Direct evaluation of multicomponent phase equilibria using flat-histogram methods

We present a method for directly locating density-driven phase transitions in multicomponent systems. Phase coexistence conditions are determined through manipulation of a total density probability distribution evaluated over a density range that includes both coexisting phases. Saturation quantities are determined through appropriate averaging of density-dependent mean values of a given property of interest. We discuss how to implement the method in both the grand-canonical and isothermal-isobaric semigrand ensembles. Calculations can be conducted using any of the recently introduced flat-histogram techniques. Here, we combine the general algorithm with a transition-matrix approach to produce an efficient self-adaptive technique for determining multicomponent phase equilibrium properties. To assess the performance of the new method, we generate phase diagrams for a number of binary and ternary Lennard-Jones mixtures.     

A computational study of hydration, solution structure, and dynamics in dilute carbohydrate solutions

We report results from a molecular simulation study of the structure and dynamics of water near single carbohydrate molecules (glucose, trehalose, and sucrose) at 0 and 30 degrees C. The presence of a carbohydrate molecule has a number of significant effects on the microscopic water structure and dynamics. All three carbohydrates disrupt the tetrahedral arrangement of proximal water molecules and restrict their translational and rotational mobility. These destructuring effects and slow dynamics are the result of steric constraints imposed by the carbohydrate molecule and of the ability of a carbohydrate to form stable H bonds with water, respectively. The carbohydrates induce a pronounced decoupling between translational and rotational motions of proximal water molecules.     

Free radical-mediated vinyl amination: access to N,N-dialkyl enamines and their beta-stannyl and beta-thio derivatives

[reaction: see text] The first examples of free radical-mediated vinyl amination are described by nonconventional vinyl radical addition to azomethine nitrogen. This new vinyl amination protocol is mild and provides convenient synthetic access to nonstabilized N,N-dialkyl enamines and tandem bond-forming processes.     

RAMPED-UP NMR: multiplexed NMR-based screening for drug discovery

The crucial step in drug discovery is the identification of a lead compound from a vast chemical library by any number of screening techniques. NMR-based screening has the advantage of directly detecting binding of a compound to the target. The spectra resulting from these screens can also be very complex and difficult to analyze, making this an inefficient process. We present here a method, RAMPED-UP NMR, (Rapid Analysis and Multiplexing of Experimentally Discriminated Uniquely Labeled Proteins using NMR) which generates simple spectra which are easy to interpret and allows several proteins to be screened simultaneously. In this method, the proteins to be screened are uniquely labeled with one amino acid type. There are several benefits derived from this unique labeling strategy: the spectra are greatly simplified, resonances that are most likely to be affected by binding are the only ones observed, and peaks that yield little or no information upon binding are eliminated, allowing the analysis of multiple proteins easily and simultaneously. We demonstrate the ability of three different proteins to be analyzed simultaneously for binding to two different ligands. This method will have significant impact in the use of NMR spectroscopy for both the lead generation and lead optimization phases of drug discovery by its ability to increase screening throughput and the ability to examine selectivity. To the best of our knowledge, this is the first time in any format that multiple proteins can be screened in one tube.     

Variable coordination geometries at the diiron(II) active site of ribonucleotide reductase R2

The R2 subunit of Escherichia coli ribonucleotide reductase contains a dinuclear iron center that generates a catalytically essential stable tyrosyl radical by one electron oxidation of a nearby tyrosine residue. After acquisition of Fe(II) ions by the apo protein, the resulting diiron(II) center reacts with O(2) to initiate formation of the radical. Knowledge of the structure of the reactant diiron(II) form of R2 is a prerequisite for a detailed understanding of the O(2) activation mechanism. Whereas kinetic and spectroscopic studies of the reaction have generally been conducted at pH 7.6 with reactant produced by the addition of Fe(II) ions to the apo protein, the available crystal structures of diferrous R2 have been obtained by chemical or photoreduction of the oxidized diiron(III) protein at pH 5-6. To address this discrepancy, we have generated the diiron(II) states of wildtype R2 (R2-wt), R2-D84E, and R2-D84E/W48F by infusion of Fe(II) ions into crystals of the apo proteins at neutral pH. The structures of diferrous R2-wt and R2-D48E determined from these crystals reveal diiron(II) centers with active site geometries that differ significantly from those observed in either chemically or photoreduced crystals. Structures of R2-wt and R2-D48E/W48F determined at both neutral and low pH are very similar, suggesting that the differences are not due solely to pH effects. The structures of these "ferrous soaked" forms are more consistent with circular dichroism (CD) and magnetic circular dichroism (MCD) spectroscopic data and provide alternate starting points for consideration of possible O(2) activation mechanisms.     

Seasonal fluctuation of DNA photodamage in marine plankton assemblages at Palmer Station, Antarctica

Ultraviolet radiation-induced DNA damage frequencies were measured in DNA dosimeters and natural plankton communities during the austral spring at Palmer Station, Antarctica, during the 1999-2000 field season. We found that the fluence of solar ultraviolet-B radiation (UV-B) at the earth's surface correlated with stratospheric ozone concentrations, with significant ozone depletion observed because of "ozone hole" conditions. To verify the interdependence of ozone depletion and DNA damage in natural microbial communities, seawater was collected daily or weekly from Arthur Harbor at Palmer Station, Antarctica, throughout "ozone season," exposed to ambient sunlight between 0600 and 1800 h and fractionated using membrane filtration to separate phytoplankton and bacterioplankton populations. DNA from these fractions was isolated and DNA damage measured using radioimmunoassay. Under low-ozone conditions cyclobutane dimer concentrations in bacterioplankton and phytoplankton communities were maximal. DNA damage measured in dosimeters correlated closely with ozone concentrations and UV-B fluence. Our studies offer further support to the theory that stratospheric deozonation is detrimental to marine planktonic organisms in the Southern Ocean.     

Uses and analyses of curtin-hammett/winstein-holness systems involving second order reactions

The Curtin-Hammett (C-H) principle and the Winstein-Holness (W-H) equation approximate the product ratio and overall rate constant of reaction for systems involving a starting material which exists in two forms, each of which reacts via first-order kinetics to give a different product. The C-H/W-H approximations are valid when the rates of isomer interconversion are significantly faster than the rates of product formation. The present treatment encompasses non-first-order reactions to product. A numerical predictor-corrector technique is used to show (1) that relative reagent concentration can affect both the product ratio and the observed rates of product formation; (2) that the absolute concentration of reagent and substrate can affect the kinetics; and (3) that factors (1) and (2) above can affect the validity of the C-H/W-H approximations for non-first-order C-H/W-H schemes.     

Synthesis and characterization of highly ordered mesoporous thin films with -COOH terminated pore surfaces

Highly ordered mesoporous inorganic-organic hybrid thin films with covalently bonded carboxylic acid (-COOH) terminal groups on the pore surfaces were synthesized by evaporation induced self-assembly of tetraethoxysilane, organosilanes, and a nonionic surfactant followed by acid hydrolysis and characterized using transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, surface acoustic wave (SAW) based N2 sorption, and thermogravimetric analysis (TGA) techniques.     

Determination of malachite green and leucomalachite green in a variety of aquacultured products by liquid chromatography with tandem mass spectrometry detection

A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed for determining the residues of malachite green (MG) and leucomalachite green (LMG) in a number of aquatic species. MG and its metabolite were extracted from homogenized tissues with a perchloric acid-acetonitrile solution; the extract was centrifuged; and an aliquot was taken, concentrated, and passed through a chemically bonded octadecyl C18 solid-phase extraction column. The compounds of interest were eluted with acetonitrile, and the eluate was evaporated to dryness. The residue was dissolved in acetonitrile and diluted with water in preparation for analysis by LC/MS/MS. MG and its metabolite were determined by reversed-phase LC using a Luna C18 column with an ammonium hydroxide-formic acid buffer in acetonitrile gradient and MS/MS detection using multiple reaction monitoring. Calibration curves were linear for all analyses between 5 and 500 pg injected for both analytes, with recoveries ranging from 81% for LMG to 98% for MG in salmon spiked at the 1 ng/g level. Detection limits of 0.1 ng/g for both MG and LMG were easily obtainable using the recommended method. The operational errors, interferences, and recoveries for spiked samples compared favorably with those obtained by established methodology. The recommended method is simple, rapid, and specific for monitoring residues of MG and LMG in a number of aquatic species.