ADLOC: An Aptamer‐Displacement Assay Based on Luminescent Oxygen Channeling

Functional nucleic acids, such as aptamers and allosteric ribozymes, can sense their ligands specifically, thereby undergoing structural alterations that can be converted into a detectable signal. The direct coupling of molecular recognition to signal generation enables the production of versatile reporters that can be applied as molecular probes for various purposes, including high‐throughput screening. Here we describe an unprecedented type of a nucleic acid‐based sensor system and show that it is amenable to high‐throughput screening (HTS) applications. The approach detects the displacement of an aptamer from its bound protein partner by means of luminescent oxygen channeling. In a proof‐of‐principle study we demonstrate that the format is feasible for efficient identification of small drug‐like molecules that bind to a protein target, in this case to the Sec7 domain of cytohesin. We extended the approach to a new cytohesin‐specific single chain DNA aptamer, C10.41, which exhibits a similar binding behavior to cytohesins but has the advantage of being more stable and easier to synthesize and to modify than the RNA‐aptamer M69. The results obtained with both aptamers indicate the general suitability of the aptamer‐displacement assay based on luminescent oxygen channelling (ADLOC) for HTS. We also analyzed the potential for false positive hits and identified from a library of 18 000 drug‐like small molecules two compounds as strong singlet‐oxygen quenchers. With full automation and the use of commercially available plate readers, we estimate that the ADLOC‐based assay described here could be used to screen at least 100 000 compounds per day.     

Charge transport in columnar mesophases of carbazole macrocycles

Charge transport properties of a columnar mesophase of carbazole macrocycles are analyzed. Realistic morphologies are sampled using all-atom molecular dynamics simulations while charge transport is simulated using the kinetic Monte Carlo method with transfer rates obtained from the high temperature nonadiabatic limit of Marcus theory. It is shown that the molecular design with side chains pointing inside the macrocycle allows close approach between molecules of neighboring columns, thus enabling three-dimensional transport and helping to circumvent charge trapping on structural defects.     

An organocatalytic approach to 2-hydroxyalkyl- and 2-aminoalkyl furanes

The first enantioselective methodology for the synthesis of electron-poor 2-hydroxyalkyl- and 2-aminoalkyl furanes is demonstrated in this study. It utilizes a highly stereoselective organocatalytic one-pot reaction cascade: epoxidation or aziridination of α,β-unsaturated aldehydes followed by Feist-Be?nary reaction of various 1,3-dicarbonyl compounds to give the target furanes. This efficient multibond forming reaction cascade benefits from low catalyst loadings and readily available starting materials. Furthermore, the possibility to interrupt the reaction sequence at the stage of the corresponding optically active 2-hydroxyalkyl- and 2-aminoalkyl 2,3-dihydrofuranes with three stereogenic centers is also presented. Finally, models which account for the formation of the optically active 2,3-dihydrofuranes are proposed.     

Kinetics of surface modification induced by submonolayer electrochemical oxygen adsorption on Pt(1 1 1)

Electrochemical oxygen adsorption/desorption below monolayer level leads to a disordering of platinum single-crystal surfaces vicinal to the (1 1 1) plane. The kinetics can be described by means of a consecutive reaction from (1 1 1)-terrace sites to (1 1 0)-defect sites, in which (1 0 0)-defects act as intermediates. The first oxidation of the electrode reflects independent contributions from terrace and step sites, the latter being structure sensitive. Oxygen adsorption charges amount to a mean value of one electron per step site.     

Local Disorder in Hydrogen Storage Compounds: The Case of Lithium Amide/Imide

Amides and imides of alkali metals are a very promising class of materials for use as a hydrogen‐storage system, as they are able to store and release hydrogen via a chemical route at controllable temperatures and pressures. We critically revise the present picture of the atomic structure of the lightest member (LiNH₂/Li₂NH) by using a combined computational and experimental approach. Specifically, ab initio path integral molecular dynamics simulations and solid‐state ¹H NMR techniques are combined. The results show that the presently assumed local structure might be inconsistent or at least incomplete and needs considerable revision. In particular, the Li atoms turn out to be more mobile and more disordered than suggested by structural data obtained from X‐ray scattering. Also, the configuration of the hydrogen atoms, which is accessible via the NMR experiment and the corresponding first‐principles calculations, is different from the previously assumed data. The computed and experimentally observed ¹H NMR parameters are in very good mutual agreement and illustrate the unusual chemical environment of the hydrogen atoms in this system. Incorporating our results on the new lithium data, we show that the effect of nuclear quantum delocalization for the hydrogen atoms is considerably reduced compared to the perfect crystal structure.     

Determination of 19 drugs of abuse and metabolites in whole blood by high-performance liquid chromatography–tandem mass spectrometry

A high-performance liquid chromatography (LC)–tandem mass spectrometry (MS/MS) method has been developed and validated for the determination of 19 drugs of abuse and metabolites and used in whole blood. The following compounds were included: amphetamine, methylenedioxyamphetamine, methylenedioxyethylamphetamine, methylenedioxymethamphetamine, methamphetamine, cocaine, benzoylecgonine, morphine, 6-acetylmorphine, codeine, methadone, buprenorphine, norbuprenorphine, ketobemidone, tramadol, O-desmethyltramadol, zaleplone, zolpidem, and zopiclone. The sample pretreatment consisted of solid-phase extraction using mixed-mode columns (Isolute Confirm HCX). Deuterated analogues were used as internal standards for all analytes, except for ketobemidone and O-desmethyltramadol. The analytes were separated by a methanol/ammonium formate gradient using high-performance LC (Agilent HPLC 1100) with a 3 mm × 100 mm Varian Pursuit 3 C₁₈ column, 3-μm particle size, and were quantified by MS/MS (Waters Quattro micro tandem quadrupole mass spectrometer) using multiple reaction monitoring in positive mode. Two transitions were used for all analytes, except for tramadol and O-desmethyltramadol. The run time of the method was 35 min including the equilibration time. For all analytes, responses were linear over the range investigated, with R ² > 0.99. One-point calibration was found to be adequate by validation, thereby saving analysis of multiple calibrators. The limits of quantification (LOQs) for the analytes ranged from 0.0005 to 0.01 mg/kg. Absolute recoveries of the analytes were from 34 to 97%, except for zaleplone (6%). Both the interday precision and the intraday precision were less than 15% (20% at the LOQ) for all analytes, except buprenorphine, norburprenorphine, and zaleplone (less than 18%). Accuracy (bias) was within ±15% (±20% at the LOQ) for all analytes, except MDMA and O-desmethyltramadol (within ±19%). No ion suppression or enhancement was seen nor was suppression from coeluted analytes seen. Matrix effects were found to be less than 23% for all analytes, except zopiclone (64%). High-concentration and low-concentration quality control samples gave acceptable values, and the method has been tried in international proficiency test schemes with good results. The present LC-MS/MS method provides a simple, specific, and sensitive solution for the quantification of some of the most frequent drugs of abuse and their metabolites in whole blood. The quantification by LC-MS/MS was successfully applied to 412 forensic cases from October 2008 to mid February 2009, where 267 cases were related to zero-tolerance traffic legislation.     

Mechanistic proposal for the zeolite catalyzed methylation of aromatic compounds

Alkylation and methylation reactions are important reactions in petrochemical production and form part of the reaction mechanism of many hydrocarbon transformation processes. Here, a new reaction mechanism is explored for the zeolite catalyzed methylation of arenes using quantum chemical calculations. It is proposed that the most substituted methylbenzenes, which will reside predominantly on the protonated form when adsorbed in a zeolite, can react directly with a neutral methanol molecule in the vicinity, thereby initiating the methylation reaction without having to return a proton to the zeolite surface. The calculated barriers are quite low, indicating that the suggested mechanism is plausible. This route might explain how the most substituted methylbenzenes can function as efficient reaction intermediates in the methanol to hydrocarbons reaction without themselves acting as catalyst poisons as a consequence of their high proton affinities.