RNA degradation in cell extracts: real-time monitoring by fluorescence resonance energy transfer

Short noncoding RNAs are increasingly recognized as key regulators of essential cellular processes such as RNA interference. A better understanding of the processes by which such RNAs are degraded is necessary to expand our knowledge of these processes and our ability to harness them. To this end we have developed a novel fluorescence resonance energy transfer (FRET) assay to monitor in real-time the degradation kinetics of short RNAs by a purified RNase and S100 cytosolic HeLa cell extract. An unstructured RNA is found to be degraded more rapidly than a stem-loop RNA under all conditions tested except for low concentrations of cell extract, showing that secondary structure confers protection against RNase activity. The assay also allows for the quantitative comparison of inhibitors such as Contrad70 and aurin tricarboxylic acid (ATA). Finally, gel electrophoretic FRET analysis confirms that HeLa cell extract is dominated by 5' to 3' exonucleolytic activity.     

UV resonance Raman spectroscopic monitoring of supramolecular complex formation: peptide recognition in aqueous solution

The formation of a supramolecular complex between a tetrapeptide and an artificial receptor , is monitored at submillimolar concentrations in water by UV resonance Raman spectroscopy. Using 275 nm excitation, we selectively probe the carboxylate binding site (CBS) within the receptor, a moiety which is very efficient in binding the carboxy terminus of peptides in aqueous media. Complexation of the receptor with the tetrapeptide involves the formation of a H-bond enforced ion pair, resulting in significant changes in the corresponding UV resonance Raman spectra. Our qualitative interpretation is based on experimental reference and calculated Raman spectra on model systems. First preliminary calculations show that for a quantitative analysis, also the distinct contributions of multiple CBS conformers must be considered in addition to the H-bond induced changes upon complexation.     

Ultraviolet photolysis of HCHO: absolute HCO quantum yields by direct detection of the HCO radical photoproduct

Absolute quantum yields for the radical (H + HCO) channel of HCHO photolysis, Phi(HCO), have been measured for the tropospherically relevant range of wavelengths (lambda) between 300 and 330 nm. The HCO photoproduct was directly detected by using a custom-built, combined ultra-violet (UV) absorption and cavity ring down (CRD) detection spectrometer. This instrument was previously employed for high-resolution (spectral resolution approximately 0.0035 nm) measurements of absorption cross-sections of HCHO, sigma(HCHO)(lambda), and relative HCO quantum yields. Absolute Phi(HCO) values were measured at seven wavelengths, lambda = 303.70, 305.13, 308.87, 314.31, 320.67, 325.59, and 329.51 nm, using an independent calibration technique based on the simultaneous UV photolysis of HCHO and Cl(2). These Phi(HCO) measurements display greater variability as a function of wavelength than the current NASA-JPL recommendations for Phi(HCO). The absolute Phi(HCO)(lambda) determinations and previously measured sigma(HCHO)(lambda) were used to scale an extensive set of relative HCO yield measurements. The outcome of this procedure is a full suite of data for the product of the absolute radical quantum yield and HCHO absorption cross-section, Phi(HCO)(lambda)sigma(HCHO)(lambda), at wavelengths from 302.6 to 331.0 nm with a wavelength resolution of 0.005 nm. This product of photochemical parameters is combined with high-resolution solar photon flux data to calculate the integrated photolysis rate of HCHO to the radical (H + HCO) channel, J(HCO). Comparison with the latest NASA-JPL recommendations, reported at 1 nm wavelength resolution, suggests an increased J(HCO) of 25% at 0 degrees solar zenith angle (SZA) increasing to 33% at high SZA (80 degrees). The differences in the calculated photolysis rate compared with the current HCHO data arise, in part, from the higher wavelength resolution of the current data set and highlight the importance of using high-resolution spectroscopic techniques to achieve a complete and accurate picture of HCHO photodissociation processes. All experimental Phi(HCO)(lambda)sigma(HCHO)(lambda) data are available for the wavelength range 302.6-331.0 nm (at 294 and 245 K and under 200 Torr of N(2) bath gas) as Supporting Information with wavelength resolutions of 0.005, 0.1, and 1.0 nm. Equivalent data sets of Phi(H(2)+CO)(lambda)sigma(HCHO)(lambda) for the molecular (H(2) + CO) photofragmentation channel, produced using the measured Phi(HCO)(lambda) sigma(HCHO)(tau) values, are also provided at 0.1 and 1.0 nm resolution.     

Ordered bimetallic coordination networks featuring rare earth and silver cations

Three lanthanide complexes of the ditopic ligand 3-cyanopentane-2,4-dionate (acacCN) have been synthesized and structurally characterized. Longer intermolecular contacts result in ninefold coordination of the cation in Ce(acacCN)(3)(H(2)O)(2), whereas mononuclear complexes of the same stoichiometry with coordination number eight are obtained for the smaller Eu(III) and Yb(III) cations. Reaction of these labile compounds with AgPF(6) leads to re-organization of the coordination sphere of the rare earth cations: neutral extended structures are formed in which the peripheric -CN moieties of Ln(acacCN)(4) anions coordinate to silver cations. The initially formed heterometallic networks show additional coordination of water or inclusion of solvent molecules; three different structure types, two of them as isomorphous pairs, have been characterized. In the case of Eu(III) and Yb(III), these solids are instable when stored in their mother liquor and undergo a slow aging process, finally resulting in phase pure stable and solvent-free 3D networks Ln(acacCN)(4)Ag.     

Added-Value Chemicals from Lignin Oxidation

Lignin is the second most abundant component, next to cellulose, in lignocellulosic biomass. Large amounts of this polymer are produced annually in the pulp and paper industries as a coproduct from the cooking process—most of it burned as fuel for energy. Strategies regarding lignin valorization have attracted significant attention over the recent decades due to lignin’s aromatic structure. Oxidative depolymerization allows converting lignin into added-value compounds, as phenolic monomers and/or dicarboxylic acids, which could be an excellent alternative to aromatic petrochemicals. However, the major challenge is to enhance the reactivity and selectivity of the lignin structure towards depolymerization and prevent condensation reactions. This review includes a comprehensive overview of the main contributions of lignin valorization through oxidative depolymerization to produce added-value compounds (vanillin and syringaldehyde) that have been developed over the recent decades in the LSRE group. An evaluation of the valuable products obtained from oxidation in an alkaline medium with oxygen of lignins and liquors from different sources and delignification processes is also provided. A review of C₄ dicarboxylic acids obtained from lignin oxidation is also included, emphasizing catalytic conversion by O₂ or H₂O₂ oxidation.     

Microhardness of a dental restorative composite resin containing nanoparticles polymerized with argon ion laser

The objective of this study was to compare the microhardness of two resin composites (microhybrid and nanoparticles). Light activation was performed with argon ion laser 1.56 J (L) and halogen light 2.6 J (H) was used as control. Measurements were taken on the irradiated surfaces and those opposite them, at thicknesses of 1, 2 and 3 mm. To evaluate the quality of polymerization, the percentages of maximum hardness were calculated (PMH). For statistical analysis the ANOVA and Tukey tests were used (p ≤ 0.05). To microhybrid was shown that the hardness with laser was inferior to the hardness achieved with halogen light, for both the 1 mm and 2 mm. The nanoparticles polymerized with laser, presented lower hardness even on the irradiated surface, than the same surface light activated with halogen light. The microhybrid attained a minimum PMH of 80% up to the thickness of 2 mm with halogen light, and with laser, only up to 1 mm. The nanoparticles attained a minimum PMH of 80% up to 3 mm thickness with halogen light and with laser this minimum was not obtained at any thickness. Based on these results, it could be concluded that light activation with argon ion laser is contra-indicated for the studied nanoparticles.     

Phenol adsorption onto powdered and granular activated carbon, prepared from Eucalyptus wood

Eucalyptus grandis sawdust, a major waste from the growing Uruguayan wood industry, was used in previous work to prepare powdered activated carbon (PAC). In the present work, granular activated carbon (GAC) was prepared by mixing PAC, carboxymethyl cellulose as a binder, and kaolin as reinforcer. Ultimate analysis and surface characterization of GAC and PAC were performed. Phenol adsorption was used as a way to compare the characteristics of different PAC and GAC preparations. Kinetics and isotherms of the different GAC and PAC were performed in a shaking bath at 100 rpm and 298 K. Phenol concentrations were determined by UV spectroscopy. Some kinetics parameters were calculated; from kinetics results, external resistance to mass transfer from the bulk liquid can be neglected as the controlling step. Isotherms were fitted to Langmuir and Freundlich models, and corresponding parameters were determined. Maximum phenol uptakes for all carbons were determined and correlated with carbon characteristics. Thermogravimertic analysis (TGA) determinations were performed in order to study adsorption characteristics and conditions for GAC regeneration after its use. The results showed that phenol is preferentially physisorbed on the carbon of the granules, though some chemisorption was detected. No adsorption was detected in the kaolin-carboxymethyl cellulose mixture.     

Inverting External Asymmetric Induction via Selective Energy Transfer Catalysis: A Strategy to β‐Chiral Phosphonate Antipodes

Enantiodivergent, catalytic reduction of activated alkenes relays stereochemical information encoded in the antipodal chiral catalysts to the pro‐chiral substrate. Although powerful, the strategy remains vulnerable to costs and availability of sourcing both catalyst enantiomers. Herein, a stereodivergent hydrogenation of α,β‐unsaturated phosphonates is disclosed using a single enantiomer of the catalyst. This enables generation of the R‐ or S‐configured β‐chiral phosphonate with equal and opposite selectivity. Enantiodivergence is regulated at the substrate level through the development of a facile E → Z isomerisation. This has been enabled for the first time by selective energy transfer catalysis using anthracene as an inexpensive organic photosensitiser. Synthetically valuable in its own right, this process enables subsequent Rh^I‐mediated stereospecific hydrogenation to generate both enantiomers of the product using only the S‐catalyst (up to 99:1 and 3:97 e.r.). This strategy out‐competes the selectivities observed with the E‐substrate and the R‐catalyst.     

Determination of vitamin E and carotenoid pigments by high performance liquid chromatography in shell of Chionoecetes opilio

This study reports the optimization of a method for the determination of vitamin E and carotenoids in shells of Chionoecetes opilio samples by online HPLC coupled with UV-vis and fluorescence detectors. The carotenoids were determined with diode-array detector (lambda 450 nm) and vitamin E with fluorescence detection (lambda(ex) 288, lambda(em) 331 nm). Two extractions methods were compared, saponification followed by an extraction step and a simple extraction with acetone. The last one was selected because allows to determine all compounds. Linearity, precisions and recoveries achieved for all compounds were satisfactory. Mean concentrations (mg per 100 g dry weight) were; 23.3 for vitamin E, 9.49 for astaxanthin and 0.2 mg for beta-carotene.