Conformations, vibrational frequencies and Raman intensities of short-chain fatty acid methyl esters using DFT with 6-31G(d) and Sadlej pVTZ basis sets

Density functional calculations, using B3LPY/6-31G(d) methods, have been used to investigate the conformations and vibrational (Raman) spectra of three short-chain fatty acid methyl esters (FAMEs) with the formula CnH2nO2 (n = 3-5). In all three FAMEs, the lowest energy conformer has a simple 'all-trans' structure but there are other conformers, with different torsions about the backbone, which lie reasonably close in energy to the global minimum. One result of this is that the solid samples we studied do not appear to consist entirely of the lowest energy conformer. Indeed, to account for the 'extra' bands that were observed in the Raman data but were not predicted for the all-trans conformer, it was necessary to add-in contributions from other conformers before a complete set of vibrational assignments could be made. Provided this was done, the agreement between experimental Raman frequencies and 6-31G(d) values (after scaling) was excellent, RSD = 12.6 cm(-1). However, the agreement between predicted and observed intensities was much less satisfactory. To confirm the validity of the approach followed by the 6-3 1 G(d) basis set, we used a larger basis set, Sadlej pVTZ, and found that these calculations gave accurate Raman intensities and simulated spectra (summed from two different conformers) that were in quantitative agreement with experiment. In addition, the unscaled Sadlej pVTZ, and the scaled 6-3 1 G(d) calculations gave the same vibrational mode assignments for all bands in the experimental data. This work provides the foundation for calculations on longer-chain FAMEs (which are closer to those found as triglycerides in edible fats and oils) because it shows that scaled 6-3 1 G(d) calculations give equally accurate frequency predictions, and the same vibrational mode assignments, as the much more CPU-expensive Sadlej pVTZ basis set calculations.

A prospective clinical trial of a real-time polymerase chain reaction assay for the diagnosis of candidemia in nonneutropenic, critically ill adults

Background. Invasive Candida infection among nonneutropenic, critically ill adults is a clinical problem that has received increasing attention in recent years. Poor performance of extant diagnostic modalities has promoted risk-based, preemptive prescribing in view of the poor outcomes associated with inadequate or delayed antifungal therapy; this risks unnecessary overtreatment. A rapid, reliable diagnostic test could have a substantial impact on therapeutic practice in this patient population.

Methods. Three TaqMan-based real-time polymerase chain reaction assays were developed that are capable of detecting the main medically important Candida species, categorized according to the likelihood of fluconazole susceptibility. Assay 1 detected Candida albicans, Candida parapsilosis, Candida tropicalis, and Candida dubliniensis. Assays 2 and 3 detected Candida glabrata and Candida krusei, respectively. The clinical performance of these assays, applied to serum, was evaluated in a prospective trial of nonneutropenic adults in a single intensive care unit.

Results. In all, 527 specimens were obtained from 157 participants. All 3 assays were run in parallel for each specimen; they could be completed within 1 working day. Of these, 23 specimens were obtained from 23 participants categorized as having proven Candida infection at the time of sampling. If a single episode of Candida famata candidemia was excluded, the estimated clinical sensitivity, specificity, and positive and negative predictive values of the assays in this trial were 90.9%, 100%, 100% and 99.8%, respectively.

Conclusions. These data suggest that the described assays perform well in this population for enhancing the diagnosis of candidemia. The extent to which they may affect clinical outcomes, prescribing practice, and cost-effectiveness of care remains to be ascertained.

Kinetic analysis of the cleavage of human protease-activated receptor-1/2/3 and 4 using quenched-fluorescent peptide substrates

Protease-activated receptors [PARs] are a family of G-protein-coupled seven-transmembrane domain receptors that are activated by proteolytic cleavage of their amino-terminal exodomain. To characterize the cleavage rate of human PAR-1 / 2 / 3 and 4 by trypsin and thrombin, four synthetic quenched-fluorescent peptide substrates have been synthesized. Each substrate consisted of a ten-residue peptide spanning the receptor activation cleavage site and using progress-curve kinetics, k(cat)/K-m values were determined.