Nanomaterials as Pseudocatalysts in the Construction of Electrochemical Nonenzymatic Sensors for Healthcare: A Review

Enzymes, primarily different types of oxidases and most commonly peroxidase, are often used in the construction of biosensors. Enzymatic biosensors, due to their small size, easy to handle construction, accuracy and specificity, are powerful healthcare tools commonly used for the diagnosis of diseases for more than 20 years. Unfortunately, the loss of enzymatic activity during the immobilization of enzymes into biosensors has been a recent major problem. Hence, nonenzymatic electrochemical sensors based on organic and inorganic nanostructures have gained great attention in the last few years. In this short review, different types of nanostructures and nanocomposites and their practical applications in the construction of nonenzymatic electrochemical sensors in healthcare and diagnosis are described and summarized.     

Structure Analysis of Montmorillonite Intercalated with Cetylpyridinium and Cetyltrimethylammonium: Molecular Simulations and XRD Analysis

Molecular mechanics and molecular dynamics simulations combined with X-ray powder diffraction were used in structure investigation of montmorillonite intercalated with cetylpyridinium (CP) and cethyltrimethylammonium (CTA) cations. Molecular modeling revealed the interlayer structure and differences in intercalation behavior of CP and CTA cations in montmorillonite. The experimental and calculated values of basal spacing were in good agreement for both intercalates: in the case of CP-montmorillonite d(exp)=20.59 ?, d(calc)=20.60 ?; for CTA-montmorillonite d(exp)=18.00 ? and d(calc)=18.10 ?. CTA-montmorillonite exhibits significantly higher total sublimation energy and higher host-guest interaction energy than the CP-montmorillonite. The main difference between both intercalates is in charge distribution on the host layers and guest species. The charge transfer from the guest species to the host layer is higher in CTA-montmorillonite than in CP-montmorillonite, and consequently the charge polarization between the host and guest layers is much higher in CTA-montmorillonite. This leads to much stronger host-guest electrostatic interaction in the case of CTA-montmorillonite. Copyright 2001 Academic Press.     

Differential Invariants of Immersions of Manifolds with Metric Fields

The problem of finding all r^th order differential invariants of immersions of manifolds with metric fields, with values in a left (G¹_m×G¹_n)-manifold is formulated. For obtaining the basis of higher order differential invariants the orbit reduction method is used. As a new result it appears that r^th order differential invariants depending on an immersion f:M N of smooth manifolds M and N and metric fields on them can be factorized through metrics, curvature tensors and their covariant differentials up to the order (r–2), and covariant differentials of the tangent mapping Tf up to the order r. The concept of a covariant differential of Tf is also introduced in this paper. The obtained results are geometrically interpreted as well.This research is supported by grants GAR 201/03/0512 and MSM 143100006.     

Comparison of two different solvents employed for pressurised fluid extraction of stevioside from <Emphasis Type="Italic">Stevia rebaudiana</Emphasis>: methanol versus water

Pressurised fluid extraction using water or methanol was employed for the extraction of stevioside from Stevia rebaudiana Bertoni. The extraction method was optimised in terms of temperature and duration of the static or the dynamic step. Extracts were analysed by liquid chromatography followed by UV and mass-spectrometric (MS) detections. Thermal degradation of stevioside was the same in both solvents within the range 70–160 °C. Methanol showed better extraction ability for isolation of stevioside from Stevia rebaudiana leaves than water within the range 110–160 °C. However, water represents the green alternative to methanol. The limit of detection of stevioside in the extract analysed was 30 ng for UV detection and 2 ng for MS detection.     

Synthesis and Cytotoxic and Antiviral Activity Profiling of All-Four Isomeric Series of Pyrido-Fused 7-Deazapurine Ribonucleosides

All four isomeric series of novel 4-substituted pyrido-fused 7-deazapurine ribonucleosides possessing the pyridine nitrogen atom at different positions were designed and synthesized. The total synthesis of each isomeric fused heterocycle through multistep heterocyclization was followed by glycosylation and derivatization at position 4 by cross-coupling reactions or nucleophilic substitutions. All compounds were tested for cytostatic and antiviral activity. The most active were pyrido[4′,3′:4,5]pyrimidine nucleosides bearing MeO, NH₂, MeS, or CH₃ groups at position 4, which showed submicromolar cytotoxic effects and good selectivity for cancer cells. The mechanism involved activation by phosphorylation and incorporation to DNA where the presence of the modified ribonucleosides causes double-strand breaks and apoptosis.     

Capillary electrophoresis–mass spectrometry as a tool for the noninvasive target metabolomic analysis of underivatized amino acids for evaluating embryo viability in assisted reproduction

Monitoring metabolite uptake and excretion in the culture medium is a noninvasive technique that is used for the metabolic study of cleaving embryos after in vitro fertilization. Low sample consumption, the versatility of the detection, and optimal sensitivity and selectivity are essential elements for extracellular metabolome analyses, and can be conveniently achieved by combining CE with mass spectrometric detection. This paper reports a method for amino acid determination in a limited volume sample (8 μL) of spent culture media collected after the cultivation of in vitro fertilized embryos. Special attention was focused on the sample preparation procedure. The sample was processed with acetonitrile, which facilitates online sample preconcentration via field-amplified sample stacking, and undesired sample evaporation was significantly reduced by the simultaneous addition of dimethyl sulfoxide. Key parameters that affected electrophoretic separation and mass spectrometric detection were investigated, including the type of buffers and organic solvent, optimization of their concentrations, and finally the settings for their ionization. The separation and quantification of 19 amino acids were achieved using 15% acetic acid as the background electrolyte with a sheath liquid consisting of an equimolar mixture of methanol and water. The applicability of the optimized system was demonstrated by determining the amino acid profile in 40 samples of spent cultivation medium in this pilot study. This developed method also has great potential for amino acid analyses in minute sample volumes of other biological matrices.     

In situ derivatization–liquid liquid extraction as a sample preparation strategy for the determination of urinary biomarker prolyl‐4‐hydroxyproline by liquid chromatography–tandem mass spectrometry

Polar analytes that possess protic functional groups have often been treated with alkyl chloroformates to decrease their polarity and increase their volatility prior to gas chromatography–mass spectrometry analysis. This derivatization reaction has two distinct advantages. It proceeds smoothly in aqueous media, and the desired reaction products are efficiently separated from interfering ionic components by their extraction into a water‐immiscible organic phase. In the present work, the derivatization–liquid liquid sample preparation was examined in detail for analysis of a potential urinary dipeptide biomarker l ‐prolyl‐4‐l ‐hydroxyproline (PHP) by downstream liquid chromatography coupled to electrospray mass spectrometry. PHP was treated with a series of alkyl and fluoroalkyl chloroformates in aqueous media, and the detected reaction products were investigated. Smooth conversion of PHP into the N‐isobutyloxycarbonyl isobutyl ester was accomplished by the coupled action of isobutanol, isobutyl chloroformate and the pyridine catalyst. This derivative afforded a highest detector response from all the derivatized forms examined, including the nonderivatized PHP. A simple isocratic elution on a common RP‐C18 HPLC column coupled with tandem mass spectrometry, and use of the synthesized heptadeuterated analog (D7‐PHP) as an internal standard, enabled validation of the method and determination of PHP in human urine in less than 5 min. The in situ derivatization–liquid liquid extraction has thus been demonstrated to be a useful sample preparation strategy for the analysis of polar metabolites by liquid chromatography–tandem mass spectrometry in the complex urine matrix. Copyright © 2012 John Wiley & Sons, Ltd.     

Synthesis of 2′-deoxy-2′-fluororibo- and 2′-deoxy-2′,2′-difluororibonucleosides derived from 6-(het)aryl-7-deazapurines

A series of novel sugar-modified derivatives of cytostatic 6-hetaryl-7-deazapurine ribonucleosides (2′-deoxy-2′-fluororibo- and 2′-deoxy-2′,2-difluororibonucleosides) bearing an aryl or hetaryl group in position 6, was prepared and screened for biological activity. The fluororibo derivatives were prepared by aqueous palladium catalyzed cross-coupling reactions of the corresponding 6-chloro-7-deazapurine 2′-deoxy-2′-fluororibonucleoside 11 with (het)arylboronic acids. The key intermediate 11 was prepared by a six-step sequence from the corresponding arabinonucleoside by selective protection of 3′- and 5′-hydroxyls by acid-labile groups followed by stereoselective S_N2 fluorination and deprotection. The difluororibo-series was prepared by non-stereoselective glycosidation of 6-chloro-7-deazapurine with benzoyl-protected 2-deoxy-2,2-difluoro-d-erythro-pentofuranosyl-1-mesylate followed by cross-couplings, separation of anomers and deprotection. The title nucleosides did not show considerable cytostatic or antiviral activity.     

Determination of insoluble avian eggshell matrix proteins

The organic components of bones and other mineralized tissues have a high impact on the organization and deposition of calcium, and consequently influence the mechanical properties of those tissues. The extractable proteins of avian eggshells have been studied extensively and many of them have been identified; insoluble (non-extractable) proteins have been sparsely studied, however. In the work discussed in this paper we studied EDTA-insoluble proteins by gradual decalcification of eggshell with EDTA. The insoluble proteinaceous films were chemically treated with cyanogen bromide and the mixtures of large fragments obtained were gradually precipitated with salt. The separated fractions were digested with trypsin and analyzed by HPLC–MS–MS (ion trap mass spectrometer). Analysis of the entire eggshell matrix (without precipitation steps) only enabled 6 proteins to be determined (ovocalyxins 32 and 36, ovocleidin 17 and 116, clusterin, and ovalbumin). Pretreatment of the individual eggshell layers and gradual precipitation with salt markedly increased the number of proteins identified – 28 proteins were determined. We identified for the first time collagens I (two chains) and III in the eggshell matrix, and Kunitz-like protease inhibitor as a major shell matrix protein. Besides the above mentioned proteins we can also mention EDIL3, fibronectin, sulfhydryl oxidase, tubulin alpha 1, lysozyme, Dickkopf-related protein 3, keratins, and ovotransferrin. The relative abundances of proteins in all eggshell layers were determined using the exponentially modified protein abundance index (emPAI). In the cuticle layer seven proteins were identified, whereas 16 proteins were described in the palisade layer and 23 in the mammillary layer.