High-performance liquid chromatographic determination of (R)-and (S)-proxyphylline in human plasma

A reversed-phase high-performance liquid chromatographic assay has been developed for determination of (R)-(--)-and (S)-(+)-proxyphylline in human plasma. The procedure is based on liquid-solid extraction of proxyphylline from plasma followed by derivatization of extracted proxyphylline with (--)-camphanoyl chloride. The ratio between the enantiomers is calculated from the peak areas of the corresponding diastereoisomeric proxyphylline camphanates after injection into the liquid chromatograph. The recovery of proxyphylline from plasma was 88% (coefficient of variation = 4%) and proxyphylline was detectable from a plasma concentration of 0.2 micrograms/ml. Three different plasma extraction procedures for proxyphylline using Extrelut, Bond Elut, and Chem Elut columns have been developed and compared, and the rate of derivatization of the proxyphylline enantiomers with camphanoyl chloride has been studied.     

Development of small molecule inhibitors and probes of human SUMO deconjugating proteases

Sentrin specific proteases (SENPs) are responsible for activating and deconjugating SUMO (Small Ubiquitin like MOdifier) from target proteins. It remains difficult to study this posttranslational modification due to the lack of reagents that can be used to block the removal of SUMO from substrates. Here, we describe the identification of small molecule SENP inhibitors and active site probes containing aza-epoxide and acyloxymethyl ketone (AOMK) reactive groups. Both classes of compounds are effective inhibitors of hSENPs 1, 2, 5, and 7 while only the AOMKs efficiently inhibit hSENP6. Unlike previous reported peptide vinyl sulfones, these compounds covalently labeled the active site cysteine of multiple recombinantly expressed SENP proteases and the AOMK probe showed selective labeling of these SENPs when added to complex protein mixtures. The AOMK compound therefore represents promising new reagents to study the process of SUMO deconjugation.     

Non-reflecting boundary conditions applicable to general purpose CFD simulators

In simulations of propagating blast waves the effects of artificial reflections at open boundaries can seriously degrade the accuracy of the computations. In this paper, a boundary condition based on a local approximation by a plane traveling wave is presented. The method yields small artificial reflections at open boundaries. The derivation and the theory behind these so-called plane-wave boundary conditions are presented. The method is conceptually simple and is easy to implement in two and three dimensions. These non-reflecting boundary conditions are employed in the three-dimensional computational fluid dynamics (CFD) solver FLACS, capable of simulating gas explosions and blast-wave propagation in complex geometries. Several examples involving propagating waves in one and two dimensions, shock tube and an example of a simulation of a propagating blast wave generated by an explosion in a compressor module are shown. The numerical simulations show that artificial reflections due to the boundary conditions employed are negligible. © 1998 John Wiley & Sons, Ltd.     

Shape resonances in slow electron scattering by aromatic molecules. I. Anthraquinone derivatives

A series of anthraquinone (C(14)O(2)H(8)) derivatives has been studied by means of electron capture negative ion mass spectrometry (ECNI-MS), photoelectron spectroscopy (PES), and AM1 quantum chemical calculations. Mean lifetimes of molecular negative ions M(-.) (MNI) have been measured. The mechanism of long-lived MNI formation in the epithermal energy region of incident electrons has been investigated. A simple model of a molecule (a spherical potential well with the repulsive centrifugal term) has been applied for the analysis of the energy dependence of cross sections at the first stage of the electron capture process. It has been shown that a temporary resonance of MNI at the energy approximately 0.5 eV corresponds to a shape resonance with lifetime 1-2.10(-13) s in the f-partial wave (l = 3) of the incident electron. The next resonant state of MNI at the energy approximately 1.7 eV has been associated with the electron excited Feshbach resonance (whose parent state is a triplet npi* transition). In all cases the initial electron state of the MNI relaxes into the ground state by means of a radiationless transition, and the final state of the MNI is a nuclear excited resonance with a lifetime measurable on the mass spectrometry timescale. Copyright 1999 John Wiley & Sons, Ltd.     

A finite element formulation for global linear stability analysis of a nominally two‐dimensional base flow

A stabilized finite element method, to carry out the linear stability analysis of a two‐dimensional base flow to three‐dimensional perturbations that are periodic along span, is presented. The resulting equations for the time evolution of the disturbance requires a solution to the generalized eigenvalue problem. The analysis is global in nature and is also applicable to non‐parallel flows. Equal‐order‐interpolation functions for velocity and pressure are utilized. Stabilization terms are added to the Galerkin formulation to admit the use of equal‐order‐interpolation functions and to eliminate node‐to‐node oscillations that might arise in advection‐dominated flows. The proposed formulation is tested on two flow problems. First, the mode transitions in the circular Couette flow are investigated. Two scenarios are considered. In the first one, the outer cylinder is at rest, while the inner one spins. Two linearly unstable modes are identified. The primary mode is real and represents the axisymmetric Taylor vortices. The second mode is complex and consists of spiral vortices. For the counter‐rotating cylinders, the primary transition is via the appearance of spiral vortices. Excellent agreement with results from earlier studies is observed. The formulation is also utilized to investigate the parallel and oblique modes of vortex shedding past a cylinder for the Re = 100 flow. It is found that the flow is associated with a large number of unstable oblique shedding modes. The parallel mode of vortex shedding is a special case of this family of modes and is associated with the largest growth rate. Copyright © 2014 John Wiley & Sons, Ltd.     

Identification and evaluation of small molecule pan-caspase inhibitors in Huntington's disease models

Huntington's Disease (HD) is characterized by?a mutation in the huntingtin (Htt) gene encoding an expansion of glutamine repeats on the N terminus of the Htt protein. Numerous studies have identified Htt proteolysis as a critical pathological event in HD postmortem human tissue and mouse HD models, and proteases known as caspases have emerged as attractive HD therapeutic targets. We report the use?of the substrate activity screening method against caspase-3 and -6 to identify three novel, pan-caspase inhibitors that block proteolysis of Htt at caspase-3 and -6 cleavage sites. In HD models these irreversible inhibitors suppressed Hdh(111Q/111Q)-mediated toxicity and rescued rat striatal and cortical neurons from cell death. In this study, the identified nonpeptidic caspase inhibitors were used to confirm the role of caspase-mediated Htt proteolysis in HD. These results further implicate caspases as promising targets for HD therapeutic development.     

On specifications,subset types and interpretation of proposition in type theory

In this paper we discuss some practical aspects of using type theory as a programming and specification language, where the viewpoint is to use it not only as a basis for program synthesis but also as a programming language with a programming logic allowing us to do ordinary verification.The subset type has been added to type theory in order to avoid irrelevant information in programs. We give an example of a proof which illustrates the problems that may occur if the subset type is used in specifications when we have the standard interpretation of propositions as types. Harrop-formulas and Squash are then discussed as solutions to these problems. It is argued that they are not acceptable from a practical point of view.An extension of the theory to include the two new judgment forms:A is a proposition, andA is true, is then given and explained in terms of the old theory. The logical constants are no longer identified with the corresponding type theoretical constants, but propositions are interpreted as Gödel formulas, which allow us to introduce and justify logical rules similar to rules for classical logic. The interpretation is extended to include predicates defined by using reflections of the ordinary definition of Gödel formulas in a type of small propositions.The programming example is then revisited and stronger elimination rules are discussed.     

An optimized activity-based probe for the study of caspase-6 activation

Although significant efforts have been made to understand the mechanisms of caspase activation during apoptosis, many questions remain regarding how and when executioner caspases get activated. We describe the design and synthesis of an activity-based probe that labels caspase-3/-6/-7, allowing direct monitoring of all executioner caspases simultaneously. This probe has enhanced in vivo properties and reduced cross-reactivity compared to our previously reported probe, AB50. Using this probe, we find that caspase-6 undergoes a conformational change and can bind substrates even in the absence of cleavage of the proenzyme. We also demonstrate that caspase-6 activation does not require active caspase-3/-7, suggesting that it may autoactivate or be cleaved by other proteases. Together, our results suggest that caspase-6 activation proceeds through a unique mechanism that may be important for its diverse biological functions.