Glycoprofiling as a novel tool in serological assays of systemic sclerosis: A comparative study with three bioanalytical methods

Systemic sclerosis (SSc) is an autoimmune disease seriously affecting patient’s quality of life. The heterogeneity of the disease also means that identification and subsequent validation of biomarkers of the disease is quite challenging. A fully validated single biomarker for diagnosis, prognosis, disease activity and assessment of response to therapy is not yet available. The main aim of this study was to apply an alternative assay protocol to the immunoassay-based analysis of this disease by employment of sialic acid recognizing lectin Sambucus nigra agglutinin (SNA) to glycoprofile serum samples. To our best knowledge this is the first study describing direct lectin-based glycoprofiling of serum SSc samples. Three different analytical methods for glycoprofiling of serum samples relying on application of lectins are compared here from a bioanalytical point of view including traditional ELISA-like lectin-based method (ELLA), novel fluorescent lectin microarrays and ultrasensitive impedimetric lectin biosensors. Results obtained by all three bioanalytical methods consistently showed differences in the level of sialic acid present on glycoproteins, when serum from healthy people was compared to the one from patients having SSc. Thus, analysis of sialic acid content in human serum could be of a diagnostic value for future detection of SSc, but further work is needed to enhance selectivity of assays for example by glycoprofiling of a fraction of human serum enriched in antibodies for individual diagnostics.     

The activity and thermal stability of RhOx/CeO2 nanocomposites prepared by radio-frequency plasma sputtering

The model RhO_x/CeO₂ systems were prepared by radio-frequency (RF) plasma sputtering of Rh electrode in O₂ or Ar/O₂ atmosphere. Thermal stability of the systems and their reaction probability towards CO oxidation were studied by X-ray photoelectron spectroscopy. It was shown that the small oxidized Rh nanoparticles on the CeO₂ surface (RhO_x/CeO₂) obtained by RF sputtering in O₂ have spectroscopic characteristics close to those of Rh³⁺ ions highly dispersed in ceria lattice. The RhO_x/CeO₂ system remains stable upon heating in vacuum at 450°C and shows reactivity towards CO oxidation at T > 200°C. RF sputtering in Ar/O₂ atmosphere results in the formation of larger rhodium nanoparticles that are close to Rh₂O₃ oxide. The Rh₂O₃/CeO₂ system demonstrates lower activity in CO oxidation and cannot be reduced at a temperature below 300°C.     

Photoprotective properties of Prunella vulgaris and rosmarinic acid on human keratinocytes

UVA radiation provokes the generation of reactive oxygen species (ROS), which induce oxidative stress in the exposed cells leading to extensive cellular damage and cell death either by apoptosis or necrosis. One approach to protecting human skin against the harmful effects of UV radiation is by using herbal compounds as photoprotectants. This study evaluated the protective effects of Prunella vulgaris L. (Labiatae) and its main phenolic acid component, rosmarinic acid (RA), against UVA-induced changes in a human keratinocyte cell line (HaCaT). Human keratinocytes exposed to UVA (10-30 J/cm(2)) were treated with an extract of P. vulgaris (1-75 mg/l) or RA (0.9-18 mg/l) for 4h. P. vulgaris and RA exhibited ability to reduce the UVA-caused decrease in a cell viability monitored by neutral red retention and by LDH release into medium. The P. vulgaris extract and RA significantly suppressed UVA-induced ROS production, which manifests as a decrease in intracellular lipid peroxidation, elevation of ATP and reduced glutathione. Post-treatment with P. vulgaris extract and RA also significantly reduced DNA damage. In addition, UVA-induced activation of caspase-3 was inhibited by treatment with P. vulgaris and RA. The P. vulgaris extract and RA demonstrated a concentration-dependent photoprotection (maximum at 25-50 mg/l and 9 mg/l, respectively). These results suggest that P. vulgaris and RA, used in skin care cosmetics, may offer protection against UVA-induced oxidative stress and may be beneficial as a supplement in photoprotective dermatological preparations.     

Asymmetric synthesis of (S,R)- and (R,R)-methiin stereoisomers

Abstract

An asymmetric synthesis of (+)- and (–)-methiine (S-methyl-(R)-cysteine sulfoxide) diastereomers has been developed. These natural sulfur compounds were isolated from a variety of Brassica vegetables. As the starting compound, (R)-cysteine was used, which was methylated to form (R)-S-methylcysteine. Then the oxidation of S-methylcysteine with tert-butyl hydroperoxide catalyzed by the chiral tetra(isopropylate)titanium/(S)- or (R)-Binol complex led to the formation of (1?R,2S)-(+)- or (1?R,2R)-(–)-methiin stereomers.     

Glycan analysis: scope and limitations of different techniques—a case for integrated use of LC-MS(/MS) and NMR techniques

The structure of glycans from glycoproteins is highly relevant for their function. We tightly integrate liquid chromatography–mass spectrometry (LC-MS), MS/MS, and nuclear magnetic resonance (NMR) data to achieve a complete characterization of even isobaric glycans differing in only one linkage position or in the substitution in one branch. As example, we analyzed ten desialylated underivatized glycans from bovine fibrinogen. The molecules were separated on a PGC column, and LC-MS data allowed an assignment of the compositions of the glycans. MS/MS data of the same glycans allowed elucidation of sequence and to some extent of branching and linkage. All MS/MS fragmentation methods led to multiple dissociations, resulting in several cases in ambiguous data. The MS/MS data were interpreted both by scientists and automatically by software, and the differential results are compared. Additional data from a tight integration of LC-MS and NMR data resulted in a complete structural characterization of the glycans. The acquisition of simple 1D ¹H NMR data led—in combination with LC-MS and MS/MS data—to an unambiguous assignment of the isobaric glycans. Compounds that were not separated in the chromatography could easily be assigned structurally by applying the 3D cross-correlation (3DCC) technology to arrive at NMR spectra of the pure components—without actually separating them. By applying LC-MS, MS/MS, 1D ¹H NMR, and 3DCC together, one can assign glycan structures from glycoconjugates with high confidence affording only 200 pmol of glycan material.     

Graphane Nanostripes

Graphane, the hydrogenated counterpart of graphene, was shown to exhibit properties such as tunable band gaps through varied degrees of hydrogenation, fluorescence, or ferromagnetism. Graphane nanostripe properties have also been theoretically predicted. Herein, we show that graphane nanostripes can be prepared by opening carbon nanotubes using Birch reduction in liquid ammonia utilizing potassium as a reducing agent and water as a proton donor. The prepared graphane nanostripes exhibit several exceptional properties when coupled with trace metal dopants. The interplay of metallic nanoparticles and defects lead to a spin polarization and induction of ferromagnetic moment, as well as to enhanced electrocatalytic properties in the hydrogen evolution reaction when compared to non‐hydrogenated carbon nanotubes.     

Ferrite supports for isolation of DNA from complex samples and polymerase chain reaction amplification

The influence of cobalt ferrite particles, with non-modified or modified surface, on the course of polymerase chain reaction (PCR) was investigated. DNA isolated from bacterial cells of Bifidobacterium bifidum was used in PCR evaluation of magnetic microspheres. The presence of cobalt ferrite particles inhibits PCR amplification. The effect is not dependent on the functional groups of the modifying reagents used (none, amino, carboxyl). Amplification was improved after the magnetic separation of magnetic particles. Proposed indirect method enabled verification of the suitability of designed particles for their application in PCR assays. Magnetic particles coated with alginic acid under high PEG and sodium chloride concentration were used for the isolation of PCR-ready bacterial DNA from various dairy products. DNA was isolated from crude bacterial cell lysates without phenol extraction of samples. Bifidobacterium and Lactobacillus DNAs were identified in dairy products using PCR.     

Towards Asymmetrical Methylene Blue Analogues: Synthesis and Reactivity of 3-N′-Arylaminophenothiazines

The search for new ways to obtain analogues of the well-known Methylene Blue dye is an important synthetic task. Herein, we proposed and developed an approach to the synthesis of 3-N′-arylaminophenothiazines and asymmetrical 3,7-di(N′-arylamino)phenothiazines. This approach included the optimization of synthetic strategy by quantification analysis of the positive charge distribution in the cation of 3-N′-arylaminophenothiazine derivative. The obtained experimental data are confirmed by DFT studies. Two synthetic routes for asymmetrical phenothiazine diarylamino derivatives were suggested and verified. The developed convenient and versatile synthetic approach makes it easy to obtain aromatic Methylene Blue isostructural analogues with various substituents. As a result, a series of novel 3-N′-arylaminophenothiazines and asymmetrical 3,7-di(N′-arylamino)phenothiazines containing ester, tert-butoxycarbonyl, sulfonic acid, hydroxyl and amine groups were obtained in high yields.     

Indomethacin: The Interplay between Structural Relaxation,Viscous Flow and Crystal Growth

Non-isothermal differential scanning calorimetry (DSC) was used to study the influences of particle size (d_aver) and heating rate (q⁺) on the structural relaxation, crystal growth and decomposition kinetics of amorphous indomethacin. The structural relaxation and decomposition processes exhibited d_aver-independent kinetics, with the q⁺ dependences based on the apparent activation energies of 342 and 106 kJ·mol⁻¹, respectively. The DSC-measured crystal growth kinetics played a dominant role in the nucleation throughout the total macroscopic amorphous-to-crystalline transformation: the change from the zero-order to the autocatalytic mechanism with increasing q⁺, the significant alteration of kinetics, with the storage below the glass transition temperature, and the accelerated crystallization due to mechanically induced defects. Whereas slow q⁺ led to the formation of the thermodynamically stable γ polymorph, fast q⁺ produced a significant amount of the metastable α polymorph. Mutual correlations between the macroscopic and microscopic crystal growth processes, and between the viscous flow and structural relaxation motions, were discussed based on the values of the corresponding activation energies. Notably, this approach helped us to distinguish between particular crystal growth modes in the case of the powdered indomethacin materials. Ediger’s decoupling parameter was used to quantify the relationship between the viscosity and crystal growth. The link between the cooperativity of structural domains, parameters of the Tool-Narayanaswamy-Moynihan relaxation model and microscopic crystal growth was proposed.