Production of meiosis-activating sterols from metabolically engineered yeast

Meiosis-activating sterols (MAS), a class of potent regulators of reproductive processes, are difficult to obtain by chemical synthesis or isolation from natural sources. We demonstrate the development of metabolically engineered strains of Saccharomyces cerevisiae that accumulate MAS as the predominant sterol product. Homologous recombination was used to construct an erg24Delta erg25Delta hem1Delta mutant RXY4.3, which lacked sterol Delta14 reductase, C-4 oxidase, and delta-aminolevulinate synthase. The HEM1 deletion allowed sterol import and rendered RXY4.3 viable under aerobic conditions. This mutant accumulated 4,4-dimethyl-5alpha-cholesta-8,14,24-trien-3beta-ol (FF-MAS), and a similar erg25Delta hem1Delta mutant produced 4,4-dimethyl-5alpha-cholesta-8,24-dien-3beta-ol (T-MAS). Based on consistent yields of approximately 5 mug of FF-MAS per mL of culture, fermentation of genetically modified yeast compares favorably with other approaches to produce MAS.     

A phenoxyl radical complex of copper(II)

A new N,O-bidentate pro-ligand (HL), [ML2] (M = Cu, Zn) and [CuL2][BF4] have been synthesised; [CuL2].4DMF and [CuL2][BF4].2CH2Cl2 have been crystallographically and spectroscopically characterised; these data indicate that [CuL2]+ cations are constituted as [Cu2+(L.)(L-)]+ and involve the phenoxyl radical L..     

Composition-dependent microhardness and its relationship to bonding in sodium tungsten bronzes

The technique of microhardness measurements using diamond indenters is outlined and assessed for its potential use in quantifying bonding changes and studying reactions in nonstoichiometric crystals. Results are presented for both Vickers and Knoop hardness values on {001} and {011} crystal planes of cubic sodium tungsten bronzes, Na_xWO₃, with x in the range 0.5 to 0.75. The Knoop data show that in only one direction, 〈110〉 on {001}, is the hardness sensitive to changes in composition. Hardness in the 〈110〉 directions and the degree of anisotropy increase as the sodium content of the bronze increases. All the crystal faces examined showed marked anisotropic behavior, with 〈110〉 being about 50% harder than 〈100〉 on {001} faces, while on {011} planes hardness increases in the sequence $\text{〈100〉:〈21}\text{1}\text{〉:〈11}\text{1}\text{〉 ≈ 〈01}\text{1}\text{〉}$. Hardness results from isomorphous and isoelectronic ReO₃ are considered with the Na_xWO₃ data to show the dominant role played by Na⁺WO₃ matrix interactions in determining the properties of these materials. The results are discussed in terms of current bonding theories for bronzes.     

Determination of bradykinin and arg-bradykinin in rat muscle tissue by microdialysis and capillary column-switching liquid chromatography with mass spectrometric detection

Quantification of bradykinin peptides in limited amounts of rat muscle tissue dialysate has been performed using a packed capillary LC-ESI-TOF-MS method. The micro dialysate samples (450 microL) with added internal standard were loaded onto a 1 mm x 5 mm loading column packed with 5 microm Kromasil C18 particles by a carrier solution of 0.1% formic acid in ACN/water (5:95, v/v) at a flow rate of 250 microL/min for online preconcentration of the analytes. Back-flushed elution onto a 150 mm x 0.5 mm Zorbax C18 column packed with 5 microm particles was conducted using a linear solvent ACN/H2O gradient containing 0.1% formic acid. (Tyr8)-bradykinin was used as an internal standard and was added to the dialysis sample prior to injection. Baseline separation of bradykinin, arg-bradykinin and (tyr8)-bradykinin was achieved within 10 min. Positive ESI was performed in the m/z range of 200-1300. The method was validated in the range 0.2-1.0 ng/mL dialysate, yielding correlation coefficients of 0.995 and 0.990 for bradykinin and arg-bradykinin, respectively. The within-assay and between-assay precisions were between 4.3-9.6% and 6.2-10.6%, respectively. Both arg-bradykinin and bradykinin were detected in dialysate from rat muscle tissue, at concentrations of 0.1 and 0.4 ng/mL for bradykinin and arg-bradykinin, respectively, confirming the presence of arg-bradykinin in rat muscles.     

On the structure of Sidon sets

LetG be any connected compact group with dual object. We give in this paper a new proof that the union of any two Sidon sets in is again a Sidon set. We also show that any Sidon subset of is the union of a set whose elements have bounded degree with a finite union of sets which satisfy a quasi-independence condition.     

Metabonomic Studies Comparing Capillary and Conventional HPLC-oa-TOF MS for the Analysis of Urine from Zucker Obese Rats

The application of capillary HPLC and orthogonal acceleration (oa) TOF-MS to a metabonomic investigation of the urinary metabolic profiles of samples obtained from male and female Zucker rats is described. The results obtained using capillary HPLC for performing the chromatographic separation are also compared with the analysis of the same samples by conventional HPLC. Capillary HPLC-MS gave increased sensitivity (ca. 100 fold) despite using a fraction of the sample volume consumed by HPLC (0.5 L vs 10 L). Capillary HPLC-MS also provided a greater peak count (ca. 3000 ions compared to 1500 for HPLC) for the same analysis time compared to conventional HPLC-MS. These data were processed using partial least squares discriminant analysis (PLS-DA)) enabling markers of diurnal variation to be detected for urine samples from both male and female rats.     

Thermal and photochemistry of a pyrene dihydrodioxin (PDHD) and its radical cation: a photoactivated masking group for ortho-quinones

Pyrene dihydrodioxins (1 and 2) have been synthesized and shown to be effective photochemical blocking groups for pyrene-4,5-dione (3). The mechanism of quinone release proceeds through the formation of a remarkably stable radical cation. Direct evidence is provided that this radical cation is not only thermally labile but also photochemically labile, and that both pathways lead to quinone extrusion. Once initiated with UV light, the pyrene quinone product serves as an electron-transfer photosensitizer for the further release of quinone with visible light.     

Micellar electrokinetic capillary chromatography of algal toxins

Recent methods employed for the analysis of algal toxins have focused on high performance liquid chromatography. However these methods suffer from poor resolution, poor efficiency, and long analysis times. This study involves the investigation of a number of toxins including nodularin, microcystin LR, YR, and RR which are cyclic peptides produced by strains of blue-green algae. The electroseparation mode was micellar electrokinetic capillary chromatography (MECC) using a borate buffer containing sodium dodecyl sulfate (SDS) as the surgactant of choice. The method was optimized with standard toxin compounds and employed for the screening opf toxins in supercritical fluid extracts (SFE) of freeze-thawed algal scum samples.