be used as chiral sensors and their behavior was compared with that of [Eu(pda)₂]^? (pda=1,10‐phenanthroline‐2,9‐dicarboxylic acid). As these Eu^III complexes have achiral D_2d structures under ordinary conditions, there were no CPL signals in the emission assigned to f–f transitions. However, when the solutions contained particular amino acids they exhibited detectable CPL signals with g_lum values of about 0.1 (g_lum=CPL/2 TL; TL=total luminescence). On examining 13 amino acids with these three Eu^III complexes, it was found that whether an amino acid induced a detectable CPL depended on the Eu^III complex ligands. For example, when ornithine was used as a chiral agent, only [Eu(dppda)₂]^? exhibited intense CPL in aqueous solutions of 10^?2 mol dm^?3. Steep amino acid concentration dependence suggested that CPL in [Eu(dppda)₂]^? and [Eu(pzpda)₂]^? was induced by the association of four or more amino acid molecules, whereas CPL in [Eu(pda)₂]^? was induced by association of two arginine molecules.     

A laser flash photolysis study of amino acids and dipeptides using 4-nitroquinoline 1-oxide as a photosensitizer: The pH dependence

The pH effects on the photochemical reaction of amino acids and related dipeptides with 4-nitroquinoline 1-oxide (4NQO) as a photosensitizer have been investigated by laser flash photolysis. The obtained kinetic parameters show that the electron transfer from Tryptophan (Trp), Tyrosine (Tyr) as well as dipeptides containing Trp and/or Tyr residue to triplet 4NQO (^T4NQO) are efficient, but inefficient from methionine (Met) and dipeptides containing neither Trp nor Tyr. The result was supported by the calculated values of the free energy change from measured oxidation potentials for the electron transfer. It was demonstrated that Trp and Tyr residues are initial reaction sites with ^T4NQO, while Tyr/O^? radical may be final species for Trp-Tyr dipeptide. In acidic aqueous solutions, the self-quenching rate constants of ^T4NQO and the rate constants of electron transfer from amino acids to ^T4NQO decrease with decreasing pH. In alkaline solutions, amino acids are easily oxidized by 4NQO under irradiation of laser pulse, and no transient absorption signal was observed.     

PHOSPHORESCENCE LIFETIME STUDIES OF INTERACTIONS BETWEEN SERUM ALBUMINS AND SODIUM DODECYL SULFATE

Binding of sodium dodecyl sulfate (SDS) to bovine serum albumin (BSA) and human serum albumin (HSA) in aqueous solutions at room temperature induces significant changes in the phosphorescence lifetime of tryptophan (Trp) residues. A steep rise of the phosphorescence lifetime from 1.9 ms to 10.0 ms for BSA and from 1.9 ms to 5.5 ms for HSA is observed when the total SDS concentration increased from 0.0 mM to 0.22 mM at 1 mg/mL protein concentration. As the total SDS concentrationis further inccreased to 2.2 mM, a slower increase in the phosphorescence lifetime is observed, from 10.0 ms to 19.5 ms for BSA and from 5.5 ms to 7.2 ms for HSA. It appears that the phosphorescence lifetime modifications are mainly due to an increase of protein matrix rigidity around Trp residues. The observed differences (between HSA and BSA) allow us to distinguish the contribution of the two Trp residues to the BSA phosphorescence.     

Quenching of the Singlet and Triplet Excited States of Pterin by Amino Acids

Unconjugated oxidized pterins accumulate in the skin of patients suffering from vitiligo and, under UVA irradiation, photosensitize the oxidation of amino acids. In this work, we study the interaction of the singlet and triplet excited states of pterin (Ptr), the parent compound of oxidized pterins, with four oxidizable amino acids: tryptophan (Trp), tyrosine (Tyr), histidine (His) and methionine (Met). Steady‐state and time‐resolved fluorescence measurements and laser flash photolysis experiments were performed to investigate the quenching of the Ptr excited states by the amino acids in aqueous solution. The singlet excited states of Ptr are quenched by Met mainly via a dynamic process and by Trp via a combination of dynamic and static processes. His does not quench singlet excited states of Ptr, and quenching by Tyr could not be investigated due to the low solubility of this amino acid. The triplet excited states of Ptr are quenched by the four studied amino acids, and the corresponding bimolecular quenching rate constants are in the range of diffusion controlled limit. The assessment of the results in the context of the Ptr‐photosensitization of amino acids suggests that triplet excited state of Ptr is the species that initiates the photochemical processes.     

DRUG SENSITIZATION: PHOTOBINDING OF SULFANILAMIDE TO BOVINE SERUM ALBUMIN

Abstract— Photobinding of sulfanilamide to bovine serum albumin (BSA) was investigated by irradiating BSA and buffered BSA/drug solutions with UV light (Λ= 300 nm) under anaerobic conditions. The protein solutions were then denatured and the unbound sulfanilamide removed. Marked differences in the UV and fluorescence spectra of the solutions before and after irradiation were observed, suggesting covalent binding of the drug to BSA. This was confirmed using [¹⁴C]labelled sulfanilamide. The extent of photobinding of sulfanilamide determined using the radiolabeled drug, was concentration dependent. The binding ratio varied from 3 mol drug per mol BSA for a 10^-4 M drug concentration, to 10 mol drug per mol BSA for 10^-2 M drug concentration.
The protein solutions were hydrolysed under acid conditions and the amino acids obtained were analysed by ion exchange chromatography. The hydrolysate of irradiated BSA (10^-4 M ) -sulfanilamide (10^-2 M ) mixture lost about 10 mol of cystine per mol of BSA. This loss was not observed after hydrolysis of irradiated alone or non-irradiated BSA. Irradiation of cystine with [¹⁴C]sulfanilamide in HC1 solutions produced the same compound as was found after hydrolysis of irradiated BSA/sulfanilamide mixtures. This was demonstrated by autoradiography of paper chromatograms. The same compound was also detected in an irradiated [³⁵S]cystine non-labelled sulfanilamide mixture. It was not detected, however, after irradiation of a mixture of all amino acids of BSA excluding cystine. These data suggest that cystine residues are involved in the photobinding of sulfanilamide (or its photoproducts) to BSA.     

The effect of glucose on bovine serum albumin denatured aggregation kinetics at high concentration

The effect of glucose (0–15 mass%) on the kinetics of bovine serum albumin (BSA) denatured aggregation at high concentration in aqueous solution has been studied by differential scanning calorimetry. The observed denatured aggregation process was irreversible and could be characterized by a denaturation temperature (T _m), apparent activation energy (E _a), the approximate order of reaction, and pre-exponential factor (A). As the glucose concentration increased from 0 to 15 mass%, T _m increased, E _a also increased from 514.59409±6.61489 to 548.48611±7.81302 kJ mol⁻¹, and A/s⁻¹ increased from 1.24239E79 to 5.59975E83. The stabilization increased with an increasing concentration of glucose, which was attributed to its ability to alter protein denatured aggregation kinetics. The kinetic analysis was carried out using a composite procedure involving the iso-conversional method and the master plots method. The iso-conversional method indicated that denatured aggregation of BSA in the presence and absence of glucose should conform to single reaction model. The master plots method suggested that the simple order reaction model best describe the process. This study shows the combination of iso-conversional method and the master plots method can be used to quantitatively model the denatured aggregation mechanism of the BSA in the presence and absence of glucose.     

Study of interaction of Potassium Dodecatangestato Cobaltate(III) with Bovine Serum Albumin using Fluorescence Spectroscopy

The binding of potassium dodecatangestato cobaltate(III) (PDC) as a water-soluble polyoxometal with bovine serum albumin (BSA) as a major transporting protein of plasma, has been investigated at pH 7.2, 5?mM phosphate buffer, 27°C and various ionic strength by fluorescence spectroscopy.

The results show that the binding of PDC to BSA quenches BSA emission and the Stern–Volmer linear relationship can be applied for the quenching process.

The values of Stern–Volmer constant, K _sv, which can be considered as a binding constant for formation of 1:1 complex at 0.01, 0.1 and 0.2?M NaCl are 8.56 × 10⁵, 5.72 × l0⁵ and 9.60 × 10⁵, respectively. The interpretation of the results represents that binding affinity depends on both electrostatic forces and conformational stability of BSA. A step-by-step aggregation model, which has been developed by Borissevich et al., was used to determine the average aggregation number of BSA, ?J?, from the fluorescence quenching. The results show that the binding of PDC to BSA does not induce any considerable aggregation in BSA molecules. Therefore, it can be concluded that there are no considerable conformational changes in BSA molecules during its interaction with PDC.     

Interaction of albumin with polysaccharides containing ionic groups

It is very important to clarify the mechanisms of the interaction between components of organisms. In this report, the interaction between bovine serum albumin (BSA) and ionic polysaccharides were discussed. The fluorescence spectrum of tryptophan (Trp) in BSA changed as its conformation changed. On adding polysaccharide containing sulfonic acid residues (sulfonic‐type) at pH 7.4, a remarkable blue shift of the emission maximum (λ_em) of Trp was observed. This blue shift was decreased by adding NaCl. In contrast, polysaccharide containing carboxylic acid residues (carboxy‐type) scarcely changed Trp fluorescence at the same pH. At a pH lower than the isoelectric point (PI = 4.7–4.9) of BSA, a remarkable blue shift was observed not only by adding sulfonic‐type polysaccharide but also by adding carboxy‐type polysaccharide. Moreover, using the energy transfer method, in the pH region higher than the PI of BSA, carboxylic‐type polysaccharides may interact relatively weakly with the binding site II of BSA, but sulfonic‐type ones can selectively interact with binding site I weakly and with binding site II strongly. And in the pH region lower than the PI of BSA, carboxylic‐type polysaccharides interact with binding site II strongly. On the other hand, sulfonic‐type polysaccharides are bound to both binding site I and binding site II very strongly. Copyright © 1999 John Wiley & Sons, Ltd.     

Highly Efficient Nondoped OLEDs with Negligible Efficiency Roll‐Off Fabricated from Aggregation‐Induced Delayed Fluorescence Luminogens

Purely organic emitters that can efficiently utilize triplet excitons are highly desired to cut the cost of organic light‐emitting diodes (OLEDs), but most of them require complicated doping techniques for their fabrication and suffer from severe efficiency roll‐off. Herein, we developed novel luminogens with weak emission and negligible delayed fluorescence in solution but strong emission with prominent delayed components upon aggregate formation, giving rise to aggregation‐induced delayed fluorescence (AIDF). The concentration‐caused emission quenching and exciton annihilation are well‐suppressed, which leads to high emission efficiencies and efficient exciton utilization in neat films. Their nondoped OLEDs provide excellent electroluminescence efficiencies of 59.1 cd A⁻¹, 65.7 lm W⁻¹, and 18.4 %, and a negligible current efficiency roll‐off of 1.2 % at 1000 cd m⁻². Exploring AIDF luminogens for the construction of nondoped OLEDs could be a promising strategy to advance device efficiency and stability.     

Spectroscopic studies,structural characterization and electrochemical studies of two cobalt (III) complexes with tridentate hydrazone Schiff base ligands: Evaluation of antibacterial activities,DNA‐binding,BSA interaction and molecular docking

Co(III) complexes of tridentate Schiff base ligands derived from N‐(2‐hydroxybenzylideneamino)benzamide (H ₂ L ¹ ) and 2‐(2‐hydroxybenzylidene)hydrazine‐1‐carboxamide ( H ₂ L ² ) were synthesized and characterized using IR, Raman, ¹H–NMR and UV–Vis spectroscopies. X‐ray single crystal structures of complexes 1 and 2 have also been determined, and it was indicated that these Co(III) complexes are in a distorted octahedral geometry. The cyclic voltammetry (CV) of the complexes indicates an irreversible redox behavior for both complexes 1 and 2 . The antibacterial effects of the synthesized compounds have been tested by minimum inhibitory concentration and minimum bactericidal concentration methods, which suggested that the metal complexes exhibit better antibacterial effects than the ligands against Gram‐positive bacteria. The effects of the drug (drug  =  ligands and complexes) on bovine serum albumin (BSA) were examined using circular dichroism (CD) spectropolarimetry, and it was revealed that the BSA (BSA, as a carrier protein) secondary structure changed in the presence of the drug. Interaction of the drug with calf‐thymus DNA (CT‐DNA) was investigated by UV–Vis absorption, fluorescence emission, CV and CD spectroscopy. Binding constants were determined using UV–Vis absorption. The results indicated that the studied Schiff bases bind to DNA, with the hyperchromic effect and non‐intercalative mode in which the metal complexes are more effective than ligands. Furthermore, molecular docking simulation was used to obtain the energetic and binding sites for the interaction of the complexes with Mycobacterium tuberculosis enoyl‐acyl carrier protein reductase (InhA), and results showed that complex 1 has more binding energy.     

Multi-spectroscopic Study on the Interaction between CdTe Quantum Dots and Bovine Serum Albumin

The interaction between CdTe quantum dots (QDs) and bovine serum albumin (BSA) was systematically investigated by fluorescence, UV‐vis absorption and circular dichroism (CD) spectroscopy under physiological conditions. The experimental results showed that the fluorescence of BSA could be quenched by CdTe QDs with a static quenching mechanism, indicating that CdTe QDs could react with BSA. The quenching constants according to the modified Stern‐Volmer equation were obtained as 1.710×10⁶, 1.291×10⁶ and 1.010×10⁶ L·mol^?1 at 298, 304, and 310 K, respectively. ΔH, ΔS and ΔG for CdTe QDs‐BSA system were calculated to be ?33.68 kJ·mol^?1, 6.254 J·mol^?1·K^?1 and ?35.54 kJ·mol^?1 (298 K), respectively, showing that electrostatic interaction in the system played a major role. According to F?rster theory, the distance between Trp‐214 in BSA and CdTe QDs was given as 2.18 nm. The UV‐vis, synchronous fluorescence and CD spectra confirmed further that the conformations of BSA after addition of CdTe QDs have been changed.     

Tetrahedraphene: A Csp3-centered 3D Molecular Nanographene Showing Aggregation-Induced Emission

The bottom-up synthesis of 3D tetrakis(hexa-peri-hexabenzocoronenyl)methane, “tetrahedraphene”, is reported. This molecular nanographene constituted by four hexa-peri-hexabenzocoronene (HBC) units attached to a central sp³ carbon atom, shows a highly symmetric arrangement of the HBC units disposed in the apex of a tetrahedron. The X-ray crystal structure reveals a tetrahedral symmetry of the molecule and the packing in the crystal is achieved mostly by CH⋅⋅⋅π interactions since the interstitial solvent molecules prevent the π⋅⋅⋅π interactions. In solution, tetrahedraphene shows the same electrochemical and photophysical properties as the hexa-^tBu-substituted HBC (^tBu-HBC) molecule. However, upon water addition, it undergoes a fluorescence change in solution and in the precipitated solid, showing an aggregation induced emission (AIE) process, probably derived from the restriction in the rotation and/or vibration of the HBCs. Time-Dependent Density Functional Theory (TDDFT) calculations reveal that upon aggregation, the high energy region of the emission band decreases in intensity, whereas the intensity of the red edge emission signal increases and presents a smoother decay, compared to the non-aggregated molecule. All in all, the excellent correlation between our simulations and the experimental findings allows explaining the colour change observed in the different solutions upon increasing the water fraction.     

Nano‐Porous Light‐Emitting Silicon Chip as a Potential Biosensor Platform

Abstract

Nano‐porous silicon (PS) offers a potential platform for biosensors with benefits both in terms of light emission and the large functional surface area. A light emitting PS chip with a stable and functional surface was fabricated in our laboratory. When protein was deposited on it, the light emission was reduced in proportion to the protein concentration. Based on this property, we developed a rudimentary demonstration of a label‐free sensor to detect bovine serum albumin (BSA). A serial concentration of BSA was applied to the light chip and the reduction in light emission was measured. The reduction of the light intensity was linearly related to the concentration of the BSA at concentrations below 10^?5 M. The detection limit was 8×10^?9 M.     

Deciphering the role of pH in the binding of ciprofloxacin hydrochloride to bovine serum albumin

The effect of the added fluoroquinolone, Ciprofloxacin Hydrochloride (CpH), on structural properties of Bovine Serum Albumin (BSA) was investigated by Circular Dichroism (CD), steady-state, time-resolved and Dynamic Light Scattering (DLS) spectroscopic approaches. The intrinsic fluorescence of the Tryptophan (Trp) amino acid residue in the globular protein BSA was made use of and the effect of pH at two different temperatures was thoroughly investigated. CD results indicate that CpH induces some structural changes in BSA and this has been well-supported by steady-state, lifetime and DLS data. The fluorescence intensity of Trp gradually decreases with the rise in concentration of CpH and we have conclusively proved that at pH 7.4 and 9.2, the mechanism of fluorescence quenching is mostly dynamic in nature, whereas at pH 4.5 mainly static quenching is operational. Thermodynamic parameters have been studied to rationalize the nature of binding of CpH to BSA, and we have concluded that hydrophobic and van der Waals forces play an important role in the process of drug-protein interaction at three different pH values. The lifetime of Trp was found to decrease with the rise in CpH concentration and the percentage reduction in lifetime was found to be a function of the pH of the medium under investigation.     

Synthesis,structure and in vitro antiproliferative activities of oxamido‐bridged dicopper(II) complexes: A comparative study of experimental evidence and molecular docking of DNA/protein binding

Two μ‐oxamido‐bridged dicopper(II) complexes, namely [Cu₂(hmpoxd)(H₂O)(phen)](ClO₄) ( 1 ) and [Cu₂(papo)(H₂O)(phen)](ClO₄)·2H₂O ( 2 ), where H₃hmpoxd and H₃papo represent N‐(2‐hydroxy‐5‐methylphenyl)‐N′‐[3‐(dimethylamino)propyl]oxamide and N‐(2‐hydroxylphenyl)‐N′‐(3‐aminopropyl)oxamide, respectively, and phen represents 1,10‐phenanthroline, were synthesized. Single‐crystal X‐ray crystallography and other methods revealed that the two copper(II) ions in complex 1 are bridged by the cis‐hmpoxd^3? with Cu···Cu separation of 5.1896(7) Å, in which the inner (Cu1) and outer (Cu2) copper(II) atoms are located in square‐planar and square‐pyramidal geometries, respectively. To evaluate the effects of bridging ligand hydrophobicity on DNA/protein binding and potential anticancer activities, comparative studies of the reactivity towards herring sperm DNA and protein bovine serum albumin (BSA) as well as cytotoxicity of complex 1 with our previously reported complex 2 were conducted theoretically and experimentally. The results indicate that the two complexes can interact interactively with DNA, and bind to BSA via the binding sites Trp213 for 1 and Trp134 for 2 . Interestingly, the in vitro anticancer activities and DNA/protein binding affinities consistently follow the order of 1 > 2 .     

Evaluation of the influence of arsenic species on the nitrogen metabolism of a model angiosperm: nasturtium,Tropaeolum majus

How the various organic and inorganic arsenic species affect the nitrogen metabolism of a model plant, Tropaeolum majus, was studied in order to evaluate the toxicological impact of the various chemical forms of arsenic. For this purpose, the effects on the (a) entire nitrogen pool, (b) protein fraction, and (c) non‐protein fraction were distinguished. The arsenic‐dependent effects on the nitrogen cycle were assessed by using ¹⁵N‐labelled KNO₃ as a nutritive substance and optical emission spectroscopy to analyse how ¹⁵N is incorporated into the nitrogen cycle. In addition to the ¹⁵N‐tracer experiments, the uptake and metabolization of the arsenic compounds were examined. The work shows that biochemical indicator systems like ¹⁵N‐tracer studies are able to characterize the degree of the influence of metabolic processes by arsenic species. For example, the incorporated ¹⁵N concentration decreased linearly and independently of the ¹⁵N fraction with increasing dimethylarsinate (DMA) concentrations. This behaviour indicates that DMA has prevented the uptake of ¹⁵N and hence the formation of amino acids and proteins. Arsenite‐treated plants exhibited an elevated concentration of non‐protein ¹⁵N, which could be an indication either for a stimulated uptake of nitrate or for an interrupted amino acid/protein synthesis. Copyright © 2005 John Wiley & Sons, Ltd.