Capillary electrophoretic separation of serum proteins of workers occupationally exposed to heavy metals

Metallurgy processes are associated with many hazardous and toxic factors, including heavy metals. Exposure to heavy metals can cause damage to different organs, which can be observed through variation in the concentration of proteins in serum. The aim of this study was to evaluate the changes in a serum protein profile of copper smelters exposed to As, Cd and Pb ions, and xenobiotics present in tobacco smoke. A 2.3-fold higher Pb concentration in the blood and a 2.8-fold greater As concentration have been observed in the urine of non-smoking smelters compared to a control group. In the blood of smoking smelters, Cd concentration was 2-fold higher than in non-smoking ones. Serum proteins were separated by capillary electrophoresis, and in the group of non-smoking smelters, a higher amount of α₁-globulins was observed. In the group of smoking smelters, fewer α₁-globulins were noted. Furthermore, a greater amount of α₂-globulins in the serum of smoking and non-smoking workers in relation to the control group was revealed. A positive correlation between the concentration of Cd in the blood and the content of a fraction containing α₁- and α₂-globulins was revealed. Urine Cd concentration was found to be negatively associated with the α₁- and α₂-globulins fraction. Observed abnormalities in the proteins profiles of smelters can be important markers when assessing exposure to heavy metals and in the early diagnosis of diseases caused by them.     

Thermal And Structural Characterization of Commercial α-, β-, and γ-Cyclodextrins

α-, β-, and γ-cyclodextrins (CDs) marketed by five different companies were characterized from the thermal and structural point of view. Three αCD samples showed two-step DSC dehydration profiles and their XRD patterns were characteristic for αCD⋅6H₂O form I, whereas one brand with an apparent three-step DSC dehydration behaviour was a mixture of αCD⋅6H₂O form I and anhydrous αCD. The differences in the DSC profiles after dehydration and EGA onset decomposition temperatures recorded for the five βCD brands were attributed to different manufacturing and purification processes. The five γCDs brands showed a common thermal behaviour and very similar XRD patterns. The patterns did not match the idealized pattern of γCD⋅14.1H₂O, indicating the occurrence of two different hydrated crystal structures. This revised version was published online in August 2006 with corrections to the Cover Date.     

Practical aspects of splitless injection of semivolatile compounds in fast gas chromatography

Possibilities and practical aspects of implementation of splitless injection of larger volumes for fast GC purposes utilizing narrow-bore column, hydrogen as carrier gas, fast temperature programming under programmed flow conditions and commercial instrumentation were searched. As a model sample semivolatile compounds of a broad range of volatility and polarity (7 n-alkanes and 19 pesticides) were chosen. Peak shapes, peak broadening and peak areas and its repeatability were evaluated under various experimental set-ups (liner/injection technique combinations). Various factors, such as liner design, injection technique, retention gap length, compound volatility and polarity, the solvent used, initial oven temperature influenced compound focusation and/or maximal injection volume. Combination of analytical column (CP-Sil 13 CB 25 m long, 0.15 mm i.d., film thickness 0.4 microm) with normal-bore retention gap (1 m long, 0.32 mm i.d.) allowed maximal injection volume 8 microl for 4 mm i.d. liner used without any peak distortion when solvent recondensation in the retention gap was employed.     

A differential microcalorimetric study of whey proteins and their behaviour in oil-in-water emulsions

Oil-in-water emulsions (20% soya oil, 1% protein) have been prepared containing lysozyme or isolates of -lactalbumin and β-lactoglobulin from whey protein. The structural characteristics of these proteins adsorbed at an oil/water interface were determined by following their thermal transitions using differential scanning microcalorimetry. Thermograms of the proteins in the adsorbed state differed markedly from the corresponding transitions seen for the proteins in solution. This suggests that the proteins underwent substantial changes in secondary and tertiary structure upon adsorption. In general, for all the proteins studied, a net decrease in the total energy absorbed during denaturation was found when the proteins were in an adsorbed state. Both lysozyme and -lactalbumin were in part “surface denatured”, and they showed a certain degree of reversibility between solution and the adsorbed state. β-Lactoglobulin showed the highest degree of denaturation upon adsorption and the conformational change was irreversible.     

Molecular Hybridization Strategy on the Design,Synthesis, and Structural Characterization of Ferrocene-N-acyl Hydrazones as Immunomodulatory Agents

Immunomodulatory agents are widely used for the treatment of immune-mediated diseases, but the range of side effects of the available drugs makes necessary the search for new immunomodulatory drugs. Here, we investigated the immunomodulatory activity of new ferrocenyl-N-acyl hydrazones derivatives (SintMed(141–156). The evaluated N-acyl hydrazones did not show cytotoxicity at the tested concentrations, presenting CC₅₀ values greater than 50 µM. In addition, all ferrocenyl-N-acyl hydrazones modulated nitrite production in immortalized macrophages, showing inhibition values between 14.4% and 74.2%. By presenting a better activity profile, the ferrocenyl-N-acyl hydrazones SintMed149 and SintMed150 also had their cytotoxicity and anti-inflammatory effect evaluated in cultures of peritoneal macrophages. The molecules were not cytotoxic at any of the concentrations tested in peritoneal macrophages and were able to significantly reduce (p < 0.05) the production of nitrite, TNF-α, and IL-1β. Interestingly, both molecules significantly reduced the production of IL-2 and IFN-γ in cultured splenocytes activated with concanavalin A. Moreover, SintMed150 did not show signs of acute toxicity in animals treated with 50 or 100 mg/kg. Finally, we observed that ferrocenyl-N-acyl hydrazone SintMed150 at 100 mg/kg reduced the migration of neutrophils (44.6%) in an acute peritonitis model and increased animal survival by 20% in an LPS-induced endotoxic shock model. These findings suggest that such compounds have therapeutic potential to be used to treat diseases of inflammatory origin.     

A peptidomic approach for monitoring and characterising peptide cyanotoxins produced in Italian lakes by matrix-assisted laser desorption/ionisation and quadrupole time-of-flight mass spectrometry

In recent years, the occurrence of cyanobacterial blooms in eutrophic freshwaters has been described all over the world, including most European countries. Blooms of cyanobacteria may produce mixtures of toxic secondary metabolites, called cyanotoxins. Among these, the most studied are microcystins, a group of cyclic heptapeptides, because of their potent hepatotoxicity and activity as tumour promoters. Other peptide cyanotoxins have been described whose structure and toxicity have not been thoroughly studied. Herein we present a peptidomic approach aimed to characterise and quantify the peptide cyanotoxins produced in two Italian lakes, Averno and Albano. The procedure was based on matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry mass spectrometry (MALDI-TOF-MS) analysis for rapid detection and profiling of the peptide mixture complexity, combined with liquid chromatography/electrospray ionisation quadrupole time-of- flight tandem mass spectrometry (LC/ESI-Q-TOF-MS/MS) which provided unambiguous structural identification of the main compounds, as well as accurate quantitative analysis of microcystins. In the case of Lake Averno, a novel variant of microcystin-RR and two novel anabaenopeptin variants (Anabaenopeptins B(1) and Anabaenopeptin F(1)), presenting homoarginine in place of the commonly found arginine, were detected and characterised. In Lake Albano, the peculiar peptide patterns in different years were compared, as an example of the potentiality of the peptidomic approach for fast screening analysis, prior to fine structural analysis and determination of cyanotoxins, which included six novel aeruginosin variants. This approach allows for wide range monitoring of cyanobacteria blooms, and to collect data for evaluating possible health risks to consumers, through the panel of the compounds produced along different years.     

A Coumarin Triflate Reagent Enables One-Step Synthesis of Photo-Caged Lipid Metabolites for Studying Cell Signaling

Photorelease of caged compounds is among the most powerful experimental approaches for studying cellular functions on fast timescales. However, its full potential has yet to be exploited, as the number of caged small molecules available for cell biological studies has been limited by synthetic challenges. Addressing this problem, a straightforward, one-step procedure for efficiently synthesizing caged compounds was developed. An in situ generated benzylic coumarin triflate reagent was used to specifically functionalize carboxylate and phosphate moieties in the presence of free hydroxy groups, generating various caged lipid metabolites, including a number of GPCR ligands. By combining the photo-caged ligands with the respective receptors, an easily implementable experimental platform for the optical control and analysis of GPCR-mediated signal transduction in living cells was developed. Ultimately, the described synthetic strategy allows rapid generation of photo-caged small molecules and thus greatly facilitates the analysis of their biological roles in live cell microscopy assays.     

XANES speciation of mercury in three mining districts – Almadén,Asturias (Spain), Idria (Slovenia)

The mobility, bioavailability and toxicity of mercury in the environment strongly depend on the chemical species in which it is present in soil, sediments, water or air. In mining districts, differences in mobility and bioavailability of mercury mainly arise from the different type of mineralization and ore processing. In this work, synchrotron‐based X‐ray absorption near‐edge spectroscopy (XANES) has been taken advantage of to study the speciation of mercury in geological samples from three of the largest European mercury mining districts: Almadén (Spain), Idria (Slovenia) and Asturias (Spain). XANES has been complemented with a single extraction protocol for the determination of Hg mobility. Ore, calcines, dump material, soil, sediment and suspended particles from the three sites have been considered in the study. In the three sites, rather insoluble sulfide compounds (cinnabar and metacinnabar) were found to predominate. Minor amounts of more soluble mercury compounds (chlorides and sulfates) were also identified in some samples. Single extraction procedures have put forward a strong dependence of the mobility with the concentration of chlorides and sulfates. Differences in efficiency of roasting furnaces from the three sites have been found.     

Interaction of Naproxen with Crystalline and Amorphous Methylated β-Cyclodextrin in the Liquid and Solid State

Abstract

Interactions of naproxen (NAP) with amorphous, randomly methylated β-cyclodextrin at a degree of substitution per anhydroglucose unit of 1.8 (RAMEB) and with crystalline heptakis-(2,6-di-O-methyl)-β-cyclodextrin (DIMEB) were studied in aqueous solution and in the solid state using, respectively, phase-solubility analysis (at 25 °C, 37 °C and 47 °C) and differential scanning calorimetry (DSC) supported by X-ray powder diffractometry. RAMEB and DIMEB displayed similar solubilizing and complexing abilities towards NAP, suggesting analogous inclusion modes of the drug in the host cavity in aqueous solution. Differences were instead observed in interactions in the solid state, where the amorphizing capacity of RAMEB toward NAP (evaluated by DSC) was about twice that of DIMEB at each drug-to-carrier ratio. Assuming that inclusion complexation is also involved in solid-state interactions, molecular modelling accounted for the experimental results in terms of structural features of DIMEB, i.e. the particular inwards orientation of O-6-C-8 groups of three alternate glucoses on the primary hydroxyl side which hampers a deep penetration of NAP in the DIMEB cavity in the solid state. On the contrary, no obstruction of the cavity apparently occurs with RAMEB due its noncrystalline state. The aqueous dissolution rate of NAP from NAP-RAMEB and NAP-DIMEB blends containing 0.59, 0.73, 0.85, and 0.92 mass fraction of carrier linearly increased at decreasing drug-to-carrier ratios. The improvement was 5 to 20 times (from powders) and 50 to 200 times (from discs) the dissolution rate of NAP alone for both carrier. Therefore the choice of the amorphous RAMEB in pharmaceutical formulations can be recommended mainly for economic reasons, though the anhydrous and non-hygroscopic nature of crystalline DIMEB might be of particular advantage in case of moisture sensitive formulations.