A Far-Red Emitting Fluorescent Chemogenetic Reporter for In Vivo Molecular Imaging

Far-red emitting fluorescent labels are highly desirable for spectral multiplexing and deep tissue imaging. Here, we describe the generation of frFAST (far-red Fluorescence Activating and absorption Shifting Tag), a 14-kDa monomeric protein that forms a bright far-red fluorescent assembly with (4-hydroxy-3-methoxy-phenyl)allylidene rhodanine (HPAR-3OM). As HPAR-3OM is essentially non-fluorescent in solution and in cells, frFAST can be imaged with high contrast in presence of free HPAR-3OM, which allowed the rapid and efficient imaging of frFAST fusions in live cells, zebrafish embryo/larvae, and chicken embryos. Beyond enabling the genetic encoding of far-red fluorescence, frFAST allowed the design of a far-red chemogenetic reporter of protein–protein interactions, demonstrating its great potential for the design of innovative far-red emitting biosensors.     

Quantitative Concentration Inequalities for Empirical Measures on Non-compact Spaces

We establish quantitative concentration estimates for the empirical measure of many independent variables, in transportation distances. As an application, we provide some error bounds for particle simulations in a model mean field problem. The tools include coupling arguments, as well as regularity and moment estimates for solutions of certain diffusive partial differential equations.     

Elution of organic solutes from different polarity sorbents using subcritical water

The intermolecular interactions between organic solutes and sorbent matrices under subcritical water conditions have been investigated at a pressure of 50 bar and temperatures ranging from 50 to 250°C. Both polar and nonpolar organics (chlorophenols, amines, n-alkanes, and polycyclic aromatic hydrocarbons) and five different sorbent matrices (glass beads, alumina, Florisil, silica-bonded C₁₈, and polymeric XAD-4 resins) were used. From the same matrix, the polar solutes always eluted at lower temperatures, while the moderately polar and nonpolar solutes only eluted at higher temperatures. Similar to matrix effects previously observed using supercritical carbon dioxide, the sorbent type greatly influenced the elution efficiency under subcritical water conditions. Lower temperatures are sufficient to elute a particular solute from glass beads, alumina, and Florisil, but higher temperatures (less polar water) are needed to elute the same solute from silica-bonded C₁₈. The highest temperatures were required to elute aromatic organics from XAD-4. These matrix effects demonstrate that, while low temperature water can break inert or dipole interactions between solutes and glass beads, alumina, and Florisil, higher temperature water is required to interrupt the van der Waals attractions between solutes and silica-bonded C₁₈, and even higher temperatures needed to overcome the π-electron interactions between aromatic solutes and XAD-4.     

Possible Role of the Iron Coordination Sphere in Hemoprotein Electron Transfer Self-Exchange: (1)H NMR Study of the Cytochrome c-PMe(3) Complex

The rates of self-exchange electron transfer in the trimethylphosphine complex of cytochrome c have been measured by an NMR technique over a large range of ionic strengths. The rate constant is 1.56 x 10(4) M(-)(1) s(-)(1) at 23 degrees C (&mgr; = 0.34 M) at pH 6.9. Dependence on ionic strength of the rate constant is treated by van Leeuwen theory. Extrapolation of the rate constant to infinite ionic strength gives a rate constant of 3.9 x 10(5) M(-)(1) s(-)(1). This rate constant is compared with others reported for myoglobin and cytochrome b(5)(). The values for these systems range over 2 orders of magnitude with myoglobin-PMe(3) < cytochrome b(5)() < cytochrome c-PMe(3) < cytochrome c. Analysis of the data in terms of Marcus theory gives a reorganization energy, lambda, for self-exchange of 0.75 eV mol(-)(1) for cytochrome c-PMe(3). Substitution of Met-80 by PMe(3) appears to influence only weakly the rearrangement barrier to electron transfer.     

Determination of ambroxol in an automated multi-pumping pulsed flow system.

A new flow methodology exploiting the multi-pumping approach was developed for the spectrophotometric determination of ambroxol hydrochloride in pharmaceutical preparations. The flow manifold was implemented by using, exclusively, multiple solenoid-actuated micro-pumps, which acted simultaneously as sample insertion, solutions propelling and reagents commutation units. Linear calibration plots were obtained over an ambroxol concentration ranging from 10 to 200 mg l(-1) (r.s.d. < 0.5%, n = 15) and a sampling rate of about 60 samples per hour (flow rate = 1.92 ml min(-1), sample volume = 80 microl).     

Le cation méthylènecyclopropyle: Etude théorique. Solvolyse des halogénocyclopropènes et des halogénométhylenecyclopropanes

The solvolysis in acid solution of allylic halides and acetates of methylenecyclopropanes and cyclopropylmethanes does not lead to ring opening when there is an ethoxycarbonyl substituent on the ring methylene. Only the cyclopropenyl derivative is obtained. A study of the electronic structure and the geometry of the possible allylic cation intermediate has been carried out using the MNDO method. In addition, the influence of the substituents on the ring opening of this cation has beeen examined. It appears that the ring opening is disfavoured upon substitution of an alkyl group by alkoxymethyl on the ring methylene group.     

Synthesis and complexing properties of a double-calix[4]arene crown ether

The template synthesis of copper(II) phthalocyanine by the condensation of dicyanobenzene within the supercages of Cu(II) exchanged X and Y type zeolites was studied using differential scanning calorimetry. The exotherms for intrazeolite and surface complexation have been resolved. These thermal events exhibit a dependence on the copper loading and hydration level which has been interpreted as a pressure effect.     

Development of cross-linked polystyrene-supported chiral amines featuring a fluorinated linker for gel-phase 19F NMR spectrometry monitoring of reactions

Ten cross-linked polystyrene-supported, protected chiral amines featuring both a spacer, comprising from 5 to 15 atoms, and a fluorinated linker have been successfully prepared. The development of the monitoring technique by gel-phase 19F NMR spectrometry on cross-linked polystyrene derivatives proved to be of high value in four steps of the process, as shown by the comparison of data gathered from both a classic NMR spectrometer and elemental analysis. Gel-phase 19F NMR spectrometry, thus, constitutes a useful technique that complements IR and 13C NMR spectrometries for the qualitative monitoring of reactions. In addition, quantitative determination of the conversion in a given transformation is possible, provided that 19F chemical shifts of the substrate and the product be different enough (Deltadelta>base width), as illustrated by the Mitsunobu coupling process (16-->17). The technique is nondestructive, and the samples used to monitor the reactions may be returned to the reaction medium. Deprotection of the above amines was achieved and furnished eight of the final resins in good to acceptable purity for future applications.