Glutamate receptor incorporated in a mixed hybrid bilayer lipid membrane array, as a sensing element of a biosensor working under flowing conditions

The realization of a reliable receptor biosensor requires stable, long-lasting, reconstituted biomembranes able to supply a suitable biomimetic environment where the receptor can properly work after incorporation. To this end, we developed a new method for preparing stable biological membranes that couple the biomimetic properties of BLMs (bilayer lipid membranes) with the high stability of HBMs (hybrid bilayer membranes); this gives rise to an innovative assembly, named MHBLM (mixed hybrid bilayer lipid membrane). The present work deals with the characterization of biosensors achieved by embedding an ionotropic glutamate receptor (GluR) on MHBLM. Thanks to signal (transmembrane current) amplification, which is typical of natural receptors, the biosensor here produced detects glutamate at a level of nmol L(-1). The transmembrane current changes linearly vs glutamate up to 100 nmol L(-1), while the limit of detection is 1 nmol L(-1). In addition, the biosensor response can be modulated both by receptor agonists (glycine) and antagonists (Mg(2+)) as well, and by exploiting the biosensor response, the distribution of different kinds of ionotropic GluR present in the purified sample, and embedded in MHBLM, was also evaluated. Finally, one of the most important aspects of this investigation is represented by the high stability of the biomimetic system, which allows the use of biosensor under flowing conditions, where the solutions flow on both biomembrane faces.     

Surface enhanced Raman scattering investigation of the halide anion effect on the adsorption of 1,2,3-triazole on silver and gold colloidal nanoparticles

The halide anion effect on the adsorption of 1,2,3-triazole on Ag and Au colloidal nanoparticles has been investigated by means of surface enhanced Raman scattering (SERS), UV-visible absorption spectroscopy, and scanning electron microscopy. To interpret the SERS spectra, the vibrational spectra of 1,2,3-triazole were assigned with the help of density functional theoretical (DFT) calculations of the two tautomers of 1,2,3-triazole, both free and bound to Ag and Au adatoms. Upon addition of halide anions, both tautomers interact with the Ag surface through one nitrogen atom. Analogous behavior is observed in the case of basified Au colloids, whereas at the usual pH of these hydrosols (approximately 6) the adsorption of 1,2,3-triazole is the same of that observed in halide-free colloids.     

Removal of humic matter interference in the determination of Cr(VI) in soil extracts by the diphenylcarbazide method

Alkaline digestion of soil samples, which is recommended for minimizing Cr(III)–Cr(VI) interconversions during the extraction of Cr(VI), may also solubilize humic matter (HM). The latter is responsible for both positive and negative interference in the analysis of Cr(VI) in the extract by the diphenylcarbazide (DPC) method. Humic compounds indeed absorb light at 540 nm as the Cr-DPC product and are also able to rapidly reduce Cr(VI) under the pH conditions of the standard DPC method. To prevent any risk of interference and make the DPC method applicable to soil extracts, a new protocol is proposed. This consists of three successive steps: (1) extraction of Cr(VI) by the U.S.EPA method 3060A in the presence of Mg²⁺; (2) batch removal of solubilised HM by the XAD-7 sorbent at pH 3.0 ± 0.1 with 3–5 min contact time; (3) analysis of Cr(VI) with the DPC method at pH 3.0 ± 0.1. The application of this new protocol to the soil product SQC-012 Lot 4 certified by R.T. Corporation (RTC, USA) gave significantly lower Cr(VI) concentrations and smaller variability compared to certified values (46.5 ± 2.3 instead of 153 ± 32.6 mg/kg). The new protocol was validated by Cr(VI) and Cr(III) spikes either to the soil/extractant suspension or to the exctract. Cr(VI) results in the wide range of acceptance limits (104–202 mg/kg) reported by RTC were only obtained when the U.S.EPA method 3060A without the addition of Mg²⁺ and DPC analysis at pH 1.0 were applied. The latter procedure appears questionable since it leads to largely variable results which reflect the complex role played by humic matter in the determination of Cr(VI) and the instability of Cr(III) during the extraction.     

FLUORESCENCE OF Euglena gracilis PHOTORECEPTOR PIGMENT: AN in vivo MICROSPECTROFLUOROMETRIC STUDY

Abstract— The spectroscopic characterization of the photoreceptor pigment is one of the main questions in the study of the photosensory transduction chains in photomotile microorganisms. One of the possible techniques that can be used is in vivo microspectrofluorometry. By means of a tunable dye-laser microspectrofiuorometer developed by us, we have investigated some of the spectroscopic properties of the photoreceptor pigment of the green flagellate Euglena gracilis. The in vivo fluorescence excitation spectrum has been determined and the fluorescence quantum yield has been measured. The results show that flavins are indeed present in the paraflagellar body of E. gracilis and that their fluorescence quantum yield is much lower than that of a free flavin. An estimate of the order of magnitude of the rate constants for primary molecular reactions is tentatively given.     

Diradicals Photogeneration from Chloroaryl-Substituted Carboxylic Acids

With the aim of generating new, thermally inaccessible diradicals, potentially able to induce a double-strand DNA cleavage, the photochemistry of a set of chloroaryl-substituted carboxylic acids in polar media was investigated. The photoheterolytic cleavage of the Ar−Cl bond occurred in each case to form the corresponding triplet phenyl cations. Under basic conditions, the photorelease of the chloride anion was accompanied by an intramolecular electron-transfer from the carboxylate group to the aromatic radical cationic site to give a diradical species. This latter intermediate could then undergo CO₂ loss in a structure-dependent fashion, according to the stability of the resulting diradical, or abstract a hydrogen atom from the medium. In aqueous environment at physiological pH (pH=7.3), both a phenyl cation and a diradical chemistry was observed. The mechanistic scenario and the role of the various intermediates (aryl cations and diradicals) involved in the process was supported by computational analysis.     

PMMA/PC Blends: Effect of Mixing Conditions on Compatibility

In this work the effect of melt mixing condition and of a trans-esterification catalyst on miscibility of poly(methyl methacrylate) (PMMA)/polycarbonate of bisphenol A (PC) blends is studied. In particular, at high temperature chemical reactions between PMMA and PC phases can take place; these strongly change the compatibility in the blend and materials having single T_g can be obtained. FT-IR analyses, coupled with solvent extraction, suggest that a grafting reaction of PC on PMMA is involved. SEC and DSC data are consistent with spectroscopic results, and some decrement of the molar weight distribution (MWD) of PC phase is observed. On the other hand, the presence of a fraction of modified material having higher MWD of starting PMMA is also noticed. The single T_g characteristic of some materials has been confirmed by experimental data of structural relaxation performed by differential scanning calorimetry (DSC). These materials showed optical clarity and the morphological analysis performed by scanning electron microscopy (SEM) confirm the homogeneity of these materials.     

Interaction of lipophilic gemcitabine prodrugs with biomembrane models studied by Langmuir-Blodgett technique

The stability and bioavailability of anticancer agents, such as gemcitabine, can be increased by forming prodrugs. Gemcitabine is rapidly deaminated to the inactive metabolite (2('),2(')-difluorodeoxyuridine), thus to improve its stability a series of increasingly lipophilic gemcitabine prodrugs linked through the 4-amino group to valeroyl, lauroyl, and stearoyl acyl chains were synthesized. Studies of monolayer properties are important to improve understanding of biological phenomena involving lipid/gemcitabine or lipid/gemcitabine derivative interactions. The interfacial behavior of monolayers constituted by DMPC plus gemcitabine or lipophilic gemcitabine prodrugs at increasing molar fractions was studied at the air/water interface at temperatures below (10 degrees C) and above (37 degrees C) the lipid phase transition. The effect of the hydrophobic chain length of gemcitabine derivatives on the isotherm of pure DMPC was investigated by surface tension measurement, and the results are reported as molar fractions as a function of mean molecular area per molecule. The results show that the compounds interact with DMPC producing mixed monolayers that are subject to an expansion effect, depending on the prodrug chain length. The results give useful hints of the interaction of these prodrugs with biological membranes and increase knowledge on the incorporation site of such compounds, as a function of their lipophilicity, in a lipid carrier; they may lead to improved liposomal formulation design.     

An electrochemically driven molecular shuttle controlled and monitored by C60

Herein we describe a fullerene rotaxane, in which shuttling between two well-defined and distant co-conformations is both induced and monitored by the C60 stopper.     

One-electron reduction of methanesulfonyl chloride. The fate of MeSO2Cl(.)(-) and MeSO2(.) intermediates in oxygenated solutions and their role in the cis-trans isomerization of mono-unsaturated fatty acids

The one-electron reduction of methanesulfonyl chloride (MeSO2Cl) leads, in the first instance, to an electron adduct MeSO2Cl(.)(-) which lives long enough for direct detection and decays into sulfonyl radicals MeSO2(.) and Cl(-), with k = 1.5 x 10(6) s(-1). Both, MeSO2Cl(.)(-) and MeSO2(.) showed a similar absorption in the UV with lambdamax of 320 nm. In the presence of oxygen, MeSO2Cl(.)(-) transfers an electron to O(2) and establishes an equilibrium with superoxide. The rate constant for the forward reaction was measured to 4.1 x 10(9) M(-1) s(-1), while for the back reaction only an interval of 1.7 x 10(5) to 1.7 x 10(6) M(-1) s(-1) could be estimated, with a somewhat higher degree of confidence for the lower value. This corresponds to an equilibrium constant in the range of 2.4 x 10(3) to 2.4 x 10(4). With reference to E degrees (O2/O2(.)(-)) = -155 mV, the redox potential of the sulfonyl chloride couple, E degrees (MeSO2Cl/MeSO2Cl(.)(-)), thus results between being equal to -355 and -414 mV (vs NHE). MeSO2Cl(.)(-) reduces (besides O2) 4-nitroacetophenone. The underlying electron transfer took place with k = 1.5 x 10(9) M(-1) s(-1), corroborating an E degrees for the sulfonyl chloride couple significantly exceeding the above listed lower value. The MeSO2(.) radical added to oxygen with a rate constant of 1.1 x 10(9) M(-1) s(-1). Re-dissociation of O2 from MeSO2OO(.) occurred only very slowly, if at all, that is, with k < 10(5) s(-1). MeSO2(.) radicals can act as the catalyst for the cis-trans isomerization of several Z- and E-mono-unsaturated fatty acid methyl esters in homogeneous solution. The effectiveness of the isomerization processes has been addressed, and in the presence of oxygen the isomerization is completely suppressed.