Binding mode of proflavine to DNA probed by polarized light spectroscopy

It is well known that proflavine binds to DNA. Here we investigate the binding mode of proflavine to native DNA using absorption, circular dichroism (CD), and linear dichroism (LD) spectroscopy as well as by fluorescence techniques. The observed changes of proflavine upon complexation with DNA can be summarized as a red shift and hypochromism in the absorption spectrum. The negative LD^r in the proflavine absorption region has a magnitude comparable to or larger than that of the DNA absorption region, confirming the intercalative binding mode of proflavine to DNA. Saturation of the LD spectrum in the proflavine absorption region at R = 0.25 and a decrease in the fluorescence intensity provide further evidence of intercalation. Furthermore, the coupling of electric transition of intercalated proflavine is observed. Although proflavine has been reported to position itself along the DNA stem at high [proflavine]/[DNA base] ratios, the spectral characteristics, which include a clear isosbestic point in the absorption spectrum and proportionality in the LD magnitude in the proflavine absorption region, do not show any possibility of exterior binding.     

Femtosecond fluorescence upconversion studies of excited-state proton-transfer dynamics in 2-(2′-hydroxyphenyl)benzoxazole (HBO) in liquid solution and DNA

A femtosecond fluorescence upconversion study is reported for HBO in solution, as well as for HBO incorporated in DNA. The typical time for the excited-state intramolecular proton-transfer reaction of the syn-enol tautomer in solution and in DNA has been determined to be 150 fs. In addition, the lifetimes of the keto, the anti-enol and the ‘solvated enol’ tautomer forms were determined in protic solvents, aprotic solvents and DNA. Picosecond rise and decay components in the fluorescence transients with characteristic times between 3 and 25 ps are also observed and attributed to the effects of vibrational cooling.     

Ratiometric detection of pH fluctuation in mitochondria with a new fluorescein/cyanine hybrid sensor

The homeostasis of mitochondrial pH (pH_m) is crucial in cell physiology. Developing small-molecular fluorescent sensors for the ratiometric detection of pH_m fluctuation is highly demanded yet challenging. A ratiometric pH sensor, Mito-pH, was constructed by integrating a pH-sensitive FITC fluorophore with a pH-insensitive hemicyanine group. The hemicyanine group also acts as the mitochondria targeting group due to its lipophilic cationic nature. Besides its ability to target mitochondria, this sensor provides two ratiometric pH sensing modes, the dual excitation/dual emission mode (D_ex/D_em) and dual excitation (D_ex) mode, and its linear and reversible ratiometric response range from pH 6.15 to 8.38 makes this sensor suitable for the practical tracking of pH_m fluctuation in live cells. With this sensor, stimulated pH_m fluctuation has been successfully tracked in a ratiometric manner via both fluorescence imaging and flow cytometry.     

Rapid separation of four probiotic bacteria in mixed samples using microchip electrophoresis with laser-induced fluorescence detection

Microchip electrophoresis (MCE) coupled to laser-induced fluorescence detection was applied to the rapid separation of Bifidobacterium, Lactobacillus casei, Lactobacillus acidophilus, and Enterococccus faecalis. All bacteria were quickly separated within 150?s using a running buffer of pH 8.5 containing Tris, borate, EDTA, and poly(ethylene oxide). The latter was crucial to reduce the bacterial adsorption on the walls of the microchannels. The pH of 8.5 warrants that bacteria carry a negative charge at their surface and thus display good electrophoretic performance. The method was used to analyze medical samples containing these probiotics, and the results showed that the identification and detection of bacteria by MCE is advantageous in terms of sample consumption, waste production, time of analysis, and instrumental effort.

Diphotonic one-electron oxidation of NADH on laser excitation at 353 nm

The primary steps in one-electron oxidation of NADH (the reduced form of nicotinamide adenine dinucleotide) were studied by means of nanosecond laser photolysis combined with kinetic spectrophotometry. High-intensity excitation at 353 nm of NADH in aqueous solution (pH 10.0) was found to produce two-photon ionization at the nicotinamide group. The absorption spectrum of the -NADH⁺ radical cation formed in this reaction was determined. This species disappeared by deprotonation at a rate of 3.6 × 10⁶ s^?1 at pH 10 forming -NAD.     

Regularities of the sorption behavior of actinide ions on mineral colloid particles

It is shown that an alternative to K _d in describing sorption at low degrees of surface saturation of colloid particles is pH₅₀ which takes into account both the properties of the sorbent and sorbate. The correlations of pH₅₀ with cation charge density for the An(III)-An(IV)-An(V)-An(VI) series and with hydrolysis constants are presented. The redox reactions with plutonium that accompany its sorption onto Fe(III) oxides are discussed.     

Spin probe study of acidity of inorganic materials

Acidic characteristics of various inorganic materials, such as silica gel, aluminum, and titanium(IV) oxides, original and modified with F^? and SO ₄ ^2? ions, in the form of hydrogels and nanostructured powders doped by copper(II) ions, are studied. It is shown that pH values inside pores of the studied sorbents (pH_intr) differ from the pH of external solutions by 0.5–1.1 units. In the pores of original γ-Al₂O₃, the pH_intr values are higher, while, in pores of γ-Al₂O₃ samples modified with F^? и SO ₄ ^2? , as well as nonmodified α-Al₂O₃, SiO₂ gel, and TiO₂ hydrogels, pH_intr are lower compared to the pH of the external solution. It is found that pH_intr decreases, as the content of Cu²⁺ ions in the TiO₂ phase grows, and, beginning with a certain moment, it coincides with pH of an equilibrium solution. The pK _a values of functional group dissociation in some materials are determined. It is shown that, using spin probes, that is, pH-sensitive nitroxyl radicals, the electrostatic potential of the surface of nanostructured TiO₂ can be estimated.     

Time-programmable pH: decarboxylation of nitroacetic acid allows the time-controlled rising of pH to a definite value

In this report it is shown that nitroacetic acid 1 (O₂NCH₂CO₂H) can be conveniently used to control the pH of a water solution over time. Time-programmable sequences of the kind pH_1(high)–pH_2(low)–pH_3(high) can be achieved, where both the extent of the initial pH jump (pH_1(high)–pH_2(low)) and the time required for the subsequent pH rising (pH_2(low)–pH_3(high)) can be predictably controlled by a judicious choice of the absolute and relative concentrations of the reagents (acid 1 and NaOH). Successive pH_1(high)–pH_2(low)–pH_3(high) sequences can be obtained by subsequent additions of acid 1. As a proof of concept, the method is applied to control over time the pH-dependent host–guest interaction between alpha-cyclodextrin and p-aminobenzoic acid.

Predictable and time-programmable sequences of the kind pH_1(high)–pH_2(low)–pH_3(high) in water solution are obtained by a judicious choice of the concentration of nitroacetic acid undergoing decarboxylation.     

Influence ofpH and concentration of complexing agents on fluorescence of samarium(III), gadolinium(III), and terbium(III)—Ethylenediaminetetraacetic acid or nitrilotriacetic acid complexes in aqueous solutions

The fluorescence intensity of aqueous solutions of Sm(III), Gd(III) and Tb(III) complexes withEDTA orNTA was investigated as a function of complexone concentration over a widepH range. For Sm(III) and Tb(III) complexes the ratio of intensities of hypersensitive and allowed bands was calculated in order to obtain information about intensity borrowing of the fluorescence bands as a result of vibronic coupling.     

Challenges to computational quantum chemistry from contemporary advances in polyatomic molecular electronic spectroscopy

Molecular electronic spectroscopy featuring intramolecular proton transfer and twisted intramolecular charge transfer poses a whole new range of problems for computational quantum chemistry. The development of the four-level laser based on the intramolecular proton-transfer focuses on the subtleties of the interaction of the singlet and triplet electronic state manifolds of the two different tautomeric species. Examples are given of the sensitive variation of proton-transfer fluorescence with chemical substitution. A competing excitation channel is shown to exist when internal molecular torsion couples with sudden polarization to yield a twisted intramolecular charge transfer configuration. In such systems, three competing fluorescences can be observed. Several electronic puzzles will be presented that can provide fertile territory for quantum chemical computations. © 1993 John Wiley & Sons, Inc.     

The complexation of apo-metallothionein with cadmium ion and its conformation study

The circular dichroism was used to study the complexation of two isoforms of rabbit liver apo-metallothioneins (apo-MT1 and apo-MT2) with Cd²⁺ ion and the influence of Cd²⁺ ion on the conformation of reconstituted MTs. The stability of sulfhydryls of apo-MT has been investigated at the room temperature in the presence of air. The reconstitutions of apo-MT1 with Cd²⁺ ion were carried out atpH 4.71 (stable state) andpH 7.9 (with 90% sulfhydryls oxidated) respectively. it was found that the characteristic CD band at 257 nm(+), 238 nm(?), 226 nm(+) of reconstituted MT with Cd²⁺ ion was the same as native MT atpH 4.71, however only one peak at 243 nm(+) appeared on the CD spectra atpH 7.9 which arose from mononuclear complexes with four separated thiolate ligands per Cd²⁺ ion. The CD spectra of apo-MTs+7 eq Cd²⁺ system were measured at variouspH values. It was found that the peak at 256 nm of apo-MT1 binding Cd²⁺ ion split into two small peaks atpH between 2.42 and 3.02, and became one peak atpH 3.32, while the shapes of Cd peaks of apo-MT2 binding Cd²⁺ ion did not change withpH, indicating that the binding sites and pathway of apo-MT1 binding Cd²⁺ ion were different from those of apo-MT2. A possible mechanism was suggested.     

Determination of pH value of isoelectric solution and identification of passive iron surface phases using immersion method

Using the method of phase modeling, the pH values of solutions corresponding to the uncharged surface of passive iron and ferric oxide γ-Fe₂O₃ (pH₀) are compared. According to the theory of connected places, the charge of metal oxide surface is determined by the adsorption or desorption of hydrogen ions leading to a change in the potential drop at the oxide/solution interface. Preliminarily passivated iron electrode was washed with twice-distilled water and placed into 0.5 M NaNO₃ solution with various pH values; the variation in the potential (ΔE) with time was studied. The pH₀ value for passive electrode under the open-circuit conditions was determined by the dependence of ΔE on the pH value (pH₀ 6.2 ± 0.1). The pH₀ value was close to that for γ-Fe₂O₃ (pH₀ 6.2), which was determined by the method of potentiometrical titration of oxide suspension in the nitrate solution. The introduction of surface-active ions Ba²⁺ and Cl^? changes the charge of passive iron surface: Ba²⁺ ions increase the electrode potential, while Cl^? ions decrease it. Comparing the pH₀ values for passive electrode and metal oxides, one can identify the composition of passive electrode surface.     

Tetrakis(4-sulfonatophenyl)porphyrin fluorescence as reporter of human serum albumin structural changes induced by guanidine hydrochloride

The interaction of the drug carrier protein human serum albumin (HSA) with the ionic, free base porphyrin tetrakis(4-sulfonatophenyl)porphyrin (TSPP) was investigated under chemical denaturation conditions using guanidine hydrochloride (Gdn-HCl) in aqueous solution at pH 7 and 2.5. Protein stability was studied by fluorescence spectroscopy using intrinsic tryptophan fluorescence, whereas far-UV circular dichroism gave information regarding conformational changes. Steady-state and time-resolved fluorescence as well as extinction and induced visible CD of TSPP were also monitored in the presence of the denaturant.The addition of 1.0 M Gdn-HCl inhibited the FRET process between the sole tryptophan residue of HSA and the porphyrin as inferred by an increase in the intrinsic fluorescence of the former together with a drop in the fluorescence of the latter. Simultaneously, an induced bisignate CD band was detected in the Soret region of TSPP extinction following the changes in the monomer ? aggregate equilibrium of TSPP. The features in the extinction spectra pointed to the formation of J-aggregates at pH = 2.5 and were confirmed by fluorescence lifetime measurements. At pH = 7, no TSPP dimers were detected in the absence of the protein or in the presence of native HSA. However, H-dimers or higher aggregates of TSPP associated to HSA were induced at concentrations of Gdn-HCl below 2 M.The main unfolding transition probed by HSA intrinsic fluorescence took place between 2 and 3 M Gdn-HCl at pH = 7, whereas at pH = 2.5 it was detected only above 2.8 M Gdn-HCl, coinciding with TSPP release into solution which occurs at high denaturant concentration for both pH studied. The results suggest that the chemical unfolding of HSA is a multistep process. The free base porphyrin contributes to an increase in the protein stability, particularly important under acidic conditions, where the protein is known to be in an expanded form in the absence of TSPP.The analysis of TSPP fluorescence fluctuations in the autocorrelation functions obtained using fluorescence correlation spectroscopy (FCS) in the presence of HSA at different denaturant concentrations showed that the porphyrin only interacts with the native form of the protein.Both fluorescence and circular dichroism data confirmed that in the noncovalent complex HSA–TSPP the free base porphyrin can act as a reporter of the protein structural changes induced either by pH or chemical denaturation.     

c-di-GMP can form remarkably stable G-quadruplexes at physiological conditions in the presence of some planar intercalators

The ubiquitous bacterial biofilm regulator, c-di-GMP can form G-quadruplexes at physiological conditions in the presence of some aromatic compounds, such as acriflavine and proflavine. The fluorescence of these compounds is quenched upon c-di-GMP binding and some of the formed c-di-GMP G-quadruplexes are stable even at 75 °C.     

Studies with dithizone: Part XXI. The Relationship between pH12 and Molar Solubility

The effect of changes of ionic strength on the partition of an analytically important weak acid between a variety of aqueous and immiscible organic phases has been studied with respect to new measurements on dithizone (3-mercapto-1,5-diphenylformazan) and di(p-tolyl)thiocarbazone. A linear relationship has been deduced between the pH at which 50% extraction takes place and S_r,o (the solubility of the reactant in the organic phase), and confirmed by measurements of pH_{$\text{1}\text{2}$}, and S_r,o for a large number of organic solvents. The solubilities of the reagents in water, S_r, are also reported. The form of the relationship between pH and log D, and between pH_{$\text{1}\text{2}$} and log S_r,o is the same whether a solution of dithizone consists of a single species or a mixture of thiol and thione forms in tautomeric equilibrium.     

Fluorescence properties of tetraphenylporphyrin-polypeptide complexes as model photosynthetic systems

The steady-state and time-resolved fluorescence spectra of complexes formed from α,β,γ,δ-tetrakis(4-carboxyphenyl) porphin and the synthetic sequential polytripeptides poly(L-lysyl-L-leucyl-L-alanine)_n (I) (MW 7900) and poly(L-lysyl-L-alanyl-L-alanine) _n (II) (MW 6900) are studied in phosphate buffer (pH=7.2). The peptide matrices were found to bond 4-5 pigment molecules per one polypeptide. The complexed pigment fluorescent decay kinetics is two-exponential with components τ₁ =240 ps and τ₂=2400 (I) and 3500 ps (II). The relative contributions of τ₁ correlate with the degree of α-helicity of the polypeptide matrix.     

Characteristics of the hydrogen ion-sensitive field effect transistors with sol-gel-derived lead titanate gate

The sol-gel-derived lead titanate (PbTiO₃) membrane has been successfully applied as a pH sensitive layer to form the PbTiO₃ gate ion-sensitive field-effect transistor (ISFET). There exhibit the excellent quasi-Nernstian response of 56-59 mV pH⁻¹, good surface adsorption and anticorrosion characteristics via the C-V measurement of the EIS structure. At a specific pH concentration, the output and transfer characteristics are very similar to the behaviours of MOSFETs, and the ISFET model can be derived by the modified MOSFET model. As it operated in the nonsaturation region, there exhibits a linear pH response of about 56-59 mV pH⁻¹. On the other hand, as it operated in the saturation region, the pH response and linearity can be controlled by adjusting the V_GS values, e.g. the pH responses of −4.2, −24.8 and −31.3 μA pH⁻¹ and the correlation coefficients of 0.9491, 0.9995 and 0.9996 at V_GS=1, 3 and 5 V can be obtained, respectively. Besides, in order to get the best pH response and the minimized leakage current, the heat treatment temperature of the PbTiO₃ membrane must be limited between 350 and 450 °C.     

UV laser induced proton-transfer of protein molecule in the gas phase produced by droplet-beam laser ablation

Droplet-beam laser-ablation mass-spectrometry was applied for a study of the UV-laser induced proton-transfer reaction of protonated lysozyme hydrated clusters in the gas phase. Protonated lysozyme hydrated clusters were produced by irradiation of an IR laser onto a droplet-beam of an aqueous solution of lysozyme and were subsequently irradiated by a UV laser. It is found that H⁺ and H₃O⁺ are produced through photodissociation of protonated lysozyme hydrated clusters. The mechanism of the proton-transfer reaction is discussed.     

Gleichgewichte und Kinetik zur 1:1-Komplexbildung von Aluminium(III) mit Kojisäure in wäßriger Lösung

The reaction of aluminium(III) with kojic acid was investigated in the range ofpH=(1.6 ... 3.4) at excess of Aluminium(III). Only the complex AlL ²⁺ was formed with β₁₁=7.6. The results of equilibrium and kinetic measurements indicate that only the species Al³⁺, AlOH²⁺ and the neutral form of the ligand contribute to the reaction. Reaction values and rate constants are given. A general acid base catalysis was detected.     

Understanding the mechanism of the N-heterocyclic carbene-catalyzed ring-expansion of 4-formyl-β-lactams to succinimide derivatives

The mechanism of the N-heterocyclic carbene (NHC)-catalyzed ring-expansion of 4-formyl-β-lactams to succinimides has been studied using DFT methods at the B3LYP/6-31G^∗∗ level. The first step is the nucleophilic attack of NHC to the aldehyde to yield the zwitterionic intermediate, which by a proton-transfer process affords the Breslow intermediate. The lactam N-C breaking bond in this intermediate yields an enol-amidate, which by a keto-enol type equilibrium becomes the ketone form. The subsequent ring-closure achieved by the nucleophilic attack of the amidate to carbonyl carbon allows the formation of the five-membered ring. Finally, elimination of NHC affords the succinimide. Analysis of the nucleophilicity index correctly explains the behaviors of the NHCs and the Breslow intermediates in the umpolung reactivity of aldehydes.