Adaption of a fragment analysis technique to an automated high-throughput multicapillary electrophoresis device for the precise qualitative and quantitative characterization of microbial communities

The analysis of microbial communities is of increasing importance in life sciences and bioengineering. Traditional techniques of investigations like culture or cloning methods suffer from many disadvantages. They are unable to give a complete qualitative and quantitative view of the total amount of microorganisms themselves, their interactions among each other and with their environment. Obviously, the determination of static or dynamic balances among microorganisms is of fast growing interest. The generation of species specific and fluorescently labeled 16S ribosomal DNA (rDNA) fragments by the terminal restriction fragment length polymorphism (T-RFLP) technique is a suitable tool to overcome the problems other methods have. For the separation of these fragments polyacrylamide gel sequencers are preferred as compared to capillary sequencers using linear polymers until now because of their higher electrophoretic resolution and therefore sizing accuracy. But modern capillary sequencers, especially multicapillary sequencers, offer an advanced grade of automation and an increased throughput necessary for the investigation of complex communities in long-time studies. Therefore, we adapted a T-RFLP technique to an automated high-throughput multicapillary electrophoresis device (ABI 3100 Genetic Analysis) with regard to a precise qualitative and quantitative characterization of microbial communities.     

Effect of particle size in preparative reversed-phase high-performance liquid chromatography on the isolation of epigallocatechin gallate from Korean green tea

To isolate epigallocatechin gallate (EGCG) of catechin compounds from Korean green tea (Bosung, Chonnam), a C18 reversed-phase preparative column (250x22 mm) packed with packings of three different sizes (15, 40-63, and 150 microm) was used. The sample extracted with water was partitioned with chloroform and ethyl acetate to remove the impurities including caffeine. The mobile phases in this experiment were composed of 0.1% acetic acid in water, acetonitrile, methanol and ethyl acetate. The injection volume was fixed at 400 microl and the flow rate was increased as the particle size becomes larger. The isolation of EGCG with particle size was compared at a preparative scale and the feasibility of separation of EGCG at larger particle sizes was confirmed. The optimum mobile phase composition for separating EGCG was experimentally obtained at the particle sizes of 15 and 40-63 microm in the isocratic mode, but EGCG was not purely separated at the particle size of 150 microm.     

Torsionsabhängige Beiträge zur Spin-Bahn-Kopplung in organischen Molekülen

Zusammenfassung Die Spin-Bahn-Kopplung der niedrigen elektronischen Anregungszustände von organischen Molekülen wird untersucht. Es wird gezeigt, daß die Zerstörung der aromatischen Ebene durch Torsionen oder Schwingungen zu nicht-verschwindenden Zweizentrenbeiträgen führt. Am Beispiel des Triphenylmethyl-Kations wird mit Hilfe der LCAO MO SCF-Methode die Größenordnung der auftretenden Wechselwirkungen ermittelt. Diese haben wahrscheinlich großen Einfluß auf die strahlungslosen Prozesse in Molekülen mit leicht beweglichen Gruppen.

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Spin-orbit coupling of the lower excited electronic states of organic molecules has been investigated. It is shown that any destruction of the aromatic plane through torsions or vibrations leads to non-vanishing contributions from two-centre integrals. The order of magnitude of this type of interactions is determined for the triphenylmethyl-cation with the aid of the LCAO MO SCF method. These interactions probably have much influence on radiationless transitions in molecules containing mobile groups.

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Résumé Etude du couplage spin-orbite dans les états électroniques exités les plus bas des molécules organiques. Toute non planéité aromatique créée par torsions ou vibrations entraîne une contribution non nulle de la part des intégrales bicentriques. L'ordre de grandeur de ce type d'interaction est detérminé, à l'aide de la méthode L.C.A.O. M.O. SCF, pour le cation triphenylméthyle. Ces interactions ont probablement beaucoup d'influence sur les transitions non radiatives dans les molécules contenant des groupements mobiles.

     

Phosphine-mediated reductive condensation of gamma-acyloxy butynoates: a diversity oriented strategy for the construction of substituted furans

Exposure of gamma-acyloxy butynoates to stoichiometric quantities of triphenylphosphine results in reductive condensation to afford substituted furans, by way of allenic ester intermediates. As gamma-acyloxy butynoates are readily obtained through condensation of ethyl propiolate with aldehydes followed by acylation, this method represents a powerful and mechanistically novel protocol for the convergent three-component construction of substituted furans.     

Characterization and epitope mapping of two monoclonal antibodies against human CD99

CD99 plays a critical role in the diapedesis of monocytes, T cell differentiation, and the transport of MHC molecules. Engagement of CD99 by agonistic monoclonal antibodies has been reported to trigger multifactorial events including T cell activation as well as cell-cell adhesion during hematopoietic cell differentiation. In this study, to identify the functional domains participating in the cellular events, we mapped the epitopes of CD99, which are recognized by two agonistic CD99 monoclonal antibodies, DN16 and YG32. Using recombinant fusion proteins of GST with whole or parts of CD99, we found that both antibodies interact with CD99 molecules independently of sugar moieties. DN16 mAb detected a linear epitope located in the amino terminal region of CD99 while YG32 mAb bound another linear epitope in the center of the extracellular domain. To confirm that the identified epitopes of CD99 are actually recognized by the two mAbs, we showed the presence of physical interaction between the mAbs and the fusion proteins or synthetic peptides containing the corresponding epitopes using surface plasmon resonance analyses. The dissociation constants of DN16 and YG32 mAbs for the antigen were calculated as 1.27 x 10(-7) and 7.08 x 10(-9) M, respectively. These studies will help understand the functional domains and the subsequent signaling mechanism of CD99.     

Crystal structure, morphology and composition of magnetron sputtered tungsten carbide films

Summary It is shown that small variations of the deposition parameters during magnetron sputtering of tungsten carbide thin films may result in drastic changes of film properties. An increasing working gas pressure for example lowers stress and hardness values. Simultaneously, the texture of the WC_1–x cristallites turns from 200 preferential orientation to 111, whereas the composition of the films does not change. In reactive sputtering with a tungsten target there is a narrow range from 2 to 3% C₂H₂ gas admixture to the working gas where the films are stochiometric (WC) and hard, and grain size and morphology are similar to that of non-reactively sputtered films. The generation of different crystallite structures and orientations in the range of 0–3% C₂H₂ admixtures are used to produce a multiphase thin film with extremely low crack propagation.