Development of a new nonpeptidic self-immolative spacer. Application to the design of protease sensing fluorogenic probes

The design and synthesis of novel self-immolative spacer systems aiming at the release of phenol-containing compounds are described. The newly designed traceless linkers proved to be conveniently stable under physiological conditions and operate through spontaneous decomposition of an hemithioaminal intermediate under neutral aqueous conditions. Their utility was then illustrated by the preparation of original fluorogenic substrates of penicillin amidase whose strong fluorescence is unveiled through enzyme-initiated domino reactions.     

Thermostability of salt bridges versus hydrophobic interactions in proteins probed by statistical potentials

The temperature dependence of the interactions that stabilize protein structures is a long-standing issue, the elucidation of which would enable the prediction and the rational modification of the thermostability of a target protein. It is tackled here by deriving distance-dependent amino acid pair potentials from four datasets of proteins with increasing melting temperatures (Tm). The temperature dependence of the interactions is determined from the differences in the shape of the potentials derived from the four datasets. Note that, here, we use an unusual dataset definition, which is based on the Tm values, rather than on the living temperature of the host organisms. Our results show that the stabilizing weight of hydrophobic interactions (between Ile, Leu, and Val) remains constant as the temperature increases, compared to the other interactions. In contrast, the two minima of the Arg--Glu and Arg--Asp salt bridge potentials show a significant Tm dependence. These two minima correspond to two geometries: the fork--fork geometry, where the side chains point toward each other, and the fork--stick geometry, which involves the N(epsilon) side chain atom of Arg. These two types of salt bridges were determined to be significantly more stabilizing at high temperature. Moreover, a preference for more-compact salt bridges is noticeable in heat-resistant proteins, especially for the fork--fork geometry. The Tm-dependent potentials that have been defined here should be useful for predicting thermal stability changes upon mutation.     

Very weak, generalized and strong solutions to the Stokes system in the half-space

In this paper, we study the Stokes system in the half-space , with N?2. We give existence and uniqueness results in weighted Sobolev spaces. After the central case of the generalized solutions, we are interested in strong solutions and symmetrically in very weak solutions by means of a duality argument.     

Isolation of a new class of ecdysteroid conjugates (glucosyl-ferulates) using a combination of liquid chromatographic methods

The Polynesian medicinal fern Microsorum membranifolium contains very large amounts of ecdysteroids, including ecdysone, 20-hydroxyecdysone, 2-deoxy-20-hydroxyecdysone, and 2-deoxyecdysone. It also contains large amounts of unusual ecdysteroids which have been unambiguously identified by mass spectrometry and nuclear magnetic resonance. A new class of ecdysteroid conjugates (3-glucosyl-ferulates of 2-deoxyecdysone and 2-deoxy-20-hydroxyecdysone) is isolated, together with a new glycoside (2-deoxyecdysone 25-rhamnoside). The simultaneous presence of a sugar and an aromatic moiety results in a very particular chromatographic behavior of these conjugates. They behave like flavonoids and polyphenols when using the classical purification on polyamide, aimed at removing the latter from crude plant extracts, and would therefore be lost. They elute as non-polar ecdysteroids on reversed-phase high-performance liquid chromatography (RP-HPLC), whereas their behavior on normal-phase (NP) HPLC is strongly dependent on the mobile phase composition. Our data highlight the importance of selectivity in the choice of HPLC methods used for ecdysteroid separations.     

Optimization of solid-phase extraction for the liquid chromatography-mass spectrometry analysis of harpagoside, 8-para-coumaroyl harpagide, and harpagide in equine plasma and urine

Solid-phase extraction cartridges among those usually used for screening in horse doping analyses are tested to optimize the extraction of harpagoside (HS), harpagide (HG), and 8-para-coumaroyl harpagide (8PCHG) from plasma and urine. Extracts are analyzed by liquid chromatography coupled with multi-step tandem mass spectrometry. The extraction process retained for plasma applies BondElut PPL cartridges and provides extraction recoveries between 91% and 93%, with RSD values between 8 and 13% at 0.5 ng/mL. Two different procedures are needed to extract analytes from urine. HS and 8PCHG are extracted using AbsElut Nexus cartridges, with recoveries of 85% and 77%, respectively (RSD between 7% and 19%). The extraction of HG involves the use of two cartridges: BondElut PPL and BondElut C18 HF, with recovery of 75% and RSD between 14% and 19%. The applicability of the extraction methods is determined on authentic equine plasma and urine samples after harpagophytum or harpagoside administration.     

Optimized palladium‐based approaches to analogues of PK 11195

The peripheral‐type benzodiazepine receptor ligands such as PK 11195 and Ro 5‐4864 were found more than twenty years ago in the course of research on neurobiology. These ligands were instrumental in pointing out an involvement of the peripheral‐type benzodiazepine receptor (PBR) in apoptosis processes. With in mind an improvement of the solubility of PK 11195 in biological media, we report here improved reaction conditions for the palladium‐based arylation reaction of alkyl 1‐bromoisoquinoline‐3‐carboxylates and its ethyl 4‐bromoquinoline‐2‐carboxylate isomer. The use of [1,1′‐bis(diphenylphosphino) ferrocene] dichloropalladium as a precatalyst enabled a much improved preparation of an array of the 1‐arylisoquinoline‐3‐carboxylates as well as 4‐arylquinoline‐3‐carboxylates. This work should pave the way for the design of chemical probes aiming at the elucidation of the PBR biological role(s).     

Novel diastereoselective synthesis of bicyclic beta-lactams through radical cyclization and their reduction toward 2-(1-alkoxy-2-hydroxyethyl)piperidines and 2-(1-alkoxy-2-hydroxyethyl)azepanes

1-Allyl- and 1-(3-phenylallyl)-substituted 4-(2-bromo-1,1-dimethylethyl)azetidin-2-ones were transformed into 3-substituted 7-alkoxy-5,5-dimethyl-1-azabicyclo[4.2.0]octane-8-ones through radical cyclization by means of n-tributyltin hydride and AIBN in toluene with excellent diastereocontrol (>or=99%). The radical cyclization of 4-(2-bromo-1,1-dimethylethyl)-1-(2-methylallyl)azetidin-2-ones afforded 8-alkoxy-3,6,6-trimethyl-1-azabicyclo[5.2.0]nonan-9-ones in good diastereomeric excess (75-78%). The reductive ring opening of 1-azabicyclo[4.2.0]octane-8-ones and 1-azabicyclo[5.2.0]nonan-9-ones with lithium aluminum hydride resulted in novel 2-(1-alkoxy-2-hydroxyethyl)piperidines and -azepanes, which were isolated as single isomers.     

CEC for Studying the Retention and Separation of Pesticides on a Humic Acid Stationary Phase

A humic acid stationary phase is evaluated for studying by capillary electrochromatography (CEC) the binding of four pesticides with humic acid (HA), the main organic component in soil. It was demonstrated that the humic acid capillary was stable in terms of efficiency and retention during a long period of time. It appeared that HA has a lower affinity for neutral than for charged pesticides. The influence of the mobile phase constitution on the pesticide retention was also investigated in order to provide valuable information about the binding mechanism. The results suggest that the interactions of the pesticides during CEC are solely based on chromatographic mechanisms and that electrophoresis only plays a minor role. As well, it was shown that the HA stationary phase was able to separate these pesticides by CEC with a low analysis time. This HA capillary column could soon become attractive to determining the risk assessment of pesticides.     

Inducing spin crossover in metallo-supramolecular polyelectrolytes through an amphiphilic phase transition

A phase transition in an amphiphilic mesophase is explored to deliberately induce mechanical strain in an assembly of tightly coupled metal ion coordination centers. Melting of the alkyl chains in the amphiphilic mesophase causes distortion of the coordination geometry around the central transition metal ion. As a result, the crystal field splitting of the d-orbital subsets decreases resulting in a spin transition from a low-spin to a high-spin state. The diamagnetic-paramagnetic transition is reversible. This concept is demonstrated in a metallo-supramolecular coordination polyelectrolyte-amphiphile complex self-assembled from ditopic bis-terpyridines, Fe(II) as central transition metal, and dialkyl phosphates as amphiphiles. The magnetic properties are studied in a Langmuir-Blodgett multilayer. The modularity of this concept provides extensive control of structure and function from molecular to macroscopic length scales and gives access to a wide range of new molecular magnetic architectures such as nanostructures, thin films, and liquid crystals.