Unconventional salt trend from soft to stiff in single neurofilament biopolymers

We present persistence length measurements on neurofilaments (NFs), an intermediate filament with protruding side arms, of the neuronal cytoskeleton. Tapping mode atomic force microscopy enabled us to visualize and trace at subpixel resolution photoimmobilized NFs, assembled at various subunit protein ratios, thereby modifying the side-arm length and chain density charge distribution. We show that specific polyampholyte sequences of the side arms can form salt-switchable intrafilament attractions that compete with the net electrostatic and steric repulsion and can reduce the total persistence length by half. The results are in agreement with present X-ray and microscopy data yet present a theoretical challenge for polyampholyte interchain interactions.     

Development and validation of an improved reversed‐phase liquid chromatographic method combined with pulsed electrochemical detection for the analysis of netilmicin

Netilmicin is one of the aminoglycoside antibiotics that lacks a strong UV absorbing chromophore. However, the application of pulsed electrochemical detection has been used successfully for the direct analysis of aminoglycoside antibiotics. This study describes an improved LC method combined with pulsed electrochemical detection for the analysis of netilmicin. Using a Zorbax SB C‐18 column (250 mm×4.6 mm id, 5 μm), isocratic elution was carried out with a mobile phase containing sodium sulfate (20 g/L), sodium octanesulfonate (0.3 g/L), THF (20 mL/L), and 0.2 M phosphate buffer pH 3.0 (50.0 mL/L). The robustness of the method was examined by means of an experimental design. The method proved to be sensitive, repeatable, linear, and robust. The method has also been used to analyze some commercial netilmicin samples.     

Evaluation of comprehensive on-line liquid chromatography thermally assisted hydrolysis and methylation-gas chromatography-mass spectrometry for characterization of sulfonated lignins

Sulfonated lignins, used as dispersants in agrochemical formulations, have been characterized by hyphenation of ion-pair RPLC and thermally assisted hydrolysis and methylation-GC-MC (THM-GC-MS). The chemical structure of a series of selected lignins was evaluated, both in terms of overall composition and in terms of composition as a function of molecular size. Some sulfonated lignins give rise to unstable formulations. In the compositional analysis, these samples were found to yield additional fragments. A newly developed comprehensive method made it possible to monitor the occurrence of these fragments as a function of molecular size. In addition to differences in the molecular size between "good" and "bad" batches, clear differences in chemical composition were established.     

A rapid and sensitive CE method with field-enhanced sample injection and in-capillary derivatization for selenomethionine metabolism catalyzed by flavin-containing monooxygenases

A rapid and sensitive electrophoretically mediated microanalysis method with field-enhanced sample injection (FESI) for in-capillary derivatization was developed to determine selenomethionine (SeMet) and selenomethionine selenoxide (SeOMet). Phthalic anhydride (PA) was selected as the derivatization reagent due to the fast reaction at room temperature and the stability of derivatives. The in-capillary derivatization was accomplished by electrophoretically mixing PA and sample plugs. PA reagent was introduced hydrodynamically into the capillary, whereas the sample solution was injected electrokinetically, thus allowing a selective preconcentration of the analytes by FESI. For FESI, the optimum sample solvent was 2 mM borate solution. The borate buffer was suitable for both in-capillary derivatization and separation of the derivatives. The combination of electrophoretically mediated microanalysis with FESI for in-capillary derivatization was successfully achieved with about 800-fold concentration sensitivity enhancement compared to direct CE-UV detection in the same setup. The present method is miniaturized and fully automated, which ensures the on-line derivatization, stacking, separation and detection in 10 min. Finally, the developed method was successfully applied to measure enzyme activities by analyzing the reaction mixtures of SeMet with human flavin-containing monooxygenases (FMO). The results showed that both FMO1 and FMO3, but not FMO5 could catalyze the Se-oxygenation of SeMet.     

Impurity profiling of acetylspiramycin by liquid chromatography–ion trap mass spectrometry

Investigation of acetylspiramycin (ASPM) and its related substances was carried out using a reversed-phase liquid chromatography/tandem mass spectrometry method. The identification of impurities in the ASPM complex was performed with a quadrupole ion trap mass spectrometer, with an electrospray ionization (ESI) source in the positive ion mode which provides MSⁿ capability. A total of 83 compounds were characterized in commercial samples, among which 31 impurities that had never been reported and 31 partially characterized impurities were deduced using the collision-induced dissociation (CID) spectra of major ASPM components as templates. Most of the major impurities arise from the starting materials and the synthesis process. This work provides very useful information for quality control of ASPM and evaluation of its synthesis process.