S-nitrosylation is emerging as an important signaling mechanism that regulates a broad range of cellular functions. The recognition of Cysteine residues that undergo S-nitrosylation is crucial to elucidate how this modification modulates protein activity. We report here a novel strategy, defined His-tag switch, which allows the purification and identification of S-nitrosylated proteins and the unambiguous localization of the modified cysteine residues by mass spectrometry analysis.
A rapid confirmatory method for determining 12 sulfonamide (SAs) antibacterials in whole milk and eggs is presented. This method is based on the matrix solid-phase dispersion technique with hot water as extractant followed by liquid chromatography (LC)-mass spectrometry (MS). The LC-MS instrument was equipped with an electrospray ion source and a single quadrupole. After 4 mL of a milk sample containing the analytes had been deposited on sand (crystobalite), this material was packed into an extraction cell. SAs were extracted by flowing 4 mL of water through the cell heated at 75 degrees C. With some modifications, this procedure was applied also to eggs. After pH adjustment and filtration, 0.5 mL of the final extracts was then injected into the LC column. MS data acquisition was performed in the positive-ion mode and by monitoring at least three ions for each target compound. The in-source collision-induced dissociation process produced confirmatory ions. At the 50 ng/g level, recovery of the analytes in milk and eggs was 77-92% with relative standard deviations ranging between 1 and 11%. Estimated limits of quantification (S/N = 10) were 1-3 ng/g of SAs in milk and 2-6 ng/g in eggs. With both matrices, attempts to reduce the analysis time by using a short chromatographic run time caused severe ion signal suppression for the early-eluted SAs. This effect was traced to competition effects by polar endogenous coextractives, maybe proteinaceous species, which are eluted in the first part of the chromatographic run. This unwelcome effect was almost completely removed by simply adopting more selective chromatographic conditions.
This paper presents a comprehensive review of European Union (EU) legislation addressing the safety of chemical substances, and possibilities within each piece of legislation for applying grouping and read-across approaches for the assessment of nanomaterials (NMs). Hence, this review considers both the overarching regulation of chemical substances under REACH (Regulation (EC) No 1907/2006 on registration, evaluation, authorization, and restriction of chemicals) and CLP (Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures) and the sector-specific pieces of legislation for cosmetic, plant protection and biocidal products, and legislation addressing food, novel food, and food contact materials. The relevant supporting documents (e.g. guidance documents) regarding each piece of legislation were identified and reviewed, considering the relevant technical and scientific literature. Prospective regulatory needs for implementing grouping in the assessment of NMs were identified, and the question whether each particular piece of legislation permits the use of grouping and read-across to address information gaps was answered.ARTICLE HISTORY
Responsible development of any technology, including nanotechnology, requires protecting workers, the first people to be exposed to the products of the technology. In the case of nanotechnology, this is difficult to achieve because in spite of early evidence raising health and safety concerns, there are uncertainties about hazards and risks. The global response to these concerns has been the issuance by authoritative agencies of precautionary guidance to strictly control exposures to engineered nanomaterials (ENMs). This commentary summarizes discussions at the “Symposium on the Health Protection of Nanomaterial Workers” held in Rome (25 and 26 February 2015). There scientists and practitioners from 11 countries took stock of what is known about hazards and risks resulting from exposure to ENMs, confirmed that uncertainties still exist, and deliberated on what it would take to conduct a global assessment of how well workers are being protected from potentially harmful exposures.
The low molecular weight (LMW) region of the circulatory proteome, thought to contain a rich source of biomarkers, resides in vivo, in a complexed state with larger, highly abundant resident proteins. Consequently, serum fractionation approaches that deplete the high-abundance proteins under native conditions will remove much of the LMW proteome. We describe a new strategy to systematically collect, isolate and enrich the LMW molecules that would be otherwise eliminated during the depletion of high-abundance circulatory proteins based on continuous elution electrophoresis. We employ strong denaturing conditions to disrupt association with the high-abundance carrier proteins followed by fractionation and removal of SDS. Under denaturation, the LMW molecules were effectively stripped from the highly abundant carrier proteins. We then removed the SDS by ion exchange matrix sequestration and concentrated the fractions. The outcome is a series of SDS-free fractions of LMW molecules. The isolated fractions were then analyzed by enzymatic digestion followed by LC-MS/MS analysis. The yield of multiple peptide hits as well as the total number of identifications significantly increased (50%) compared to unfractionated serum. The method yielded a 30% higher number of low-abundance serum proteins compared to direct sequencing of unfractionated serum.
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