Rational design of lanthanide binding peptides

Lanthanide-binding peptides are very attractive for the design of bioprobes. Indeed, they combine the amazing properties of lanthanide ions, such as their time-resolved luminescence (Eu, Tb) or electronic relaxation (Gd) to the characteristics of the peptide scaffold, such as large solubility in water and ability to recognize biological substrates. Peptides derived from natural amino acids are reviewed in a first section. Some of their lanthanide complexes have already demonstrated their efficiency in determining protein structures and functions. Then, we will show how insertion of chelating unnatural amino acids modulates peptide-lanthanide complexes properties, such as luminescence and stability.     

On the Influence of Oxygen on the Degradation of Fe‐N‐C Catalysts

Fe‐N‐C catalysts containing atomic FeN_x sites are promising candidates as precious‐metal‐free catalysts for oxygen reduction reaction (ORR) in proton exchange membrane fuel cells. The durability of Fe‐N‐C catalysts in fuel cells has been extensively studied using accelerated stress tests (AST). Herein we reveal stronger degradation of the Fe‐N‐C structure and four‐times higher ORR activity loss when performing load cycling AST in O₂‐ vs. Ar‐saturated pH 1 electrolyte. Raman spectroscopy results show carbon corrosion after AST in O₂, even when cycling at low potentials, while no corrosion occurred after any load cycling AST in Ar. The load‐cycling AST in O₂ leads to loss of a significant fraction of FeN_x sites, as shown by energy dispersive X‐ray spectroscopy analyses, and to the formation of Fe oxides. The results support that the unexpected carbon corrosion occurring at such low potential in the presence of O₂ is due to reactive oxygen species produced between H₂O₂ and Fe sites via Fenton reactions.     

Molecularly Imprinted Polymers: Compromise between Flexibility and Rigidity for Improving Capture of Template Analogues

New synthetic strategies for molecularly imprinted polymers (MIPs) were developed to mimic the flexibility and mobility exhibited by receptor/enzyme binding pockets. The MIPs were prepared by bulk polymerization with quercetin as template molecule, acrylamide as functional monomer, ethylene glycol dimethacrylate as cross‐linker, and THF as porogen. The innovative grafting of specific oligoethylene glycol units onto the imprinted cavities allowed MIPs to be obtained that exhibit extended selectivity towards template analogues. This synthetic strategy gives promising perspectives for the design of molecular recognition of molecules based on a congruent pharmacophore, which should be of interest for drug development.     

Detection of oligonucleotide gas-phase conformers: H/D exchange and ion mobility as complementary techniques

Gas-phase hydrogen/deuterium exchange of small oligonucleotides (dTG, dC(6) and C(6)) with CD(3)OD was performed in the second hexapole of a Fourier transform ion-cyclotron resonance (FTICR) mass spectrometer. Ion activation experiments were conducted by accelerating the ions at the entrance of the H/D exchange cell under conditions promoting exclusively collisional isomerization. These experiments allowed us to assess the presence of several conformers, and to probe the height of the isomerization barrier separating these conformers. Ion mobility experiments were also performed. Their results were consistent with the H/D exchange data. A model accounting for the competing isomerization and H/D exchange reactions is proposed. Comparing the ion acceleration experiments for H/D exchange and for ion mobility reveals that the most compact conformer displays the fastest H/D exchange. This observation shows that H/D exchange and ion mobility provide us with complementary information because hydrogen accessibility and macromolecule compactness are not univocally associated.     

Optimization of a nano‐enzymatic reactor for on‐line tryptic digestion of polypeptide conjugates by capillary electrophoresis

This work aims at studying the optimization of an on‐line capillary electrophoresis (CE)‐based tryptic digestion methodology for the analysis of therapeutic polypeptides (PP). With this methodology, a mixture of surrogate peptide fragments and amino acid were produced on‐line by trypsin cleavage (enzymatic digestion) and subsequently analyzed using the same capillary. The resulting automation of all steps such as injection, mixing, incubation, separation and detection minimizes the possible errors and saves experimental time. In this paper, we first study the differents parameters influencing PP cleavage inside the capillary (plug length, reactant concentration, incubation time, diffusion and electrophoretic plugs mixing). In a second part, the optimization of the electrophoretic separation conditions of generated hydrolysis products (nature, pH and ionic strength (I) of the background electrolyte (BGE)) is described. Using the optimized conditions, excellent repeatability was obtained in terms of separation (migration times) and proteolysis (number of products from enzymatic hydrolysis and corresponding amounts) demonstrating the robustness of the proposed methodology.     

Synthesis and supramolecular organization of amphiphilic diblock copolymers combining poly(N,N-dimethylamino-2-ethyl methacrylate) and poly(epsilon-caprolactone)

Well-defined poly(epsilon-caprolactone) (PCL)/poly(N,N-dimethylamino-2-ethyl methacrylate (PDMAEMA) diblock copolymers were synthesized, and their self-assembly was investigated as micelles both in aqueous solutions and in thin solid deposits. The synthetic approach combines controlled ring opening polymerization (ROP) of epsilon-caprolactone (CL) and atom transfer radical polymerization (ATRP) of N,N-dimethylamino-2-ethyl methacrylate (DMAEMA). Diblock copolymers were prepared by ROP of CL initiated by (Al(OiPr)3), followed by quantitative reaction of the PCL hydroxy end-groups with bromoisobutyryl bromide. The alpha-isopropyloxy omega-2-bromoisobutyrate poly(epsilon-caprolactone) (PCL-Br) obtained was used as a macroinitiator for the ATRP of DMAEMA. The molecular characterization of those diblock copolymers was performed by 1H NMR spectroscopy and gel permeation chromatography (GPC) analysis. The self-assembly of the copolymers into micellar aggregates in aqueous media was followed with dynamic light scattering (DLS), as a function of concentration and the pH. In parallel, the morphology of the solid deposits of those micelles was examined with atomic force microscopy (AFM).     

Understanding J‐Modulation during Spatial Encoding for Sensitivity‐Optimized Ultrafast NMR Spectroscopy

Ultrafast (UF) NMR spectroscopy is an approach that yields 2D spectra in a single scan. This methodology has become a powerful analytical tool that is used in a large array of applications. However, UF NMR spectroscopy still suffers from an intrinsic low sensitivity, and from the need to compromise between sensitivity, spectral width, and resolution. In particular, the modulation of signal intensities by the spin–spin J‐coupling interaction (J‐modulation) impacts significantly on the intensities of the spectral peaks. This effect can lead to large sensitivity losses and even to missing spectral peaks, depending on the nature of the spin system. Herein, a general simulation package (Spinach) is used to describe J‐modulation effects in UF experiments. The results from simulations match with experimental data and the results of product operator calculations. Several methods are proposed to optimize the sensitivity in UF COSY spectra. The potential and drawbacks of the different strategies are also discussed. These approaches provide a way to adjust the sensitivity of UF experiments for a large range of applications.     

Intrinsic ratios of glucose, fructose, glycerol and ethanol 13C/12C isotopic ratio determined by HPLC-co-IRMS: toward determining constants for wine authentication

High-performance liquid chromatography linked to isotope ratio mass spectrometry (HPLC-co-IRMS) via a Liquiface? interface has been used to simultaneously determine ¹³C isotope ratios of glucose (G), fructose (F), glycerol (Gly) and ethanol (Eth) in sweet and semi-sweet wines. The data has been used the study of wine authenticity. For this purpose, 20 authentic wines from various French production areas and various vintages have been analyzed after dilution in pure water from 20 to 200 times according to sugar content. If the ¹³C isotope ratios vary according to the production area and the vintage, it appears that internal ratios of ¹³C isotope ratios ( R_¹³\textC {R_{{{}^{{13}}{\text{C}}}}} ) of the four compounds studied can be considered as a constant. Thus, ratios of isotope ratios are found to be 1.00 ± 0.04 and 1.02 ± 0.08 for R_{¹³\textC\textG/F} {R_{{{}^{{13}}{{\text{C}}_{{{\text{G/F}}}}}}}} and R_{¹³\textC\textGly/Eth} {R_{{{}^{{13}}{{\text{C}}_{{{\text{Gly/Eth}}}}}}}} , respectively. Moreover, R_{¹³\textC\textEth/Sugar} {R_{{{}^{{13}}{{\text{C}}_{{{\text{Eth/Sugar}}}}}}}} is found to be 1.15 ± 0.10 and 1.16 ± 0.08 for R_{¹³\textC\textGly/Sugar} {R_{{{}^{{13}}{{\text{C}}_{{{\text{Gly/Sugar}}}}}}}} . Additions of glucose, fructose and glycerol to a reference wine show a variation of the R_¹³\textC {R_{{{}^{{13}}{\text{C}}}}} value for a single product addition as low as 2.5 g/L⁻¹. Eighteen commercial wines and 17 concentrated musts have been analyzed. Three wine samples are suspicious as the R_¹³\textC {R_{{{}^{{13}}{\text{C}}}}} values are out of range indicating a sweetening treatment. Moreover, concentrated must analysis shows that ¹³C isotope ratio can be also used directly to determine the authenticity of the matrix.     

Comprehensive study of condensed tannins by ESI mass spectrometry: average degree of polymerisation and polymer distribution determination from mass spectra

The determination of the molecular mass distribution of tannins is still a challenge. To elucidate it, mass spectrometry is potentially interesting, but many previous studies have highlighted that the mass spectra of a tannin fraction do not always reflect the actual abundance of different chain lengths. To clarify the potentialities offered by the MS approach, a comprehensive study involving different tannin fractions analysed under different conditions was conducted with an electrospray ionization (ESI) source. This study allowed optimised ESI-MS conditions to be established for analysing tannins but also it outlines the limits of detection encountered. If the detection of high molecular weight tannins seems difficult or even impossible, the spectral distortions brought about by this limitation are not totally related to the sole average degree of polymerisation of the tannin fraction studied but greatly depend on its polymer distribution. However, ESI-MS used under optimised conditions is a suitable method to study tannin composition of vegetable extracts which contain degree of polymerisations below 26.