Accelerated on-column lysine derivatization and cysteine methylation by imidazole reaction in a deuterated environment for enhanced product ion analysis

The combination of separation techniques and mass spectrometry (MS) for peptide investigation allows superior sensitivity of detection and richer fragmentation data than available by direct MS analysis of a complex mixture. In this regard, liquid chromatography (LC) coupled to electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) MS have evolved as versatile analytical tools in proteomics. Very often, however, the product ion mass spectrum is either incomplete or overfilled with ions, thus making sequence analysis difficult. Here we report overall ion intensity improvement of C-terminal lysine-containing peptides from Lys-C digest by on-column derivatization of lysines with 2-methoxy-4,5-dihydro-1H-imidazole. The method is simple, fast and exhibits 100% efficiency of the reaction. Additionally, post-source decay carried out on derivatized peptides gave rise almost exclusively to y-series ion formation, at 100% sequence coverage and high intensity. The novelty of the method resides in the side reaction of this derivatization process, namely the methylation of cysteines. This facilitates the estimation of the disulfide bridge position in a protein and the fragmentation of cysteine-containing peptide fragments. Additionally, by using this derivatization procedure, the loss of peptides, their degradation and/or oxidation, usually occurring in digest alkylation procedures, is greatly minimized. The new on-column derivatization protocol is designed to be carried out on C18 Spin Tubes or Cleanup C18 Pipette Tips. We observed that use of buffered D2O solvent prevented unwanted oxidation and degradation reactions with respect to the stationary phase. This may be due to the fact that a deuteron is less polar than a proton, and thus the bonded silica stationary phase saturated with deuterons does not affect the reaction between epsilon-amino or cysteine thiol groups and 2-methoxy-4,5-dihydro-1H-imidazole. Complete tagging of the peptides by on-column reaction could be obtained when using D2O, as compared to water-based reaction. Methylation of cysteine residues was enhanced when beta-mercaptoethanol was added in the reactant solution.     

Phenolic Profile,Antioxidant Capacity and Antimicrobial Activity of Nettle Leaves Extracts Obtained by Advanced Extraction Techniques

Nettle is a widely known plant whose high biological activity and beneficial medicinal effects are attributed to various bioactive compounds, among which polyphenols play an important role. In order to isolate polyphenols and preserve their properties, advanced extraction techniques have been applied to overcome the drawbacks of conventional ones. Therefore, microwave-assisted extraction (MAE) has been optimized for the isolation of nettle leaves polyphenols and it was compared to pressurized liquid extraction (PLE) and conventional heat-reflux extraction (CE). The obtained extracts were analyzed for their individual phenolic profile by UPLC MS² and for their antioxidant capacity by ORAC assay. MAE proved to be the more specific technique for the isolation of individual phenolic compounds, while PLE produced extracts with higher amount of total phenols and higher antioxidant capacity. Both techniques were more effective compared to CE. PLE nettle extract showed antimicrobial activity against bacteria, especially against Gram-negative Pseudomonas fragi ATCC 4973 and Campylobacter jejuni NCTC 11168 strains. This suggests that PLE is suitable for obtaining a nettle extract with antioxidant and antimicrobial potential, which as such has great potential for use as a value-added ingredient in the food and pharmaceutical industry.     

History Marginalization Improves Forecasting in Variational Recurrent Neural Networks

Deep probabilistic time series forecasting models have become an integral part of machine learning. While several powerful generative models have been proposed, we provide evidence that their associated inference models are oftentimes too limited and cause the generative model to predict mode-averaged dynamics. Mode-averaging is problematic since many real-world sequences are highly multi-modal, and their averaged dynamics are unphysical (e.g., predicted taxi trajectories might run through buildings on the street map). To better capture multi-modality, we develop variational dynamic mixtures (VDM): a new variational family to infer sequential latent variables. The VDM approximate posterior at each time step is a mixture density network, whose parameters come from propagating multiple samples through a recurrent architecture. This results in an expressive multi-modal posterior approximation. In an empirical study, we show that VDM outperforms competing approaches on highly multi-modal datasets from different domains.     

Revision of the Absolute Configurations of Chelocardin and Amidochelocardin

Even with the aid of the available methods, the configurational assignment of natural products can be a challenging task that is prone to errors, and it sometimes needs to be corrected after total synthesis or single-crystal X-ray diffraction (XRD) analysis. Herein, the absolute configuration of amidochelocardin is revised using a combination of XRD, NMR spectroscopy, experimental ECD spectra, and time-dependent density-functional theory (TDDFT)-ECD calculations. As amidochelocardin was obtained via biosynthetic engineering of chelocardin, we propose the same absolute configuration for chelocardin based on the similar biosynthetic origins of the two compounds and result of TDDFT-ECD calculations. The evaluation of spectral data of two closely related analogues, 6-desmethyl-chelocardin and its semisynthetic derivative 1 , also supports this conclusion.     

Potential of AMnO3 (A=Ca,Sr, Ba,La) as Active Layer in Inorganic Perovskite Solar Cells

Inorganic perovskite CaMnO was proposed as a substitution for the TiO anatase in electron transport layers of solar cells containing the hybrid perovskite CH NH PbI based on increased mobility of electrons and better optical matching. Due to a suitable band gap concerning the absorption of sunlight, we investigate the potential of CaMnO and similar manganite perovskites, where Ca is replaced by either Sr, Ba or La, as an absorber layer in inorganic perovskite solar cells. In this study, we have used optical measurements on the synthesized AMnO (A=Ca, Sr, Ba, La) samples to aid density functional theory calculations (DFT) in order to accurately simulate the electronic and optical properties of AMnO compounds and gauge their potential for the role of absorber layer. Both experimental measurements and theoretical calculations show suitable band gap of 1.1-1.5 eV, depending on the compound, and absorption coefficients of the order of cm in the visible part of the spectrum.     

Phonon confinement effects in Raman spectra of porous silicon at non‐resonant excitation condition

Light emitting porous silicon samples with different porosities, i.e. crystalline sizes, were produced from the low level doped p‐type silicon wafers by the anodization process. The effects of strong phonon confinement, redshift and broadening, were found on the O(Γ) phonon mode of the Raman spectra recorded at non‐resonant excitation condition using a near infrared 1064 nm laser excitation wavelength. Similarly, the blueshift of the photoluminescence peak was observed by reducing the crystalline sizes. Vibrational and optical findings were analysed within the existing models of confinement on the vibrational and electronic states of silicon nanocrystals. Since the energy of the photoluminescence peak of small nanocrystals also depends on the oxygen content on the surface of nanocrystals, the surface oxidation states were examined using infrared and energy dispersive spectroscopy. The partial coverage of the surface of nanocrystals was found due to the sample exposure to air. As a consequence, the photoluminescence energy did not increase as would be expected from the quantum confinement model. These results further indicate that the oxygen passivation along with the quantum confinement determines the electronic states of the silicon nanocrystals in porous silicon. Copyright © 2014 John Wiley & Sons, Ltd.