A ruler for determining the position of proteins in membranes

Both the oxygen diffusion rate and the oxygen solubility vary with depth into the interior of biological membranes. The product of these two gradients generates a single gradient, a permeability gradient, which is a smooth continuous function of the distance from the center of the membrane. Using electron paramagnetic resonance and the spin-probe method, the relaxation gradient of oxygen, which is directly proportional to the permeability gradient, is the quantity that can be directly measured in membranes under physiological conditions. The gradient obtained provides a calibrated ruler for determining the membrane depth of residues either from loop regions of membrane-binding proteins or from the membrane-exposed residues of transmembrane proteins. We have determined the relaxation gradient of oxygen in zwitterionic and anionic phospholipid membranes by attaching a single nitroxide probe to a transmembrane alpha-helical polypeptide at specific residues. The peptide ruler was used to determine the depth of penetration of the calcium-binding loops of the C2 domain of cytosolic phospholipase A(2). The positions of selected residues of this membrane-binding protein that penetrate into the membrane, determined using this ruler, compared favorably with previous determinations using more complex methods. The relaxation gradient constrains the possible values of the membrane-dependent oxygen concentration and the oxygen diffusion gradients. The average oxygen diffusion coefficient is estimated to be at least 2-fold smaller in the membrane than that in water.     

Pentakis(methoxycarbonyl)cyclopentadiene,a strong organic acid: Crystal and molecular structures of HC5(CO)2Me)5 and Li[C5(CO2Me)5](H2O)

Contrary to some previously depicted structures, the proton in HC₅(CO₂Me)₅ is bonded to two oxygens rather than to carbon; the lithium ion in Li[C₅(CO₂Me)₅] (H₂O) is tetrahedrally coordinated by two carbonyl oxygens from one C₅ anion, one from a second anion, and a water molecule.     

Thermal analysis and X-ray studies of chiral ferroelectric liquid crystalline materials and their binary mixtures

Summary Thermal properties of a homologous series of ferroelectric liquid crystals S-(-)-[4-(2-n-alkoxy-propionyloxy)]biphenyl-4'-[n-alkoxy-(3,5-dimethyl)]benzoate have been investigated by polarizing optical microscopy and differential scanning calorimetry. The mesophases were identified and confirmed by X-ray too. Three binary mixtures were prepared from the individual homologues. In one of the mixtures (Mix1), the ferroelectric SmC* phase has broadened and became enantiotropic. This mesophase remained monotropic in the other two mixtures (Mix2, Mix3). The chiral nematic N* phase did not appear in Mix1, but remained monotropic for the other two mixtures. Two molecular parameters, the layer spacing and the average intermolecular distance have been calculated from the X-ray results for the homologues and their mixtures. An intercalated tail-to-tail packing of molecules was found both in the single compounds and their mixtures resulting in the layer spacing about half of the molecular length of the single compounds.     

Analytical Detectors Based on Microplasma Spectrometry

Miniaturizing all dimensions of apparatus, such as electronics and computers, is the current trend followed by scientists in various fields. The idea of Lab-on-a-Chip has significantly expanded and found its broad applications in analytical chemistry. Microplasmas can act as a sample excitation source and are the miniaturized versions of full-sized plasmas. These can be created in various forms, such as direct current, microwave induced, capacitively coupled and inductively coupled plasmas. Scaling down the size would reduce the amount of gases, liquids and consumables required, as well as the sample analysis time, which in turn would decrease the operating costs. Therefore, several research groups are involved in the development of microplasmas for utilisation in analytical instruments.     

Optimization and validation of an enzyme immunoassay for the insect growth regulator fenoxycarb

A competitive enzyme-linked immunosorbent assay (ELISA) was developed for the quantitative detection of the insect growth regulator fenoxycarb. Polyclonal rabbit antisera, raised against protein conjugates of four haptenic derivatives of fenoxycarb, were utilized in immobilized antigen-based, competitive immunoassays. With ELISA systems that were both hapten- and carrier-heterologous, most antiserum titers fell in the range of 1:1000-1:30,000. Assay conditions, including concentrations of antisera and coating antigens, were optimized. The effect of pH, organic solvents, and various blocking agents was also investigated. A hapten-homologous and two hapten-heterologous indirect ELISAs allowed fenoxycarb determination in the range of 0.1-85 ng ml⁻¹ with apparent IC₅₀ values of 1.2-2.8 ng ml⁻¹. Cross-reactivities with a number of compounds (e.g. pesticides of related structure, hapten synthesis intermediates, fenoxycarb metabolite, photodegradation products) were determined, and the assay proved highly selective for fenoxycarb. In particular, no significant interference was found with selected pyrethroid and juvenile hormone analog insecticides, phenoxyacetic acid herbicides, and photodegradation products of fenoxycarb. Using spiked water samples, assay performance was validated by SPME/GC-MS.     

Chemical applications of group theory and topology

The following procedure is described for investigating the qualitative dynamics of simple chemical systems: 1) A so-called influence diagram is generated representing the relationships between the reference reactants (phase-determining intermediates); 2) This influence diagram is used to generate a truth table indicating possible transitions between state vectors representing the signs of the time derivatives of of the reference reactant concentrations; 3) The truth table is used to determine a state transition diagram representing the flow topology around unstable equilibrium points; 4) The characteristic equation of the adjacency matrix of the influence diagram is solved in order to determine the presence of such unstable equilibrium points. The two types of qualitative dynamics possible for chemical systems containing two reference reactants and one feedback circuit are bifurcation between two attracting regions (bistability) and limit cycle oscillation. However, in two reference reactant systems oscillation requires an additional self-activating loop to generate the unstable equilibrium point required for its realization. Bistability and limit cycle oscillation are also two of the possible types of qualitative dynamics for chemical systems containing three reference reactants. However, chemical systems with three reference reactants and two or more feedback circuits can also contain interlocking limit cycles, which can lead to toroidal oscillations or chaos. The influence diagrams are given for the systems exhibiting these various types of dynamic behavior along with a summary of the important properties of all 729 possible influences for simple chemical systems containing three reference reactants.     

Semiclassical study of collision-induced predissociation: Comparison of the Landau-Zener model with the method of analytic continuation

Semiclassical calculations are carried out by two methods for the problem of collision-induced predissociation of electronically excited I₂. The first method is that of surface-hopping with the Landau-Zener model. The second method is similar to surface-hopping, except that analytic continuation of the adiabatic potential energy surfaces replaces the Landau-Zener model. Results of the calculations by the two methods compare favorably with each other and with experiment. The possible advantages of the second method are discussed.Camille and Henry Dreyfus Teacher-Scholar, Alfred P. Sloan Research Fellow.     

Comparison of quantitative methods for analysis of polyphasic pharmaceuticals

Adequate very sensitive quantification methods are needed for the development and are also now required for the monitoring of undesirable solid form(s) as routine tests. The pre-requisite for quantitation are selectivity, sensitivity and most important the purity of standards and their proper storage, what is a challenge for metastable forms. Several analytical techniques are available such as X-ray diffraction, spectroscopy, thermal analysis and microcalorimetry. The different steps of the validation of the analytical methods and problems to be solved are discussed. Examples illustrate the different techniques and compare their possible advantages and limits. The relative standard deviation of measurements should allow for checking the homogenization procedure of mixtures for calibration. The validation should be carried out following ICH guidelines for validation of analytical methods. Comparison of different techniques in adequate concentration range add confidence in the analytical results.