P.M.R. analysis of several phosphorylated aziridines

Several new phosphorylated aziridines of related structure were prepared. P.m.r. analysis via decoupling experiments provided cis H-H, trans H-H, gem H-H and PNCH coupling values. Similar to simple aziridines, the cis H-H coupling is larger than trans H-H coupling (on vicinal ring carbons) which in turn is larger than gem H-H coupling. In one example operating at 100 MHz and 0° it was possible to detect the presence of two invertomers.     

Synthesis of 1,3-dialkylpyrazolo [1,5-a]-1,3,5-triazine-2,4-diones isomers of 1,3-dialkylxanthines

The synthesis of a new series of alkylxanthine analogs containing a bridgehead nitrogen atom is reported. 1,3-Dialkylpyrazolo[1,5-a]-1,3,5-triazine-2,4-diones, were prepared by the treatment of 3-methylpyrazolo[1,5-a]-1,3,5-triazine-2,4-dione (3) with the corresponding alkyl iodide. Similarly, the reaction of 3-methyl-7-phenylpyrazolo[1,5-a]-1,3,5-dialkyl-7-phenylpyrazolo[1,5-a]-1,3,5-triazine-2,4-diones. The starting materials, 3 and 17 , were prepared via the reaction of an appropriate 3-aminopyrazole with ethoxycarbonyl isothiocyanate. Several 8-bromo derivatives were prepared by direct bromination of the 1,3-dialkylpyrazolo[1,5-a]-1,3,5-triazine-2,4-diones.     

Renewable microcolumns for automated DNA purification and flow-through amplification: from sediment samples through polymerase chain reaction

There is an increasing need for field-portable systems for the detection and characterization of microorganisms in the environment. Nucleic acids analysis is frequently the method of choice for discriminating between bacteria in complex systems, but standard protocols are difficult to automate and current microfluidic devices are not configured specifically for environmental sample analysis. In this report, we describe the development of an integrated DNA purification and polymerase chain reaction (PCR) amplification system and demonstrate its use for the automated purification and amplification of Geobacter chapellei DNA (genomic DNA or plasmid targets) from sediments. The system includes renewable separation columns for the automated capture and release of microparticle purification matrices, and can be easily reprogrammed for new separation chemistries and sample types. The DNA extraction efficiency for the automated system ranged from 3 to 25%, depending on the length and concentration of the DNA target. The system was more efficient than batch capture methods for the recovery of dilute genomic DNA even though the reagent volumes were smaller than required for the batch procedure. The automated DNA concentration and purification module was coupled to a flow-through, Peltier-controlled DNA amplification chamber, and used to successfully purify and amplify genomic and plasmid DNA from sediment extracts. Cleaning protocols were also developed to allow reuse of the integrated sample preparation system, including the flow-through PCR tube.     

Syntheses of some indazoles structurally related to equilenin

This study reports the complete synthesis and characterization of the 15, 16-diazaequilenin derivative 10,11-dihydro-3H-naphth[1,2-g]indazol-7-ol ( 2b ) as well as the methyl ether 2a of the above compound and the novel “model” compound 4,5-dihydro-1H-benz[g]indazol-7-ol ( 3b ). Indazoles 2a and 3a have demonstrated in vitro activity against a variety of microorganisms.     

Silver(I)-imidazole cyclophane gem-diol complexes encapsulated by electrospun tecophilic nanofibers: formation of nanosilver particles and antimicrobial activity

Silver(I)-imidazole cyclophane gem-diol complex, 3 [Ag2C36 N10(O)4](2+)2(x)-, where x = OH- or CO3(2-), was synthesized and well characterized. The minimum inhibition concentration tests showed that the aqueous form of 3 is 2 times less effective as an antibiotic than 0.5% AgNO3, with about the same amount of silver. The antimicrobial activity of 3 was enhanced when encapsulated into Tecophilic polymer by electrospinning to obtain mats made of nano-fibers. The fiber mats released nanosilver particles, which in turn sustained the antimicrobial activity of the mats over a long period of time. The rate of bactericidal activity of 3 was greatly improved by encapsulation, and the amount of silver used was much reduced. The amount of silver contained in the fiber mat of 3, with 75% of 3 and 25% Tecophilic, was 8 times less than that in 0.5% AgNO3 and 5 times lower than that in silver sulfadiazine cream 1%. The fiber mat was found to kill S. aureus at the same rate as 0.5% AgNO3, with zero colonies on an agar plate, and about 6 times faster than silver sulfadiazine cream. The silver mats were found effective against E. coli, P. aeruginosa, S. aureus, C. albicans, A. niger, and S. cerevisiae. Transmission electron microscopy and scanning electron microscopy were used to characterize the fiber mats. The acute toxicity of the ligand (imidazolium cyclophane gem-diol dichloride) was assessed by intravenous administration to rats, with an LD 50 of 100 mg/kg of rat.     

E. coli adhesion to silica in the presence of humic acid

The influence of humic acid on the adhesion of Escherichia coli to silica particles or glass surfaces was investigated. After adsorbing various amounts of humic acid to the particles or surfaces, bacteria were added to the sample and allowed to adhere. For the silica particles the number of bacteria-particle couplets formed were counted from video microscopy images. For the glass surfaces, a differential electrophoresis force was applied, and the force required to detach the bacteria was quantified. These experiments showed a slight increase in the number of couplets formed in the presence of humic acid, and also showed a slight increase in the force required for detachment of the bacteria. Although an increase in adhesion number and strength was measured, the magnitude of the increase was small, indicating that humic acid plays a small role in bacterial adhesion to silica or glass surfaces.     

The boron trifluoride nitromethane adduct

The separation of the boron isotopes using boron trifluoride·organic-donor, Lewis acid·base adducts is an essential first step in preparing ¹⁰B enriched and depleted crystalline solids so vital to nuclear studies and reactor applications such as enriched MgB₂, boron carbide, ZrB₂, HfB₂, aluminum boron alloys, and depleted silicon circuits for radiation hardening and neutron diffraction crystal structure studies. The appearance of this new adduct with such superior properties demands attention in the continuing search for more effective and efficient means of separation. An evaluation of the boron trifluoride nitromethane adduct, its thermodynamic and physical properties related to large-scale isotopic separation is presented. Its remarkably high separation factor was confirmed to be higher than the expected theoretical value. However, the reportedly high acid/donor ratio was proven to be an order of magnitude lower. On-going research is determining the crystal structure of deuterated and ¹¹B enriched ¹¹BF₃·CD₃NO₂ by X-ray and neutron diffraction.     

Towards a continuous dynamic kinetic resolution of 1-phenylethylamine using a membrane assisted, two vessel process

A continuous process with two separated reaction vessels provides a solution to the problems surrounding the combination of two catalysts in dynamic kinetic resolution reactions by retaining the biocatalyst in a lower temperature vessel with a microfiltration membrane and allowing the racemisation to occur efficiently in a higher temperature vessel.     

NMR-based mapping of disulfide bridges in cysteine-rich peptides: application to the mu-conotoxin SxIIIA

Disulfide-rich peptides represent a megadiverse group of natural products with very promising therapeutic potential. To accelerate their functional characterization, high-throughput chemical synthesis and folding methods are required, including efficient mapping of multiple disulfide bridges. Here, we describe a novel approach for such mapping and apply it to a three-disulfide-bridged conotoxin, mu-SxIIIA (from the venom of Conus striolatus), whose discovery is also reported here for the first time. Mu-SxIIIA was chemically synthesized with three cysteine residues labeled 100% with (15)N/(13)C, while the remaining three cysteine residues were incorporated using a mixture of 70%/30% unlabeled/labeled Fmoc-protected residues. After oxidative folding, the major product was analyzed by NMR spectroscopy. Sequence-specific resonance assignments for the isotope-enriched Cys residues were determined with 2D versions of standard triple-resonance ((1)H, (13)C, (15)N) NMR experiments and 2D [(13)C, (1)H] HSQC. Disulfide patterns were directly determined with cross-disulfide NOEs confirming that the oxidation product had the disulfide connectivities characteristic of mu-conotoxins. Mu-SxIIIA was found to be a potent blocker of the sodium channel subtype Na(V)1.4 (IC50 = 7 nM). These results suggest that differential incorporation of isotope-labeled cysteine residues is an efficient strategy to map disulfides and should facilitate the discovery and structure-function studies of many bioactive peptides.     

Localized quorum sensing in Vibrio fischeri

Quorum sensing is almost always regarded as a population density effect in three-dimensional bulk samples of bacteria. Here we create two-dimensional samples of Vibrio fischeri cells adhered onto glass surfaces to examine the effect of local population densities on quorum sensing. This is done by measuring the luminescent response. The 2-D bacterial populations enable us to simultaneously account for time and distance effects on quorum sensing, which were previously very challenging to access in typical three-dimensional bulk samples. Thus, we are able to consider quorum sensing in terms of signal diffusion. A diffusion model of quorum sensing signals guides the experiments and shows that for a given cell spacing (density) and diffusion time there exists a “true quorum”— a number of cells necessary for quorum sensing. We find that quorum sensing can occur locally in 2-D surface samples and is a function of cell population density as well as signal diffusion time.