Hydrogen evolution by co-immobilized Chlorella vulgaris and Clostridium butyricum cells

Whole cells of Chlorella vulgaris and Clostridium butyricum were co-immobilized in 2% agar gel. NADP was suitable as an electron carrier. The rate of hydrogen evolution increased with increasing NADP concentration. The optimum conditions for hydrogen evolution were pH 7.0 and 37°C. The immobilized C. vulgaris-NADP-immobilized Cl. butyricum system continuously evolved hydrogen at a rate of 0.29–1.34 μmol/h per mg Chl for 6 days. On the other hand, the system without NADP evolved only a trace amount of hydrogen.     

Measurement of sulfidopeptide leukotrienes and their metabolism in human synovial fluid of patients with rheumatoid arthritis

Leukotriene (LT)C4 in the synovial fluid of patients with osteoarthritis deformans (OA) and rheumatoid arthritis (RA) was measured by radioimmunoassay (RIA) after extraction with Sep-Pak C18 cartridge. The amounts of immunoreactive LTC4 (i-LTC4) in samples from patients with OA and RA were not significantly different, being 0.198 +/- 0.018 pmol/ml (n = 11) and 0.179 +/- 0.016 pmol/ml (n = 12), respectively. After separation by high performance liquid chromatography (HPLC) and measurement by RIA, the levels of other sulfidopeptide LTs, such as LTD4 and LTE4, in synovial fluid from patients with RA were found to be significantly higher than those in fluid from patients with OA. The leukocyte number in synovial fluids did not correlate with the i-LTC4 level. The metabolic activities of these synovial fluids were determined by incubating them with 3H-LTC4 and then separating sulfidopeptide LTs by HPLC. The conversion of LTC4 to LTD4 in synovial fluids of patients with OA and RA were similar, but the dipeptidase activity converting LTD4 to LTE4 was higher in fluid from patients with RA. It is suggested that a high level of LTE4 may contribute to exudation of synovial fluid, since LTE4 increases vascular permeability.     

Asymmetric Reduction of 4-Oxoisophorone Using Immobilized Thermophilic Bacteria Under Conditions of Controlled Cell Growth

Asymmetric reduction of 2,6,6,-trimethyl-2-cyclohexene-l,4-dione (4-oxoisophrone) to (6R)-2,2,6-trimethyl-1,4-cyclohexane-dione((3R)-dihydro-4-oxoisophorone) was catalysed by immobilized thermophilic bacteria, Thermomonospora curvata JTS 321. Because of leakage of entrapped cells from gel beads during reactions using culture medium, we optimized the medium to allow the microbial conversion under conditions of controlled cell growth. Of the media screened, liver infusion medium was found to be the most suitable and microbial conversion was achieved without cell leakage from the immobilized gels. Immobilized T. curvata cells were repeatedly used for the asymmetric reduction of 4-oxoisophorone, more than 15 times, with an extent of conversion of 50%.     

Structural characterization of lipid A component of Erwinia carotovora lipopolysaccharide

The chemical structure of the lipid A component of lipopolysaccharide excreted into the liquid medium by the plant pathogenic enterobacterium Erwinia carotovora FERM P-7576 was characterized. It consists of a -1, 6-linked glucosamine disaccharide which carries ester-and amide-bound fatty acids and phosphate similar to the lipid A from other gram-negative bacteria. The lipid A preparation was not uniform in the number and composition of the fatty acids linked to the disaccharide. Four prominent lipids A were involved, they were composed of five to seven residues of fatty acid. Among them the major component was hexa-acyl lipid A, in which the hydroxyl group at position 3 and the amino group of the non-reducing glucosamine unit carry 3-dodecanoyl-oxytetradecanoyl residues. Positions 2 and 3 of the reducing glucosamine unit were substituted by 3-hydroxytetradecanoic acid. In the hepta-acyl lipid A, an additional hexadecanoic acid was linked to the hydroxyl group of the 3-hydroxytetradecanoyl residue at position 2 of the hexa-acyl lipid A. Two penta-acyl lipids A were the homologs of the hexa-acyl lipid A with decreasing acylation. Dodecanoic acid was missing from one, and 3-hydroxytetradecanoic acid from another. 3-Dodecanoyloxytetradecanoyl residue at position 3 differentiates E. carotovora lipid A from that of other gram-negative bacteria.Abbreviations LPS lipopolysaccharide - GlcN glucosamine - KDO 3-deoxy-d-manno-octulosonic acid - FAB-MS fast atom bombardment mass spectrometry - u atomic mass unit     

NO(2) Sensor which uses immobilized nitrite oxidizing bacteria

A biosensor consisting of immobilized nitrite oxidizing bacteria and an oxygen electrode has been developed for the amperometric determination of NO(2) (nitrogen dioxide) gas. The response time for the determination of NO(2) was within 3 min. A linear relationship was observed between the current decrease and the NO(2) concentration below 255 ppm. The minimum concentration for the determination of NO(2) was 0.51 ppm. The current decrease was reproducible within +/-4% of the relative error. The selectivity of the microbial sensor for NO(2) was satisfactory. The current output of the sensor was almost constant for more than 24 days and 400 assays.     

Application of a novel apparatus, the quartz chemical analyzer, to the determination of endotoxin in blood.

A novel apparatus called a quartz chemical analyzer (QCA) has been developed using a quartz crystal resonator. This apparatus measures sample viscosity changes based on resonant frequency changes of the quartz crystal. The apparatus was used to determine bacterial endotoxin concentrations by monitoring the gelation reaction of Limulus amebocyte lysate. The QCA determined endotoxin concentrations with good accuracy and reproducibility in the range of 0.001-3 EU/ml for endotoxin standard (JP XII). For endotoxin determination in human whole blood and plasma samples, the inhibitory reaction was eliminated by pretreatment of a fourfold dilution at 60 degrees C and incubation for 30 min. There are many advantages of the QCA method compared with the turbidimetric and chromogenic methods. For example, QCA can measure sample viscosity changes with high sensitivity and accuracy because QCA detects minor resonant frequency changes and the frequency data give a numerical value for easy quantitation. QCA can examine turbid samples, and the required quantities of samples and reagents are small, since the quartz crystal detects sample viscosity changes directly. The endotoxin determination time may be shortened by raising the reaction temperature, and QCA can detect other types of coagulation reactions.     

Photo-induced activation of cytochrome P450/reductase fusion enzyme coupled with spinach chloroplasts

Summary Photoactivation of cytochrome P450 monooxygenase was studied using a combination of spinach chloroplasts and yeast microsomes containing rat P4501A1/yeast reductase fusion enzyme. Under illumination, in the reaction mixture, NADP was reduced, transferring electrons to the P450/reductase fusion enzyme to convert 7-ethoxycoumarin to 7-hydroxycoumarin.     

A comparison of screening methods for antioxidant activity in seaweeds

The inhibition of lipid peroxidation and radical scavenging effects were studied to evaluate the antioxidant activity for extracts of 17 species of seaweed. The antioxidant effect was evaluated by determination of lipoxygenase activity and by α, α-diphenyl-β-picrylhydrazyl (DPPH) decolorization. Lipoxygenase activity was depressed in the presence of aqueous and ethanol extracts of 4 algal species; Sargassum species had the highest antioxidant activity of all the species examined. The ethanol extracts of one Sargassum species showed competitive inhibition with the substrate. The same species also showed radical scavenging activity in the DPPH decolorization test. Comparison of these results shows no relationship between enzyme inhibition and radical scavenging activity. This revised version was published online in August 2006 with corrections to the Cover Date.     

Changes in eicosapentaenoic acid content of Navicula saprophila, Rhodomonas salina and Nitzschia sp. under mixotrophic conditions

Three species of microalgae able to produce eicosapentaenoic acid(EPA) were collected from brackish and sea water around Japan. The species were identified as Navicula saprophila, Rhodomonassalina and Nitzschia sp. EPA as a proportion of total fatty acids increased in the presence of acetic acid for Rhodomonas salina and Nitzschia sp. However, Navicula saprophila displayed the greatest productivity of EPA and the EPA content of its biomass was enhanced under mixotrophic conditions in the presence of acetic acid. This revised version was published online in August 2006 with corrections to the Cover Date.