Recycling and screen-segmented column isotachophoresis, two free-fluid approaches for fractionation of proteins.

Recycling and screen-segmented column isotachophoresis (ITP), two approaches for the milligrams to grams preparative-scale purification of proteins, are discussed and compared. Recycling ITP was performed in a recycling free-flow focusing apparatus. In this process, fluid flows rapidly through a narrow channel and the effluent from each channel is reinjected into the electrophoresis chamber through the corresponding input port. The residence time in the cell is of the order of 1 s per single pass, which does not allow complete separation, so recycling is essential to attain the steady state. Immobilization of the advancing zone structure is obtained via a controlled counterflow. Thirty fractions of about 4 ml each are obtained. Column ITP was executed in a Rotofor apparatus and in a similar column operated vertically and without rotation. These instruments feature a screen-segmented annular separation space with twenty subcompartments of about 2 ml each. With both approaches, the collected fractions were analysed separately for conductivity, pH and UV absorbance. Selected fractions were characterized by analytical electrophoretic methods. Examples presented include the cationic and anionic ITP behaviour of model proteins, including bovine serum albumin, ovalbumin and ribonuclease A, and the ITP removal of the major impurities from a commercial ovalbumin sample. These examples revealed that the screen-segmented column is suitable for ITP protein purification and operates optimally in a horizontal rotating mode and without internal cooling. The recycling experiments showed that counterflow improves separation and the steady-state patterns are dependent on the fluid layer thickness in the separation cell but, with a given gap, essentially independent of applied current and recycling pump rate.     

Stereoisomeric flavor compounds,part LVIII: The use of heptakis(2,3-di-O-methyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin as a chiral stationary phase in flavor analysis

The characteristics of the new chiral stationary phase heptakis(2,3-di-O-methyl-6-O-tert-butyldimethylsilyl)-β-cyclodextrin are outlined and compared with permethyl- and perethyl-β-cyclodextrins.     

Novel UDP-glycal derivatives as transition state analogue inhibitors of UDP-GlcNAc 2-epimerase

The "epimerisation" of UDP-GlcNAc to ManNAc, the first step in the biosynthesis of sialic acids, is catalyzed by UDP-GlcNAc 2-epimerase. In this paper we report the synthesis of transition state based inhibitors of this enzyme. To mimic the assumed first transition state of this reaction (TS 1), we designed and synthesized the novel UDP-exo-glycal derivatives 1-4. We also report herein the synthesis of 5 and 6, the first C-glycosidic derivatives of 2-acetamidoglucal, and the synthesis of the ketosides 7 and 8, which were designed as bis-substrate analogue and bis- product analogue, respectively, to mimic the second step of the reaction via the assumed second transition state TS 2.     

Steady-state analysis of the Anaerobic Digestion Model No. 1 (ADM1)

The steady-state behavior of the Anaerobic Digestion Model No. 1 (ADM1) with respect to dilution rate and substrate concentration is analyzed in this study. Thereby, up to ten coexisting steady-state solutions are observed under the same operating conditions. The parameter region of a methane producing operation is limited regarding high dilution rates as well as low or high substrate concentrations. The underlying mechanisms causing those limits are investigated in detail, and the common core is identified, namely a positive feedback loop between growth of acetate degraders and acetate itself. The difference lies in the activation mechanisms of this feedback loop, which differs in the three investigated cases. The comparison of the present results with literature studies of simpler two-step models reveals qualitative differences regarding the substrate concentration. Therefore, an alternative simplified model is suggested, which shows qualitatively the same bifurcation behavior as the ADM1 considering variation of the substrate concentration.     

Electrolyte system strategies for anionic isotachophoresis with electrospray‐ionization mass‐spectrometric detection. 1. Regular isotachophoresis and free‐acid isotachophoresis

The subject of this work is the definition of a simple model based on general ITP theory that allows describing and predicting the behavior of ITP systems compatible with ESI‐MS detection. The model is exemplified by anionic ITP of weak acids that represent an interesting potential application field of ITP‐ESI‐MS. Suitable ESI‐compatible electrolyte systems of very simple composition are proposed including a special free‐acid ITP arrangement. The properties of these systems are discussed using illustrative diagrams of their stacking windows. The use of anionic ITP‐ESI‐MS in negative‐ion ESI mode is reported for the first time and its suitability for sensitive trace analysis is demonstrated. The presented ITP‐ESI‐MS application example comprises a free‐acid ITP system formed of formic and propionic acids and direct injection analysis of ibuprofen and diclofenac in waters with quantitation limits of the order 10^?10 M.     

Characterisation of Taxlllaids A–G; Natural Products from Xenorhabdus indica

Six new lipodepsipeptides and an additional linear derivative named taxlllaids A–G ( 1 – 7 ) have been identified in the entomopathogenic bacterium Xenorhabdus indica. The structures of the main compounds have been solved by detailed NMR spectroscopic analysis and the structures of minor derivatives were elucidated by a combination of labelling experiments and detailed MS experiments. The absolute configuration of the taxlllaids was deduced by using the advanced Marfey method and analysis of the biosynthesis gene cluster showing the presence of epimerisation domains, which was subsequently proved to be correct by solid‐phase peptide synthesis of all taxlllaids. The exchange of a single amino acid in the adenylation domain was shown to be responsible for substrate promiscuity of the third A domain, resulting in the incorporation of leucine, phenylalanine or tyrosine. Bioactivity testing revealed the taxlllaids to be weakly active against Plasmodium falciparum and against a number of eukaryotic cell lines.     

Structure and properties of Na-magadiite dependent on temperature

The mechanism of the thermal behaviour of Na-magadiite at a temperature range from 293–573 K was developed based on X-ray diffraction, DTA, DTL, Q-TG, and gravimetric measurements. A structure model (BRANDT et al.) previously proposed was additionally used. It is possible to show that there probably is a connection between the typical modifications of basal spacing and significant shiftings of silicate sheets. Different steps of dehydration, decrystallization and structure variations can be ascribed to the several thermal transitions. The results depend on sample treatment such as the existent or not existent solid-water vapour interaction and the establishment of equilibrium. It seems that there is a predominance of the first mentioned influence.