Fluorescence "off" and "on" signalling of esculetin in the presence of copper and thiol: a possible implication in cellular thiol sensing.

The interaction of the cupric ion with esculetin, a dihydroxy coumarin derivative, was studied by absorption and fluorescence spectroscopic methods in aqueous medium. Esculetin formed a complex in the presence of the cupric ion which was characterised by the shift in its absorption band from 350 nm to 389 nm and the quenching of its fluorescence intensity at 466 nm. From Job's plot and fluorescence quenching studies, the stoichiometry of the copper ion and esculetin in the complex was estimated to be 1 : 2 respectively. Interestingly, the incubation of the Cu(ii)-esculetin complex with a thiol peptide, glutathione (GSH), showed restoration of the fluorescence intensity as well as absorption maxima to that of pure esculetin. Incubation with other common thiol and non-sulphur amino acids did not show a similar restoration of the photophysical properties of the complex except in the case of cysteine. Mechanistically, it was evident that two molecules of GSH were consumed in reducing the Cu(ii)-esculetin complex, which subsequently split into the copper ion and esculetin. In this process GSH was converted into oxidised GSH (GSSG) as evident from the mass spectroscopy and HPLC studies. The above florescence regeneration behaviour of the copper-esculetin system in the presence of GSH was also observed in the cellular system using Chinese hamster ovary (CHO) as model cells. In conclusion, these studies may find application in developing sensors for detecting the cellular thiol level.

Sensitivity-Enhanced CMOS Phase Luminometry System Using Xerogel-Based Sensors.

We present the design and implementation of a phase luminometry sensor system with improved and tunable detection sensitivity achieved using a complementary metal-oxide semiconductor (CMOS) integrated circuit. We use sol-gel derived xerogel thin films as an immobilization media to house oxygen (O2) responsive luminescent molecules. The sensor operates on the principal of phase luminometry wherein a sinusoidal modulation signal is used to excite the luminophores encapsulated in the porous xerogel films and the corresponding phase shift of the emission signals is monitored. The phase shift is directly related to excited state lifetimes of the luminophores which in turn are related to the concentration of the target analyte species present in the vicinity of the luminophores. The CMOS IC, which consists of a 16 times 16 high-gain phototransistor array, current-to-voltage converter, amplifier and tunable phase shift detector, consumes an average power of 14 mW with 5-V power supply operating at a 38-kHz modulation frequency. The output of the IC is a dc voltage that corresponds to the detected luminescence phase shift with respect to the excitation signal. As a prototype, we demonstrate an oxygen sensor system by encapsulating the luminophore tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(II) within the xerogel matrices. The sensor system showed a fast response on the order of few seconds and we obtained a detection sensitivity of 118 mV per 1% change in O2 concentration. The system demonstrates a novel concept to tune and improve the detection sensitivity for specific concentrations of the target analyte in many biomedical monitoring applications.

Chloride analysis in magnesium metal using ion chromatography with conductometric detection.

A simple, rapid and accurate method for the determination of chloride in magnesium metal has been developed. The quantitative determination of chloride was accomplished by anion exchange chromatography with conductometric determination. A Metrosep Anion Dual 2 analytical column connected in series with a Metrosep RP guard column was used for chloride separation. A solution containing a mixture of 1.3 mM Na2CO3 and 2 mM NaHCO3 was used as eluent. The method requires a sample dissolution using nitric acid. The limit of detection for the determination of chloride is 50 mg kg(-1) and the relative standard deviation was 5% for the overall method. The recovery of chloride added was 99-102%. No interference was observed from either the closely eluting "system peak" or the nitrate peak in the determination of chloride.

Study of microporosity of active carbon spheres using inverse gas chromatographic and static adsorption techniques.

Active carbon spheres (ACSs) with different porous structures prepared in the laboratory were characterized by static adsorption studies and inverse gas chromatographic (IGC) technique. Surface properties such as BET surface area, micropore volume and pore size in different regions of porosity were determined using different theoretical approaches. Thermodynamic parameters such as isosteric heat of adsorption, free energy of adsorption and dispersive component of the surface energy were determined using IGC technique from corrected retention volume of normal alkanes and corresponding branched alkanes. Thermodynamic parameters were used to assess the molecular sieving property of ACSs. It is observed that thermodynamic properties strongly depend on microporous character of ACSs. The variations observed in pore size determined by both of the techniques that is by static adsorption measurements and IGC may be attributed to the variation in analysis temperature, i.e. liquid N2 temperature for adsorption studies and elevated temperature for IGC technique.

An organically modified silicate-based ethanol biosensor.

A novel electrocatalytic ethanol biosensor using ferrocene-encapsulated palladium (Pd)-linked organically modified sol-gel glass (ormosil) is reported. The alkoxy precursors used to prepare the new ormosil-based electrocatalytic biosensor are Pd-linked glycidoxypropyltrimethoxysilane and trimethoxysilane. Pd-glycidoxypropyltrimethoxysilane (black solution) is made by mixing aqueous solutions of palladium chloride and glycidoxypropyltrimethoxysilane. The new ormosil is made using a Pd-linked silane precursor, trimethoxysilane, an aqueous solution of ferrocene monocarboxylic acid, and HCl. Alcohol dehydrogenase (ADH) is assembled over the ferrocene-ormosil layer using polyvinyl alcohol and then protecting the immobilized enzyme layer using Millipore filter membranes (pore size 1 microm). The electrocatalytic response of immobilized ADH, soluble nicotinamide adenine dinucleotide, and Pd-linked ormosil-encapsulated ferrocene is then observed. The electrocatalytic oxidation of NADH and the subsequent ADH-catalyzed formation of NADH are monitored electrochemically. Typical results recorded after the addition of varying concentrations of ethanol are reported; however, the sensor is sensitive to other alcohol and known ADH-sensitive substrates. The stability and reproducibility of the new ethanol biosensor are reported.

An algorithm and a fortran program (chemequil-2) for calculation of complex equilibria.

A computer program, CHEMEQUIL-2 (CHEMical EQUILibrium), based on interfacing an iterative algorithm with the Newton-Raphson method, for calculating equilibrium compositions in aqueous mixtures of metals and ligands, is described. The program is also capable of simulating acid-base titrations. It has been compared with MINIQUAD, COMPLEX and MINEQL with respect to execution time and memory requirements. As a result of algorithm development and program design, CHEMEQUIL-2 offers considerable savings in both execution time (by 1-2 orders of magnitude) and memory requirements, especially for large problems, compared to these programs. The computational efficiency of CHEMEQUIL-2 makes it well suited for use in hydrogeochemieal transport models.

An algorithm for reducing storage requirements in computer calculation of chemical equilibria.

An algorithm for storage of stoichiometric coefficients of the possible complexes and solids in a multi-component system of metals and ligands is described, along with FORTRAN code for its implementation. The proposed algorithm results in considerable saving in storage over the conventional use of a two-dimensional array. The saving in storage is especially useful for microcomputers, and for very large problems such as those encountered in geochemical calculations.

Study of the effect of amines on structure of coacervates formed by photochemical effect on metal ions in presence of formaldehyde.

If an aqueous mixture of ammonium molybdate, diammonium hydrogen phosphate, biological minerals, and formaldehyde is exposed to sunlight, typical "Jeewanu", mixtures are observed. However, if a little solution of p-phenylene diamine, p-nitroaniline, alpha-naphthylamine or beta-naphthyl amine is added to it, a dense white precipitate is obtained. This precipitate, under a microscope at X1200 magnification, appears as large clusters of very tiny cream-coloured spherules. But if the mixture is first exposed to sunlight for four hours, when the formation of typical microstructures appears, and then the amine solution is added, the formation of the precipitate is retarded as the exposure time of amine-free mixture is enhanced, till finally the formation of spherules as precipitate is stopped. Microscopic study in twenty days of exposure of the amine-containing mixture tube shows that the number of typical microstructures decreases; they get larger in size and gradually disintegrate, while the formation of tiny sdpherules is accelerated.

Detergent effect on metabolic changes in microorganisms. A review.

Detergents contain a hydrophobic hydrocarbon structure and a hydrophilic group which may be anionic, cationic, or neutral. Detergents form stable emulsions and are capable of trapping lipid-soluble materials in the interior of the hydrophobic portion of the miscelles. Application of the knowledge of detergents to the discipline of microbiology would provide interesting and accurate data for further studies.

Organic acids produced by self-sustaining coacervates in presence of p-nitroaniline and p-phenylene-diamine.

Oleic and malonic acids were found in control samples while p-nitroaniline addition induced production of tricarballylic acid in eight and twelve days exposed samples. Fumaric and citric acids were found in sporadic instances. Addition of p-phenylene diamine to the mixture showed a degeneration process in organic acid production. In some cases production of fumaric acid was positive.

Amino acid changes in abiogenic mixtures containing p-nitroaniline and p-phenylene diamine.

Mixtures containing diammonium hydrogen phosphate, ammonium molybdate, and minerals in presence of p-nitroaniline resulted in formation of glycine, methionine, phenylalanine, tryptophan, and tyrosine. Proline and valine were observed in a few tubes. When p-phenylene diamine was present in the mixture, glycine was absent, while isoleucine and leucine were observed in the tubes. Predominance of proline was observed in all the studies executed.

Investigation of effect of alpha-naphthylamine and beta-naphthylamine on organic acids formation of self-sustaining coacervates.

Malonic and oleic acids were found in the typical mixtures. Samples containing alpha-naphthylamine gave oleic and malic acids which disintegrated with time of exposure. Tricarballylic acid was confirmed in presence of beta-naphthyl amine, but disappeared with time. Arachidic, tartaric, and adipic acids were found, but they disappeared in eight days' exposed samples. At later stages of exposure, malonic acid was observed along with sporadic appearances of citric , succinic, and oleic acids. Rate of disintegration of these compounds was directly proportional to time of exposure to sunlight.

Investigation of amino acid changes in self-sustaining coacervates in presence of alpha-naphthyl amine and beta-naphthyl amine.

Pronounced variation of amino acids was exhibited. Phenylalanine and tryptophan were invariably present, except at very few exposure periods. Methionine was perceived in many studies in alpha-naphthylamine, while in the study of beta-naphthylamine it was observed at four days' exposure. alpha-Naphthylamine produced cysteine along with tyrosine, leucine, isoleucine, and norleucine, while beta-naphthylamine gave valine along with tyrosine, leucine, isoleucine, and norleucine. Complete variation from blank signifies the effect of alpha- and beta-naphthyl amine on the mixture.