Reductive transformations of unsaturated azabicyclic nitrolactams

Using different reducing methods unsaturated indolizidine and quinolizidine lactams substituted with a nitro group were transformed into various alkaloid-like derivatives. Hydrogen transfer and palladium catalyzed hydrogenation gave compounds of ketolactam or lactam type meanwhile the nitro group was eliminated. On the other hand, in presence of Raney-nickel catalyst the nitro compounds were reduced to diastereomeric amino derivatives whose stereochemistry was elucidated by NMR spectroscopy. Using sodium bis-dimethoxy-ethoxy-aluminum-hydride (Red-Al) as reducing agent an unexpected tricyclic azetidine was isolated and characterized.     

Targeted identification of phosphorylated peptides by off-line HPLC-MALDI-MS/MS using LC retention time prediction

Protein phosphorylation is a type of posttranslational modification which plays an important role in cell regulation and signal transduction. Because of its biological relevance, a considerable amount of interest has been paid to the development of efficient techniques for phosphopeptide analysis. Although advances in MS control have enabled the high-throughput discovery of proteins from limited amounts of sample, automated selection of MS/MS precursor ions based on intensity alone can significantly hamper the detection of low-abundance phosphopeptides. On the basis of the observation that the introduction of a phosphate moiety does not dramatically change peptide retention time in reverse-phase chromatography, phosphopeptide specific MS/MS fragmentation attempts based on LC retention time and m/z were evaluated using a standard protein mixture, then using in vitro phosphorylated myelin basic protein. Results indicated that the majority (98%) of phosphopeptides identified eluted within a +/- 4-min window of the predicted LC elution time. While studies presented here are primarily proof of concept in nature, data suggest that the use of LC retention time prediction could be a valuable constraint for the identification of phosphopeptides within a set of off-line LC deposited sample spots. It is expected that the development of these methods will not only permit the targeted identification of protein phosphorylation sites but also allow the in-depth analysis of the dynamic events linked to the posttranslational modification.     

Dendrimers and combinatorial chemistry—tools for fluorescent enhancement in protease assays

FRET based systems are some of the best methods available to detect and monitor proteolytic activity. To enhance fluorescent signals and hence assay sensitivity, two different systems were developed using two different dendrimeric constructs. In the first case, a triple branched dendrimer bearing three dansyl groups was used to enhance assay sensitivity and showed a significant enhancement of fluorescence following enzymatic cleavage. In another example, a tris-fluorescein probe, that undergoes self-quenching, was utilized in a combinatorial library synthesis to map the substrate specificity of proteases.     

Determination of iron-porphyrin-like complexes at nanomolar levels in seawater

A new method for the non-specific determination of iron-porphyrin-like complexes in natural waters has been developed. It is based on the chemiluminescent oxidation of the luminol in the presence of dioxygen (O₂) at pH 13. The method has been implemented in a FIA manifold that allowed the direct injection of seawater. The limit of detection is 0.11 nM of equivalent hemin (Fe-protoporphyrin IX). Fe²⁺, Fe³⁺, H₂O₂, siderophore (deferoxamin mesylate), humic acid and phytic acid did not interfere when they were present at the concentrations expected in seawater. Metal free porphyrin and Mg, Cu, Co porphyrin complexes did not induce a significant chemiluminescent signal. Poisoned unfiltered samples could be stored for several weeks before analyses. The new method was successfully applied to the determination of the Fe-porphyrin complexes contained in cultured phytoplankton and in natural samples.     

Development of an analytical procedure for determination of selected estrogens and progestagens in water samples

An analytical procedure has been developed for determination of eight selected natural and synthetic hormonal steroids in surface water and in effluent samples. Several methodological points have been investigated and are discussed; they include the choice of the solid-phase extraction sorbent, the influence of flow rate on recovery, the breakthrough volume for a given sorbent (Env+ and Oasis HLB), sample clean up, and sample storage. As regards the latter point, it was found that when no preservative was added to effluent from a sewage-treatment plant, severe loss of steroids occurred—85% of progesterone and about 30% of both estrone and estradiol were found to be degraded in 24 h. The procedure developed was applied to samples from the Seine river estuary. Sex steroids were not detected in surface water; estrone was the most commonly detected steroid in sewage-treatment plant effluent, with levels ranging from 1.8 to 8.3 ng L^–1. Synthetic estrogens (ethynylestradiol and mestranol) and progestagens (levonorgestrel and norethindrone) were never detected, whatever the sampling season. Overall, for 162 out of 168 measurements levels were below the detection limits of the developed procedure.     

Computational Enzyme Design

Recent developments in computational chemistry and biology have come together in the “inside‐out” approach to enzyme engineering. Proteins have been designed to catalyze reactions not previously accelerated in nature. Some of these proteins fold and act as catalysts, but the success rate is still low. The achievements and limitations of the current technology are highlighted and contrasted to other protein engineering techniques. On its own, computational “inside‐out” design can lead to the production of catalytically active and selective proteins, but their kinetic performances fall short of natural enzymes. When combined with directed evolution, molecular dynamics simulations, and crowd‐sourced structure‐prediction approaches, however, computational designs can be significantly improved in terms of binding, turnover, and thermal stability.     

Biomimetic zinc funnel complexes based on calix[6]N3ArO ligands: an acid-base switch for guest binding

The coordination chemistry of Zn in an N(3)ArOH environment has been explored. The ligands are based on calix[6]arenes that present two imidazole arms and an amino phenol moiety at the narrow rim. Three different types of complexes have been characterized. One is dicationic with Zn(2+) coordinated to the three nitrogen atoms and to the oxygen of the phenol group of the calix[6]ligand. This complex is very sensitive to exogenous coordinating molecules and exists as a 5-coordinate species due to the endo-complexation of a guest. The second species is a monocationic complex for which the phenol group has been deprotonated. The resulting N(3)ArOZn complex can also bind a guest ligand albeit with a lower affinity than the dicationic complex. The third species is neutral. It can be obtained upon reaction with a base to yield a hydroxo complex or with an anion such as a chloride that coordinates the metal center from the outside of the calixarene cavity. The simultaneous binding of two anionic donors decreases the Zn Lewis acidity, allowing an impressive conformational reorganization of the system. One imidazole arm is released by the metal center. The other one undergoes self-inclusion into the pi-basic calixarene cavity because the low affinity of the metal center for neutral ligand does not allow the endo-coordination of an exogenous guest. Hence, the calix[6]N(3)ArOH-based Zn complexes act as an acid-base switch for guest binding. Several aspects of this system appear reminiscent of Zn-peptidases of the astacin and serralisin families.     

Co(II) and Cu(II) Schiff base complexes of bis(<Emphasis Type="Italic">N</Emphasis>-(4-diethylamino-2-methylphenyl)-3,5-di-<Emphasis Type="Italic">tert</Emphasis>-butylsalicylaldimine): Electrochemical and X-ray structural study

Copper(II) and cobalt(II) complexes of salicylaldimine obtained by the condensation of N,N-diethyl-2-methyl-1,4-phenylenediamine with 3,5-di-tert-butyl-2-hydroxybenzaldehyde have been synthesized and characterized by elemental analyses, magnetic susceptibility measurements, cyclic voltammetry, and FT-IR and UV–Vis spectroscopy. The molecular structure of the title copper(II) complex was determined by the single crystal X-ray diffraction technique. The Cu(II) center is coordinated by four atoms of the donor set in a compressed tetrahedral trans-[N₂O₂] environment, which can be essentially ascribed to the presence of bulky fragments of the ligand. The computed bond valences of the copper verify +2 oxidation state and indicate that the copper bonds, in particular Cu–N bonds, are elongated due to steric effects from bulky substituents in the ligands, N-(4-diethylamino-2-methylphenyl). Intermolecular C–H···π interactions leading to centrosymmetric synthons serve to stabilize periodic organization of the molecules.     

Compliance and Hill polarization tensor of a crack in an anisotropic matrix

This work aims at developing an efficient method to compute the compliance due to a crack modeled as a flat ellipsoid of any shape in an infinite elastic matrix of arbitrary anisotropy (Eshelby problem) when no closed-form solution seems currently available. Whereas the solution of this problem usually requires the calculation of the so-called fourth-order Hill polarization tensor if the ellipsoid is not singular, it is shown that the crack compliance can be derived from the first-order term in the Taylor expansion of the Hill tensor with respect to the smallest aspect ratio of the ellipsoidal inclusion. For a 3D ellipsoidal crack model, this first-order term is expressed as a simple integral thanks to the Cauchy residue theorem. A similar method allows to express the same term in the case of a cylindrical crack model without any integral. A numerical example is finally treated.