Identification of Shiga toxins in Shiga toxin-producing Escherichia coli using immunoprecipitation and high-performance liquid chromatography-electrospray ionization mass spectrometry

We developed a rapid and reliable identification method for Shiga toxins in Shiga toxin-producing Escherichia coli (STEC) using immunoprecipitation and high-performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS). Polyclonal antisera specific for Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) were raised in rabbits so as to be used for the immunoprecipitation. The immunoprecipitaion was carried out by mixing sample solutions with 50 microl each of the antisera to Stx1 and Stx2 followed by allowing the mixed solutions to stand for 30 min. The quantity required to obtain the immunoprecipitate was more than 0.5 microg of Shiga toxins. HPLC-ESI-MS analysis of the resulting immunoprecipitates provided accurate molecular weight information on Shiga toxins, leading to direct evidence for the presence of these toxins. It requires at most two days to perform our procedure from toxin extraction to measurement of HPLC-ESI-MS whereas the previous method using isolation procedures required about two weeks to complete. The usefulness of the present method has been demonstrated by identifying Stx1, Stx2 and a variant of Stx2 (Stx2e) in the immunoprecipitates prepared from STEC strains.     

Synthesis of polyamides by the polyaddition of bisimidazoline with dicarboxylic acids

N,N′-Dipropionylethylenediamine was synthesized by the ring-opening addition reaction of 2-ethyl-2-imidazoline with propionic acid at 220°C. By applying this reaction to polymerization, polyamides were synthesized by the ring-opening polyaddition reaction at 220°C. of 1,4-bis(imidazoline-2-yl)butane with adipic acid, succinic acid, sebacic acid, and terephthalic acid. The reaction product of 1,4-bis(imidazoline-2-yl)butane with adipic acid, which was proposed to be nylon 26, was compared with an authentic sample of nylon 26 and shown to possess a very similar infrared spectrum and melting point.     

Reaction of carbon monoxide with N-lithioketimines derived by the addition of alkyllithium to ortho-substituted benzonitriles. Diverse courses of cyclization via carbonyl anions

N-Lithioketimines, generated in situ from 2-substituted benzonitriles and t-BuLi, reacted with carbon monoxide to afford five-, six- or seven-membered cyclic products, involving participation by the ortho substituents. The reactions represent new selective transformations of carbonyllithium intermediate which previously were observed to lead to complicated reactions.     

First linear alignment of five C-Se...O...Se-C atoms in anthraquinone and 9-(methoxy)anthracene bearing phenylselanyl groups at 1,8-positions

Five Ci-Se...O...Se-Ci atoms in anthraquinone and 9-(methoxy)anthracene bearing phenylselanyl groups at 1,8-positions align linearly, the origin of which is shown to be a nonbonded 5c-6e interaction of the five atoms.     

Phase Separation in Sol-Gel System Containing Mixture of 3- and 4-Functional Alkoxysilanes

The phase separation behavior of gelling systems containing the mixture of 3-functional and 4-functional alkoxysilanes has been investigated. The relation between the starting composition and resultant macroporous morphology was examined using tetramethoxysilane (TMOS) and vinyltrimethoxysilane (VTMS) as starting alkoxysilanes, formamide (FA) as an additive, under an acidic condition. Up to TMOS:VTMS molar ratio of 0.5:0.5, the phase relation remained almost unchanged from that of pure TMOS system which exhibits morphology with well-defined co-continuous macropores in a very limited concentration region. On the VTMS-rich side typically TMOS:VTMS = 0.2:0.8, however, the co-continuous macroporous morphology was obtained in a broader composition range than those of either pure TMOS or VTMS system. A dome-like pseudo binary region was obtained with the two-phase region extending toward FA-rich direction. The domain size and pore volume of the gels with macroporous morphology could be controlled by alkoxide:water ratio and total solvent fraction, respectively.     

Circular dichroism spectra of the inclusion complexes of phlorizin in cyclodextrins

Two aromatic rings of a phlorizin molecule form inclusion complexes with -CD and -CD. Induced circular dichroism spectra of these complexes have been measured to estimate the orientation of the two aromatic rings in the hydrophobic space of CDs. Apparent complex formation constants have been also estimated for each complex. It is concluded that phlorizin forms a stronger inclusion complex with -CD than with -CD.     

Application of an Ion Beam Technique for the Design of Visible Light-Sensitive, Highly Efficient and Highly Selective Photocatalysts: Ion-Implantation and Ionized Cluster Beam Methods

As a new concept for the design of effective photocatalysts, an ion beam technology using accelerated metal ions, a metal ion implantation and an ionized cluster beam (ICB) method, have been applied to design unique photocatalysts. The metal ion implantation of TiO₂ and Ti-containing zeolites with highly accelerated metal ions (V⁺, Cr⁺, etc.) is useful in designing photocatalysts that can operate efficiently under visible light irradiation. Also, an ICB method with a low acceleration energy is useful in preparing transparent TiO₂ thin films on various types of substrates for the efficient photocatalytic degradation of pollutants diluted in water and air. The combination of the ICB method and metal ion implantation can develop the TiO₂ thin films that are able to operate not only under UV light but also under visible light irradiation.     

Photoaffinity labeling of bovine rhodopsin

Photoaffinity labeled (3-diazoacetoxy)-9-cis-retinal (1) and (9-methylenediazoacetoxy)-9-cis-retinal (20) were synthesized and bound to absorption maxima at 465 and 460 nm respectively. Binding studies established that synthetic retinals 1 and 2 bind to the natural binding site and that the integrity of the diazoacetoxy photoaffinity label is preserved in the process. Incorporation of 3-(O¹⁴COCHN₂)-labeled 9-cis retinal could be conveniently carried out in high yield using apomembrane solubilized in CHAPS as detergent to afford the pigment analog in a pure form. Photolysis of the diazoacetoxy group within the binding site led to 15–20%, crosslinking of rhodopsin as estimated by using radiocarbon containing labeled retinal 1 thus showing that this synthetic retinal is suitable for photoaffinity labeling of the active site in rhodopsin. Subsequent experiments to establish the site(s) of crosslinking by sequencing studies will then contribute to our knowledge of the structure of rhodopsin.