Isobaric vapor–liquid equilibria for the quaternary reactive system: Ethanol + water + ethyl lactate + lactic acid at 101.33 kPa

Isobaric vapor–liquid equilibrium (VLE) data of the reactive quaternary system ethanol (1) + water (2) + ethyl lactate (3) + lactic acid (4) have been determined experimentally. Additionally, the reaction equilibrium constant was calculated for each VLE experimental data. The experimental VLE data were correlated using the UNIQUAC equation to describe the chemical and phase equilibria simultaneously. For some of the non-reactive binary systems, UNIQUAC binary interaction parameters were obtained from the literature. The rest of the binary UNIQUAC parameters were obtained by correlating the experimental quaternary VLE data obtained in this work. A maximum pressure azeotrope at high water concentration for the binary reactive system ethyl lactate + water has been calculated.     

VACUUM ULTRAVIOLET NATURAL AND MAGNETIC CIRCULAR DICHROISM WITH CONVENTIONAL SOURCES AND SYNCHROTRON RADIATION

Vacuum ultraviolet natural and magnetic circular dichroism measurements have added significantly to our knowledge of the geometric and electronic structure of molecules and have provided a better understanding of the correct approach for theoretical calculations. In this review, I define natural and magnetic circular dichroism, and discuss information obtained with these techniques. The instrumentation for vacuum ultraviolet natural and magnetic circular dichroism is reviewed, from its beginning with conventional sources to the present time use of synchrotron radiation. The future possibilities and challenges for these measurements are examined with particular reference to making measurements to higher energies.     

PHOTOSENSITIZATION OF MURINE TUMOR, VASCULATURE and SKIN BY 5-AMINOLEVULINIC ACID-INDUCED PORPHYRIN

Abstract— The effects of topical and systemic administration of 5-aminolevulinic acid (ALA) were examined in several murine tumor systems with regard to porphyrin accumulation kinetics in tumor, skin and blood, vascular and tumor cell photosensitization and tumor response after light exposure. Marked, transient increases in porphyrin levels were observed in tumor and skin after systemic and topical ALA. Rapid, transient, dose-dependent porphyrin increases were also observed in blood; these were pronounced after systemic ALA injection and mild after topical application. They were highest within 1 h after ALA injection, thereafter declining rapidly. This matched the clearing kinetics of injected exogenous protoporphyrin IX (PpIX). Initially, vascular photosensitivity changed inversely to blood porphyrin levels, increasing gradually up to 5 h post-ALA, as porphyrin was clearing from the bloodstream. This pattern was again matched by injected, exogenous PpIX. After therapeutic tumor treatment vascular disruption of the tumor bed, while observed, was incomplete, especially at the tumor base. Minimal direct tumor cell kill was found at low photodynamic therapy (PDT) doses (250 mg/kg ALA, 135 J/cm² light). Significant, but limited (<1 log) direct photodynamic tumor cell kill was obtained when the PDT dose was raised to 500 mg/kg systemic ALA, followed 3 h later by 270 J/cm², a dose that was however toxic to the animals. The further reduction of clonogenic tumor cells over 24 h following treatment was moderate and probably limited by the incomplete disruption of the vasculature. Tumor responses were highest when light treatment was carried out at the time of highest tumor porphyrin content rather than at the time of highest vascular photosensitivity. Tumor destruction did not reach the tumor base, regardless of treatment conditions.     

Functionally relevant protein motions: extracting basin-specific collective coordinates from molecular dynamics trajectories

Functionally relevant motion of proteins has been associated with a number of atoms moving in a concerted fashion along so-called "collective coordinates." We present an approach to extract collective coordinates from conformations obtained from molecular dynamics simulations. The power of this technique for differentiating local structural fluctuations between classes of conformers obtained by clustering is illustrated by analyzing nanosecond-long trajectories for the response regulator protein Spo0F of Bacillus subtilis, generated both in vacuo and using an implicit-solvent representation. Conformational clustering is performed using automated histogram filtering of the inter-C(alpha) distances. Orthogonal (varimax) rotation of the vectors obtained by principal component analysis of these interresidue distances for the members of individual clusters is key to the interpretation of collective coordinates dominating each conformational class. The rotated loadings plots isolate significant variation in interresidue distances, and these are associated with entire mobile secondary structure elements. From this we infer concerted motions of these structural elements. For the Spo0F simulations employing an implicit-solvent representation, collective coordinates obtained in this fashion are consistent with the location of the protein's known active sites and experimentally determined mobile regions.     

An unexpected P-cluster like intermediate en route to the nitrogenase FeMo-co

The nitrogenase MoFe protein contains two different FeS centers, the P-cluster and the iron–molybdenum cofactor (FeMo-co). The former is a [Fe₈S₇] center responsible for conveying electrons to the latter, a [MoFe₇S₉C-(R)-homocitrate] species, where N₂ reduction takes place. NifB is arguably the key enzyme in FeMo-co assembly as it catalyzes the fusion of two [Fe₄S₄] clusters and the insertion of carbide and sulfide ions to build NifB-co, a [Fe₈S₉C] precursor to FeMo-co. Recently, two crystal structures of NifB proteins were reported, one containing two out of three [Fe₄S₄] clusters coordinated by the protein which is likely to correspond to an early stage of the reaction mechanism. The other one was fully complemented with the three [Fe₄S₄] clusters (RS, K1 and K2), but was obtained at lower resolution and a satisfactory model was not obtained. Here we report improved processing of this crystallographic data. At odds with what was previously reported, this structure contains a unique [Fe₈S₈] cluster, likely to be a NifB-co precursor resulting from the fusion of K1- and K2-clusters. Strikingly, this new [Fe₈S₈] cluster has both a structure and coordination sphere geometry reminiscent of the fully reduced P-cluster (P^N-state) with an additional μ²-bridging sulfide ion pointing toward the RS cluster. Comparison of available NifB structures further unveils the plasticity of this protein and suggests how ligand reorganization would accommodate cluster loading and fusion in the time-course of NifB-co synthesis.

The K-cluster of NifB as a key intermediate in the synthesis of the nitrogenase active site supports [Fe₄S₄] cluster fusion occurs before carbide and sulfide insertion and displays ligand spatial arrangement reminiscent to that of the P-cluster.     

Measurement of resistant starch by enzymatic digestion in starch and selected plant materials: collaborative study

Interlaboratory performance statistics was determined for a method developed to measure the resistant starch (RS) content of selected plant food products and a range of commercial starch samples. Food materials examined contained RS (cooked kidney beans, green banana, and corn flakes) and commercial starches, most of which naturally contain, or were processed to yield, elevated RS levels. The method evaluated was optimized to yield RS values in agreement with those reported for in vivo studies. Thirty-seven laboratories tested 8 pairs of blind duplicate starch or plant material samples with RS values between 0.6 (regular maize starch) and 64% (fresh weight basis). For matrixes excluding regular maize starch, repeatability relative standard deviation (RSDr) values ranged from 1.97 to 4.2%, and reproducibility relative standard deviation (RSDR) values ranged from 4.58 to 10.9%. The range of applicability of the test is 2-64% RS. The method is not suitable for products with <1% RS (e.g., regular maize starch; 0.6% RS). For such products, RSDr and RSDR values are unacceptably high.     

Fractionation of metals and metalloids by chemical bonding from particles accumulated by electrostatic precipitation in an Argentine thermal power plant

A study was undertaken to evaluate the distribution of Al, As, Cr, Cu, Fe, Mn, Ni, Pb, Ti, V and Zn in fly ashes collected in the electrostatic precipitator of a thermal power plant in San Nicolás (Argentina). Five samples were collected during one week of operation. For the fractionation, the scheme applied consisted in extracting the elements in four fractions namely (i) soluble and exchangeable elements; (ii) carbonates, oxides and reducible elements; (iii) bound to sulfidic metals; and (iv) residual elements. Metals and metalloids at μg g^− 1 level were determined in each fraction by inductively coupled plasma optical emission spectrometry (ICP OES). For validation, a standard reference material (SRM 1633 coal fly ash) from NIST was subjected to the same chemical sequential extraction procedure that the samples. X-ray diffraction powder (XRD) analysis and scanning electron microscopy (SEM) were used to characterize the major minerals present in the matrix. Total analyte concentration (in μg g^− 1) varied from 10.6 for Pb to 17,622 for Al. Minimum and maximum concentrations (in μg g^− 1) found in individual samples in the four fractions were: Al, 92.7–9668; As, < 0.3–143; Cr, 2.0–10.4; Cu, < 0.2–35.6; Fe, < 0.3–4992; Mn, < 0.1–128; Ni, < 0.3–139; Pb, < 0.5–9.1; Ti, < 0.3–2243; V, 17.0–112.9; and Zn, < 0.1–68.2. The leachability of the 11 elements under study proved to be different. Low percentages of Al (1%), V (7%) and Cr (8%) were detected in the most bioavailable fraction. Arsenic was found to be most abundant in the non-silicate phase, represented by the second and third fractions, while Cr, Fe, Ni, Pb and Zn were mostly associated to the residual fraction.     

Trapping H- bound to the nitrogenase FeMo-cofactor active site during H2 evolution: characterization by ENDOR spectroscopy

We here show that the iron-molybdenum (FeMo)-cofactor of the nitrogenase alpha-70(Ile) molybdenum-iron (MoFe) protein variant accumulates a novel S = (1)/(2) state that can be trapped during the reduction of protons to H(2). (1,2)H-ENDOR measurements disclose the presence of two protons/hydrides (H(+/)(-)) whose hyperfine tensors have been determined from two-dimensional field-frequency (1)H ENDOR plots. The two H(+/)(-) have large isotropic hyperfine couplings, A(iso)( )() approximately 23 MHz, which shows they are bound to the cofactor. The favored analysis for these plots indicates that the two H(+/)(-) have the same principal values, which indicates that they are chemically equivalent. The tensors are further related to each other by a permutation of the tensor components, which indicates an underlying symmetry of binding relative to the cofactor. At present, no model for the structure of the iron-molybdenum (FeMo)-cofactor in the S = (1)/(2) state trapped during the reduction of H(+) can be shown unequivocally to satisfy all of the constraints generated by the ENDOR analysis. The data disfavors any model that involves protonation of sulfides, and thus suggests that the intermediate instead contains two chemically equivalent bound hydrides; it appears unlikely that these are terminal monohydrides.     

pKa calculations in solution and proteins with QM/MM free energy perturbation simulations: a quantitative test of QM/MM protocols

The accuracy of biological simulations depends, in large part, on the treatment of electrostatics. Due to the availability of accurate experimental values, calculation of pKa provides stringent evaluation of computational methods. The generalized solvent boundary potential (GSBP) and Ewald summation electrostatic treatments were recently implemented for combined quantum mechanical and molecular mechanics (QM/MM) simulations by our group. These approaches were tested by calculating pKa shifts due to differences in electronic structure and electrostatic environment; the shifts were determined for a series of small molecules in solution, using various electrostatic treatments, and two residues (His 31, Lys 102) in the M102K T4-lysozyme mutant with large pKa shifts, using the GSBP approach. The calculations utilized a free energy perturbation scheme with the QM/MM potential function involving the self-consistent charge density functional tight binding (SCC-DFTB) and CHARMM as the QM and MM methods, respectively. The study of small molecules demonstrated that inconsistent electrostatic models produced results that were difficult to correct in a robust manner; by contrast, extended electrostatics, GSBP, and Ewald simulations produced consistent results once a bulk solvation contribution was carefully chosen. In addition to the electrostatic treatment, the pKa shifts were also sensitive to the level of the QM method and the scheme of treating QM/MM Coulombic interactions; however, simple perturbative corrections based on SCC-DFTB/CHARMM trajectories and higher level single point energy calculations were found to give satisfactory results. Combining all factors gave a root-mean-square difference of 0.7 pKa units for the relative pKa values of the small molecules compared to experiment. For the residues in the lysozyme, an accurate pKa shift was obtained for His 31 with multiple nanosecond simulations. For Lys 102, however, the pKa shift was estimated to be too large, even after more than 10 nanosecond simulations for each lambda window; the difficulty was due to the significant, but slow, reorganization of the protein and water structure when Lys 102 was protonated. The simulations support that Lys 102 is deprotonated in the X-ray structure and the protein is highly destabilized when this residue is protonated.     

Computational studies of the 3-Dimensional structure of cyclopenta polycyclic aromatic hydrocarbons containing a gulf region

Recently, some cyclopenta-fused polyaromatic hydrocarbons, an environmentally relevant subclass of chemicals, have been shown to have carcinogenic activity in animals. It has been suggested that benz[l] aceanthrylene ( I ), an active member of this subclass with a gulf region, has a trans dihydrodiol metabolite that is nonplanar and has two distinct spatial configurations. We have used MMP 2(85) and AM 1 to investigate the three-dimensional structure of this dihydrodiol and other similar derivatives of ( I ) and have found that although ( I ) is somewhat nonplanar the relevant derivatives are all nearly planar. Further, we have computed potential functions for the bending of the angular ring in the gulf region using MMP 2(85), AM 1, and ab initio computed energies for AM 1 spatial configurations and find that these molecules all have only a single potential minimum. We have performed the same calculations for benzo[c]phenanthren and its 1,12 dimethyl derivative, molecules with a similar gulf region for which crystallographic data exists. In agreement with that data, we find that two distinct spatial configurations exist separated by significant barries. The differences between the results generated by the three different methods of computation will be discussed.