Assessing the effects of diurnal variation on the composition of human parotid saliva: quantitative analysis of native peptides using iTRAQ reagents

Changes in salivary composition correlate with disease susceptibility, disease state, or both. However, use of saliva for diagnostic purposes is complicated by the gland-specific effects of circadian rhythm or diurnal variation. We recently characterized a suite of peptides in the < or =10-kDa fraction of human parotid saliva that included many novel species. In this study, we used novel iTRAQ labeling chemistry to investigate possible diurnal effects on peptide generation. We collected samples produced by gustatory stimulation as the ductal secretions at four time points under conditions that minimized proteolysis, pooled them according to collection time, and isolated the LMW fractions. Samples collected at each collection time were derivatized with a different isobaric iTRAQ reagent. The labeled samples were combined, separated by reversed-phase HPLC, co-spotted with matrix on MALDI targets, and analyzed by MALDI TOF/TOF mass spectrometry. With this approach, we achieved relative quantification of the parotid peptides at four time points. In several cases, abundance during the day changed dramatically. iTRAQ tagging improved the efficiency of MS/MS fragmentation, which in turn allowed the identification of several novel peptides. Our results demonstrated both the utility of this method and the importance of diurnal effects on the composition of the human parotid saliva peptidome.     

Combination of analyte protectants to overcome matrix effects in routine GC analysis of pesticide residues in food matrixes

Analyte protectants were previously defined as compounds that strongly interact with active sites in the gas chromatographic (GC) system, thus decreasing degradation, adsorption, or both of coinjected analytes. In this study, we evaluated various combinations of promising analyte protectants for the volatility range of GC-amenable pesticides using GC/quadrupole mass spectrometry (MS) and 1-microL hot splitless injection for sample introduction. A mixture of ethylglycerol, gulonolactone, and sorbitol (at 10, 1, and 1 mg/mL, respectively, in the injected samples) was found to be the most effective in minimizing losses of susceptible analytes and significantly improving their peak shapes (due to reduction of peak tailing). When added to final sample extracts and matrix-free calibration standards alike, these analyte protectants induced a similar response enhancement in both instances, resulting in effective equalization of the matrix-induced response enhancement effect even after a large number of fruit and vegetable extract injections. As compared to matrix-matched standardization, the analyte protectant approach offers a more convenient solution to the problems associated with calibration in routine GC/MS analysis of pesticide residues and possibly other susceptible analyte types in diverse samples. Moreover, the use of analyte protectants also substantially reduced another adverse matrix-related effect caused by gradual build-up of nonvolatile matrix components in the GC system, thus improving ruggedness and, consequently, reducing need for frequent maintenance.     

Acute cardio-respiratory changes induced by hyperpnoea using a respiratory muscle trainer

The aim of this study was to examine the cardio-respiratory effects of voluntary hyperpnoea using a respiratory muscle trainer (RMT) with three different sized rebreathing bags. In particular, the effects of hyperpnoea on inspired and end-tidal gas concentrations were determined. Seven adult males completed three 30 min bouts of hyperpnoea using optimal, oversized and undersized rebreathing bags. Inspired (F(I)) and expired end-tidal (F(ET)) O2 and CO2 concentrations, arterial O2 saturation (S(AO2)) and heart rate were measured during hyperpnoea. Before and after a bout of hyperpnoea, pulmonary function and blood pressure (BP) were assessed. Data were analysed using a two-way repeated-measures ANOVA, with p < 0.05 considered significant. Three subjects experienced discomfort during hyperpnoea and stopped after 20 min. During hyperpnoea, the F(ETCO2) was maintained at 4.6 +/- 0.7% irrespective of bag size. The increase in F(ICO2) over time reached 0.5 +/- 0.5% at 20 min. The F(IO2) fell to 19.4 +/- 0.8% at 20 min, and S(AO2) decreased to 97%. Heart rate and systolic BP increased slightly, but independently of rebreathing bag volume. No changes in pulmonary function or diastolic BP were found. It is concluded that the RMT maintained a constant F(ETCO2) at the expense of a mild hypoxia. The acute effects of hyperpnoea on the cardio-respiratory system are generally mild, but not always tolerable for 30 min.     

Conformal nanocoating of zirconia nanoparticles by atomic layer deposition in a fluidized bed reactor

Primary zirconia nanoparticles were conformally coated with alumina ultrathin films using atomic layer deposition (ALD) in a fluidized bed reactor. Alternating doses of trimethylaluminium and water vapour were performed to deposit Al(2)O(3) nanolayers on the surface of 26?nm zirconia nanoparticles. Transmission Fourier transform infrared spectroscopy was performed ex situ. Bulk Al(2)O(3) vibrational modes were observed for coated particles after 50 and 70?cycles. Coated nanoparticles were also examined with transmission electron microscopy, high-resolution field emission scanning electron microscopy and energy dispersive spectroscopy. Analysis revealed highly conformal and uniform alumina nanofilms throughout the surface of zirconia nanoparticles. The particle size distribution and surface area of the nanoparticles are not affected by the coating process. Primary nanoparticles are coated individually despite their high aggregation tendency during fluidization. The dynamic aggregation behaviour of zirconia nanoparticles in the fluidized bed plays a key role in the individual coating of nanoparticles.     

Two-dimensional self-assembly into multicomponent hydrogen-bonded nanostructures

By means of scanning tunneling microscopy, we have explored the two-dimensional self-assembly of functional bicomponent hydrogen-bonding dye systems, leading to well-defined patterns, different from those of the individual components, and providing design rules to immobilize multicomponent systems at the liquid-solid interface.     

2D self-assembly of oligo(p-phenylene vinylene) derivatives: from dimers to chiral rosettes

Enantiomerically pure oligo(p-phenylene vinylene) diaminotriazine derivatives and a short structurally related achiral diaminotriazine derivative, all having a rigid backbone in common, are studied to self-assemble at the solution-graphite interface by scanning tunneling microscopy. As a function of the length of the backbone, different two-dimensional motifs are formed (dimers and rosettes) that are rationalized in terms of the balance between different intermolecular interactions, in this case, intermolecular hydrogen bonding and the packing requirements of the alkyl chains on a graphite surface. In addition, the effect of molecular chirality on monolayer chirality is investigated, revealing molecular size-dependent expressions of the monolayer chirality.     

Native state kinetic stabilization as a strategy to ameliorate protein misfolding diseases: a focus on the transthyretin amyloidoses

Small molecule-mediated protein stabilization inside or outside of the cell is a promising strategy to treat protein misfolding/misassembly diseases. Herein we focus on the transthyretin (TTR) amyloidoses and demonstrate that preferential ligand binding to and stabilization of the native state over the dissociative transition state raises the kinetic barrier of dissociation (rate-limiting for amyloidogenesis), slowing and in many cases preventing TTR amyloid fibril formation. Since T119M-TTR subunit incorporation into tetramers otherwise composed of disease-associated subunits also imparts kinetic stability on the tetramer and ameliorates amyloidosis in humans, it is likely that small molecule-mediated native state kinetic stabilization will also alleviate TTR amyloidoses.     

Analysis of nucleic acids by FTICR MS

Fourier transform ion cyclotron resonance (FTICR) mass spectrometry represents a unique platform with which to study nucleic acids and non-covalent complexes containing nucleic acids moieties. In particular, systems in which very high mass measurement accuracy is required, very complex mixtures are to be analyzed, or very limited amounts of sample are available may be uniquely suited to interrogation by FTICR mass spectrometry. Although the FTICR platform is now broadly deployed as an integral component of many high-end proteomics-based research efforts, momentum is still building for the application of the platform towards nucleic acid-based analyses. In this work, we review fundamental aspects of nucleic acid analysis by FTICR, focusing primarily on the analysis of DNA oligonucleotides but also describing applications related to the characterization of RNA constructs. The goal of this review article is to give the reader a sense of the breadth and scope of the status quo of FTICR analysis of nucleic acids and to summarize a few recently published reports in which researchers have exploited the performance attributes of FTICR to characterize nucleic acids in support of basic and applied research disciplines including genotyping, drug discovery, and forensic analyses.     

Increased biological hydrogen production with reduced organic loading

An experimental matrix consisting of reactor hydraulic retention time (HRT) and glucose loading rate was tested to understand the effect of organic loading on H2 production in chemostat reactors. In order to vary the glucose loading rate over a range of 0.5-18.9 g/h, the glucose concentration in the feed was varied from 2.5 to 10 gCOD/L under conditions where the HRT varied from 1 to 10 h (30 degrees C, pH=5.5). Decreasing the glucose loading rate over this range increased the hydrogen yield from 1.7 to 2.8 mol-H2/mol-glucose. High yields of hydrogen were consistent with a high molar acetate:butyrate ratio of 1.08:1 as more hydrogen is produced with acetate as a product (4 mol-H2/mol-acetate) than with butyrate (2 mol-H2/mol-butyrate). It was thought that the decrease in yield with organic loading rate resulted from an overall decreased rate of hydrogen production. As the rate of gas production is reduced, H2 supersaturation in the liquid phase is likely reduced, relieving inhibition due to H2. Flocculation was also an important factor in the performance of the reactor. At the 5-10 gCOD/L influent glucose concentrations, substantial flocculation was observed particularly as the feeding rate was increased due to a reduction in the HRT from 10 to 2.5 h. At the HRT of 2.5 h, biomass concentrations reached as much as 25 g/L. The flocculant nature of the biomass allowed reactor operation at low HRTs with steady H2 production and >90% glucose removal. However, when the HRT was reduced to 1 h at a glucose feed concentration of 2.5 gCOD/L, there was little flocculation evident resulting in wash-out of the culture. These results suggest that hydrogen yields will be optimized for more dilute feeds and lower organic loading rates than have typically been used in biohydrogen reactor studies.     

Cryogenic laser induced U(VI) fluorescence studies of a U(VI) substituted natural calcite: implications to U(VI) speciation in contaminated Hanford sediments

Time-resolved laser-induced fluorescence spectroscopy (TRLFS) and imaging spectromicroscopy (TRLFISM) were used to examine the chemical speciation of uranyl in contaminated subsurface sediments from the U.S. Department of Energy (U.S. DOE) Hanford Site, Washington. Spectroscopic measurements for contaminant U(VI) were compared to those from a natural, uranyl-bearing calcite (NUC) that had been found via X-ray absorption spectroscopy (XAS) to include uranyl in the same coordination environment as calcium. Spectral deconvolution of TRLFS measurements on the NUC revealed the unexpected presence of two distinct chemical environments consistent with published spectra of U(VI)-substituted synthetic calcite and aragonite. Apparently, some U(VI) substitution sites in calcite distorted to exhibit a local, more energetically favorable aragonite structure. TRLFS measurements of the Hanford sediments NP4-1 and NP1-6 were similar to the NUC in terms of peak positions and intensity, despite a small CaCO3 content (1.0 to 3.2 mass %). Spectral deconvolution of the sediments revealed the presence of U(VI) in calcite and aragonite structural environments. A third, unidentified U(VI) species was also present in the NP1-6 sediment. TRLFISM measurements at multiple locations in the different sediments displayed only minor variation, indicating a uniform speciation pattern. Collectively, the measurements implied that waste U(VI), long-resident beneath the sampled disposal pond (32 y), had coprecipitated within carbonates. These findings have major implications for the solubility and fate of contaminant U(VI).