Alkylsulfanylphenyl derivatives of cytosine and 7-deazaadenine nucleosides, nucleotides and nucleoside triphosphates: synthesis, polymerase incorporation to DNA and electrochemical study

Aqueous Suzuki–Miyaura cross‐coupling reactions of halogenated nucleosides, nucleotides and nucleoside triphosphates derived from 5‐iodocytosine and 7‐iodo‐7‐deazaadenine with methyl‐, benzyl‐ and tritylsufanylphenylboronic acids gave the corresponding alkylsulfanylphenyl derivatives of nucleosides and nucleotides. The modified nucleoside triphosphates were incorporated into DNA by primer extension by using Vent(exo‐) polymerase. The electrochemical behaviour of the alkylsulfanylphenyl nucleosides indicated formation of compact layers on the electrode. Modified nucleotides and DNA with incorporated benzyl‐ or tritylsulfanylphenyl moieties produced signals in [Co(NH₃)₆]³⁺ ammonium buffer, attributed to the Brdi?ka catalytic response, depending on the negative potential applied. Repeated constant current chronopotentiometric scans in this medium showed increased Brdi?ka catalytic response, which suggests the deprotection of the alkylsulfanyl derivatives to free thiols under the conditions.     

Computer aided measurement of biomechanical characteristics of cadaverous lumbar spines

Special computer-aided equipment was designed for measurement of biomechanical characteristics of lower part of the spine (L1-L5). When the stress is applied, it is necessary to observe the shift of the sample elements, together with measurement of the spine rigidity. This shift is determined with the help of circular targets fitted to the appropriate vertebra. The targets, illuminated by lamp or laser light, are monitored and their digitalized images are scanned by CCD camera is stored as computer media. The two dimensional Fourier transform of the digital optical signal is obtained by the fast Fourier transform algorithm. The period and direction of the interference fringes determine the size and the direction of the sample shift.     

Enhancement of SEM to scanning LEEM

By introducing a cathode lens below or inside the objective lens of a scanning electron microscope, many experiments similar to those typical of the LEEM method can be performed. The conditions for the diffraction of slow electrons are modified by the convergence of the primary beam, and challenges include the necessity of managing the signal species propagating along the optical axis in a direction opposite to that of the primary beam. However, even a simple implementation, providing the integral dark-field signal only, has not only delivered plenty of results in the very low energy range below 50 eV, but the performance in the range of hundreds of eV and units of keV has also been substantially improved. The scanning LEEM method is illustrated using experimental results acquired by additionally employing multichannel detection and detection of transmitted electrons.     

Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode

In this paper, we present an electrochemical DNA–protein interaction assay based on a combination of protein-specific immunoprecipitation at magnetic beads (MBIP) with application of oligonucleotide (ON) probes labeled with an electroactive oxoosmium complex (Os,bipy). We show that double-stranded ONs bearing a dT₂₀ tail labeled with Os,bipy are specifically recognized by the tumor suppressor p53 protein according to the presence or absence of a specific binding site (p53CON) in the double-stranded segment. We demonstrate the applicability of the Os,bipy-labeled probes in titration as well as competition MBIP assays to evaluate p53 relative affinity to various sequence-specific or structurally distinct unlabeled DNA substrates upon modulation of the p53-DNA binding by monoclonal antibodies used for the immunoprecipitation. To detect the p53-bound osmium-labeled probes, we took advantage of a catalytic peak yielded by Os,bipy-modified DNA at the mercury-based electrodes, allowing facile determination of subnanogram quantities of the labeled oligonucleotides. Versatility of the electrochemical MBIP technique and its general applicability in studies of any DNA-binding protein is discussed. Figure

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Amphiphilic conetworks. IV. Poly(methacrylic acid)‐l‐polyisobutylene and poly(acrylic acid)‐l‐polyisobutylene based hydrogels prepared by two‐step polymer procedure. New pH responsive conetworks

This article describes the synthesis and characterization of new amphiphilic polymer conetworks containing hydrophilic poly(methacrylic acid) (PMAA) or poly(acrylic acid) (PAA) and hydrophobic polyisobutylene (PIB) chains. These conetworks were prepared by a two‐step polymer synthesis. In the first step, a cationic copolymer of isobutylene (IB) and 3‐isopropenyl‐α,α‐dimethylbenzyl isocyanate (IDI) was prepared. The isocyanate groups of the IB–IDI random copolymer were subsequently transformed in situ to methacrylate (MA) groups in reaction with 2‐hydroxyethyl methacrylate (HEMA). In the second step, the resulting MA‐multifunctional PIB‐based crosslinker, PIB(MA)_n, with an average functionality of approximately four methacrylic groups per chain, was copolymerized with methacrylic acid (MAA) or acrylic acid (AA) by radical mechanism in tetrahydrofuran giving rise to amphiphilic conetworks containing 31–79 mol % of MAA or 26–36 mol % of AA. The synthesized conetworks were characterized with solid‐state ¹³C‐NMR spectroscopy and differential scanning calorimetry. The amphiphilic nature of the conetworks was proven by swelling in both aqueous media with low and high pH and n‐heptane. The effect of varying pH on the swelling behavior of the synthesized conetworks is presented. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 1284–1291, 2009     

Novel Synthesis of Phosphinates by the Microwave-Assisted Esterification of Phosphinic Acids

1-Hydroxy-3-phospholene oxides (1 and 3) and phenyl-H-phosphinic acid (6) are converted to the corresponding phosphinic esters (2, 4, and 7, respectively) by reaction with simple alcohols on microwave irradiation. Under traditional heating conditions, the esterification does not take place, as in the cases of 1 and 3, or is highly incomplete, as in the case of 6. Steric hindrance in diphenylphosphinic acid prevents efficient microwave-assisted esterification.     

Synthesis and Use of 6-, 7-and 8-Membered P-Heterocycles

The synthesis of new families of P-heterocycles by reductive modifications, ring enlargement and Diels-Alder cycloaddition is described.     

Evaluation of the novel passive sampler for cyanobacterial toxins microcystins under various conditions including field sampling

In the present study, we have evaluated the effectiveness of a passive sampler for polar organic chemicals to accumulate a group of widespread and hazardous tumor-promoting toxins produced in cyanobacterial water blooms—microcystins (MC). The previously optimized configuration of the sampler based on polycarbonate membrane and Oasis HLB sorbent (2.75 mg/cm²) was validated under various exposure scenarios in laboratory and field. Calibration of the passive sampler conducted under variable conditions and concentrations of MC revealed linearity of the sampling up to 4 weeks. The sampling rates of microcystins for two different exposure scenarios were derived (e.g., MC-LR: R _s = 0.017 L/day under static and 0.087 L/d under turbulent conditions). R _s values were further used for calculations of time-weighted average concentrations in natural water. Improved sensitivity and selectivity of the in-house-made sampler was observed in comparison with the commercially available Polar Organic Compound Integrative Sampler (POCIS). Comparisons of grab and passive sampling methods were performed during cyanobacterial water bloom season in the Brno reservoir, Czech Republic in 2008. Data obtained by passive sampling provided a more relevant picture of the situation and enabled better assessment of potential risks. The present study demonstrated that the modification of POCIS is suitable for monitoring of occurrence and retrospective estimations of microcystin water concentrations, especially with respect to the control of drinking water quality.     

High-performance liquid chromatographic determination of the plasma concentration of K-27, a novel oxime-type cholinesterase reactivator

A simple and reliable HPLC method for the determination of the plasma level of K-27, an oxime type antidote of use in organophosphorus poisoning is presented. Separation was carried out by HPLC using an octyl silica stationary phase and a mobile phase consisting of 93% phosphate buffer (pH 2.6) containing octane sulfate sodium salt, and 7% methanol. Quantitative absorbance was monitored at 286 nm. The calibration curve was linear through the range of 1.25-200 microg/mL, that is well beyond the detected plasma level range of K-27. Limit of quantitation was 5 microg/mL. Intra-day and inter-day precisions of the HPLC determinations gave standard deviations as 0.77 and 2.67%, respectively. Following intramuscular administration of 50 micromol (22.31 mg) K-27 in rats, the maximum of K-27 concentration in plasma was reached at about 15 min giving 186 microg/mL and the t(1/2) was 85 min. K-27 displays initial (from 15 trough 120 min) zero order elimination kinetics. Similar results have been found after intraperitoneal administration.     

HPLC analysis of K-48 concentration in plasma

K-48 is a new oxime-type compound to be used as an enzyme reactivator in the treatment of exposure to organophosphorous compounds. Plasma concentration of K-48 can be determined using reversed-phase HPLC. Analysis using octyl silica stationary phase and ultraviolet-absorbance detection is fast and simple. K-48 displays a relatively high dose-normalized area under the curve as compared to pralidoxime, which might be beneficial for an antidote. After i.m. administration of 50 μmol K-48, the time course of the concentration can be approximated by a straight line between 15 and 120 min meaning the elimination follows zero-order kinetics.