Simple equilibrium dialysis-high-performance liquid chromatographic method for the in vitro assessment of 5-methoxypsoralen bound to human albumin

Increasingly used in therapeutics, 5-methoxypsoralen (5-MOP), a linear furocoumarin, associated with UVA irradiation (PUVA), is now an established treatment for skin diseases such as vitiligo, mycosis funcoides and particularly psoriasis. Successful PUVA therapy depends on a sufficiently high peak 5-MOP plasma concentration coinciding with the UVA irradiation. However, as with most drugs, only the free plasma fraction is able to enter the target cells and has a pharmacological effect. In this work, the binding of 5-MOP to human albumin was studied in vitro, using a dialysis chamber. Bound and free 5-MOP fractions were quantified by a modification of Stolk's high-performance liquid chromatographic method. Dialysis was performed at 37 degrees C and pH 7.4 for 2 h, against a 4% albumin solution in phosphate buffer. The 5-MOP concentrations used were from 5 x 10(-5) to 5 x 10(-2) g/l in 1 x 10(-1) g/l steps. The 5-MOP bound strongly to human albumin in an unsaturable way. The mean 5-MOP binding to albumin was 95.3%. These results are in accordance with those published by Artuc et al. and not with those of Veronese et al., who found a lower saturable fixation (91%). These two research groups used tritiated 5-MOP. The technique used in this work is simple and inexpensive. It can be employed easily in vivo, e.g., for the assessment of 5-MOP free fractions in different therapeutic conditions.     

Cell immersion and cell dipping in microfluidic devices

Combining deflective dielectrophoretic barriers with controlled pressure driven liquid flows in microfluidic devices allows accurate handling of particles such as biological cells in suspensions. Working towards cell-based lab-on-a-chip applications, a platform permitting rapid testing of devices having different dielectrophoretic and fluidic subunits was developed. The performance of such a system is shown in the cases of (A) flooding a small number of immobilised cells with a dye and (B) transient buffer swapping of a large number of cells in flow. The transition times for moving cells from one reagent to the other are below 0.5 s in the case of flow-through cell dipping.     

FURTHER CHARACTERIZATION OF ANESTHETIC-TREATED PURPLE MEMBRANES

Abstract— The absorption maximum of bacteriorhodopsin is shifted from 568 nm to 480 nm when halogenated volatile anesthetics (enflurane; halothane) are added to purple membranes. Analysis of the rate of formation of this new species upon addition of the anesthetic and of the back-formation of native bacteriorhodopsin upon its removal indicate that in purple membranes, the dark-adapted chromophore is much less reactive than its light-adapted counterpart. Lipid-soluble molecules thus have a lower accessibility to the dark-adapted chromophore.
In addition, activity of the 480 nm bacteriorhodopsin was investigated. Flash and steady-state photolysis experiments reveal that this blue shifted chromophore has full photochemical activity. It has a meta-intermediate absorbing maximally at 380 nm. The photocycle ofBR–480 is mainly characterized by a slow decay of the "O" intermediate, enabling the direct observation of the branching reaction between the "M" intermediate and the parentBR–480 pigment.     

Gentle cell trapping and release on a microfluidic chip by in situ alginate hydrogel formation

Microfluidic devices are increasingly used to perform biological experiments on a single-cell basis. However, long-term stability of cell positions is still an issue. A novel biocompatible method for cell entrapment and release on a microchip is presented. It is based on the controlled formation of an alginate hydrogel by bringing two laminar flows of alginate and calcium ions in the range of 2 mM to 40 mM into contact. The resulting growth of a gel bar is used to enclose and immobilize yeast cells. Adding ethylenediaminetetraacetic acid (EDTA) to the alginate solution allows for control of the hydrogel growth, and by varying the ratio of Ca(2+) to EDTA concentrations gel growth or gel shrinkage can be induced at will. Trapped cells are released during shrinkage of the gel. The trapping efficiency for different cell speeds is investigated and the properties of gel growth are discussed using a diffusion model. Precise positioning of a single cell is demonstrated. The technique presented allows not only the reversible immobilization of cells under gentle conditions but also offers the potential of long-term cell cultures as shown by on-chip incubation of yeast cells. The procedure may provide a simple and fully biocompatible technique for a multitude of innovative experiments on cells in microsystems.     

Use of bis(trifluoromethyl)peroxy dicarbonate as initiator in the radical homopolymerisation of vinylidene fluoride (VDF) and copolymerisation of VDF with hexafluoropropylene

The radical homopolymerisation in acetonitrile of vinylidene fluoride (or 1,1-difluoroethylene, VDF) and the copolymerisation of VDF with hexafluoropropylene (HFP) initiated by bis(trifluoromethyl)peroxy dicarbonate are presented. Different reactions and different reactants were chosen to monitor the polymerisation in terms of initiating radicals generated from this initiator. Homopolymers and copolymers thus obtained were characterised by and NMR spectroscopy. From the assignments of the characteristic signals, an overall reaction mechanism is proposed that explains each step of the polymerisation. Particularly, an interpretation of the polymer microstructures and the presence of end-groups arising from the radical initiator as well as from eventual transfers is suggested. Among some of the microstructures, the trifluoromethoxy end-group was noted to be present in both PVDF and poly(VDF-co-HFP) (co)polymers, as generated from the decomposition of the initiator. This trifluoromethoxy end-group enabled the assessment of the molecular weights of PVDF and poly(VDF-co-HFP) (co)polymers. Thermal properties of the copolymers were also determined, showing that original fluoroelastomers possessing CF₃ end-groups are obtained endowed with low T_g and good thermal stability.     

One-pot rhodium(i)-catalyzed hydroboration of alkenes: radical conjugate addition

B-Alkylcatecholboranes, prepared by rhodium(I)-catalyzed hydroboration of alkenes, are suitable radical precursors for conjugate addition to activated olefins. This procedure proved to be particularly useful for the control of the regio- and chemoselectivity of such tandem processes. Enantioselective hydroboration has also been successfully coupled with radical chain reaction in a one-pot process.     

Regioselective allylic alkylation and etherification catalyzed by in situ generated N-heterocyclic carbene ruthenium complexes

Benzimidazolium halides are used for the first time as ligand precursors in ruthenium-catalyzed substitution of allylic carbonates and chlorides by carbon nucleophiles and phenols, respectively. After generation of diaminocarbene species upon deprotonation by tBuOK, their association with [Cp*Ru(MeCN)₃]PF₆ induces a very high regioselectivity in favor of the branched isomers when cinnamyl derivatives are used as starting substrates. They also provide good regioselectivities for the allylation of phenols by unsymmetrical aliphatic allylic substrates such as 3-chloro-4-phenylbut-1-ene, and thus provide a straightforward access to new allylic phenyl ethers.     

Zeolitic properties of mixed hexacyanoferrates/II/: Adsorption isotherms and differential heats of adsorption of water vapour

The series of mixed hexacyanoferrates M₂Zn₃[Fe/CN/₆]₂.xH₂O (M=K⁺, Na⁺, Cs⁺) present zeolitic properties. The adsorption water vapour isotherms were determined by TG at 303 K. The differential heats of adsorption of water vapour were measured by microcalorimetry; the molar integral entropy of the sorbed phase was calculated. The data obtained show that (i) the H₂O-sorbent interactions are very weak at the beginning of cavity filling, whereas the sorbed molecules exhibit the same mobility as in the vapour phase; (ii) during the filling, the H₂O-H₂O interactions increase up to a maximum; (iii) close to total filling, the adsorption phenomenon is comparable to a vapour-liquid transition and the sorbed phase has the same molar entropy as that of the liquid phase.     

Synthesis of new aromatic perfluorovinyl ether monomers containing phosphonic acid functionality

The synthesis of a new perfluorovinyl ether monomer containing phosphonic acid functionality is reported. It started from 4-[(α,β,β-trifluorovinyl)oxy]bromo benzene prepared in two steps from the nucleophilic substitution of 4-bromophenate to 1,2-dibromotetrafluoroethane. The [(α,β,β-trifluorovinyl)oxy]benzene dialkyl phosphonate was prepared according to various methods of phosphonation like a Michaelis-Arbuzov or a Michaelis-Becker or a palladium catalysed arylation in the presence of various reactants. The influence of the nature of the method and of the reactants onto the yields is discussed. It was shown that reaction involving a palladium triphenyl phosphine catalyst led to the best yield. All different aromatic intermediates and fluoromonomers were characterised by , and NMR, mass spectrometry (EI), and by FTIR.