Zygomycetes from herbivore dung in the ecological reserve of Dois Irmãos, Northeast Brazil

Thirty-eight taxa of Zygomycetes distributed in 15 genera were recorded from tapir (Tapirus terrestris), camel (Camelus bactrianus), horse (Equus caballus), deer (Cervus elaphus), agouti (Dasyprocta aguti), donkey (Equus asinus), llama (Llama glama) and waterbuck (Kobus ellipsiprymnus) dung collected at the Reserva Ecológica de Dois Irmãos located in Recife, State of Pernambuco, Northeast Brazil. The samples were collected on a monthly basis from June 2005 to May 2006, taken to the laboratory and incubated in moist chambers. Higher number of taxa was observed in the excrements of tapir, followed by deer and donkey. The highest number of species was detected for Mucor, followed by Pilobolus. Statistical analyses showed significant differences in richness of Zygomycetes taxa between the herbivore dung types. Differences of species composition, however, were weak. Seasonality influenced the Zygomycetes species composition but not its richness. Variations in taxa composition between ruminants and non-ruminants dung were non significant.

Specific determination of myo-inositol in multivitamin pharmaceutical preparations by a flow injection system using a myo-inositol dehydrogenase reactor coupled with a glucose eliminating enzyme reactor.

A flow injection system for myo-inositol determination in multivitamin pharmaceutical preparations using two enzyme reactors was developed. Myo-inositol was detected using a fluorophotometer, to measure the fluorescence of NADH produced from NAD+ by a myo-inositol dehydrogenase reactor (IDR) containing myo-inositol dehydrogenase immobilized on porous glass. Enhanced interference due to excess glucose included in a multivitamin pharmaceutical preparation as a sweetener was eliminated by a glucose eliminating reactor (GER) co-immobilized with three enzymes (glucose oxidase, mutarotase and catalase). The calibration coefficient for the standard curve was 0.9993 for myo-inositol detection in the range of 1-5 microg/ml. Myo-inositol was determined even in the presence of glucose concentrations of 140-420 microg/ml. The recovery of myo-inositol added to the multivitamin pharmaceutical preparation was 99.6% (n=9).

A pratical method for screening for (beta)-galactosidase secreting microbial colonies

Microbial colonies were replicated on YNB© agar plates overlaid with soft agar containing the glucose-oxidase/peroxidase (BIOTROLr) system. The pink color developed around the colonies was the result of the reaction of this enzyme. This method proved to be very convenient for testing hundreds of colonies grown on agar plates for (beta)-galactosidase secretion by microbial cells.

Phospholipid specificity of bovine heart bc1 complex.

Bovine heart bc1 complex was reversibly inactivated by a new simple and effective chromatographic delipidation method. Upon phospholipid replenishment, catalytic activity increased from values near zero to values 2-6-times higher than those of the original preparation. Compared to original preparations maximally activated by additional phospholipid, the degree of reactivation was up to 100%. By this delipidation method, the 6.4-kDa protein subunit was removed with the phospholipid. The loss of this protein neither diminished electron transport activity nor abolished proton translocation. Two requirements were necessary to obtain quantitative data: (a) only bc1 complexes, homogeneously dissolved before and after relipidation had to be used and (b) the phospholipid bound to the complex had to be determined. The correlation of catalytic activity to bound phospholipid was studied in the range of low phospholipid/protein ratios, which had previously been insufficiently resolved. Catalytic activity increased linearly with added phospholipid up to a molar ratio of 80-100 lipid molecules/dimeric complex. This corresponds to the number of phospholipid molecules that complete a single bilayer annulus. The activating effect of phospholipid is not merely due to a hydrophobic phase effect, since it strongly depends on the nature of the polar head group of the added phospholipid. Of the three major phospholipids bound to the bc1 complex, only phosphatidylethanolamine and phosphatidylcholine activated when added as sole phospholipid. Tightly bound diphosphatidylglycerol was needed for preservation of the native complex structure.

An unusual polyanion from Physarum polycephalum that inhibits homologous DNA polymerase alpha in vitro.

From extracts of microplasmodia of Physarum polycephalum and their culture medium, an unusual substance was isolated which inhibited homologous DNA polymerase alpha of this slime mold but not beta-like DNA polymerase and not heterologous DNA polymerases. Analysis, especially NMR spectroscopy, revealed the major component to be an anionic polyester of L-malic acid and the inhibition to be due to poly(L-malate) in binding reversibly to DNA polymerase alpha. The mode of inhibition is competitive with substrate DNA and follows an inhibition constant Ki = 10 ng/mL. Inhibition is reversed in the presence of spermine, spermidine, poly(ethylene imine), and calf thymus histone H1. According to its ester nature, the inhibitor is slightly labile at neutral and instable at acid and alkaline conditions. Its largest size corresponds to a molecular mass of 40-50 kDa, but the bulk of the material after purification has lower molecular masses. The inhibitory activity depends on the polymer size and has a minimal size requirement.

Reactivation of creatine kinase activity by preincubation with buffered NAC diluent.

We have shown that several commercial control sera containing reversibly inactivated creatine kinase are reactivated by pre-incubation in N-acetyl cysteine assay diluent. The increase in activity can proceed for up to 4 h. This reactivation is not seen in patients' sera even after storage for 14 days at -20 degrees C, 7 days at 4 degrees C, or 2 days at 20 degrees C, during which time no loss of activity was evident. This study brings into question the validity of using consensus values to standardise enzyme assays.

Estandarización del inmunoanálisis enzimático / Standarization of the enzimatic immunoanalysis

Un método para poner de manifiesto anticuerpos específicos en suero es la variante indirecta de la técnica de ELISA, en la cual un problema habitual es la absorción inespecífica de las inmunoglobulinas del suero a la fase sólida del sistema, lo que dificulta la interpretación correcta de los resultados. En este trabajo se analiza la función que desempeñan varios factores involucrados en la técnica, y con base en los resultados obtenidos se proponen algunas recomendaciones generales para su buen uso

Role of cyclic adenosine monophosphate in rat lung development.

The objective of this study was to identify the biochemical mechanisms concerned with pulmonary growth and development. The data show that cyclic adenosine monophosphate (cAMP), adenylate cyclase, cAMP phosphodiesterase, and their regulation by intracellular modulators are important to the development of rat lungs. The presence in rat lung cytoplasm of factors modulating adenylate cyclase activity is described. These factors appear to be important physiologically as they are present in vivo, they appear in the cytoplasm at a specific age, and their activity is altered by diabetes and adrenalectomy and restored to original levels by administration of insulin and dexamethasone, respectively. The cytoplasmic activation of adenylate cyclase appears to be due to multiple proteins that can be resolved into less active components by DEAE-cellulose chromatography. Recombination of these proteins not only restored activity to the original level but actually resulted in more than additive activation, indicating some interdependence and positive cooperativity among the different components to maximally stimulate adenylate cyclase activity. The rat lung cytoplasmic activator protein regulates adenylate cyclase by a mechanism different from those reported for epinephrine, NaF, 5'-guanylimidophosphate, and calmodulin.

Reactivating factors of human alpha-mannosidase mutant.

In a previous study, we compared the alpha-mannosidase from mannosidosis tissues to that from normal one and we characterized the mutant. In this work, we show that the mutant inactivation by dialysis is reversible in different conditions and we investigate the nature of mannosidosis reactivating factors. The results obtained on pathological tissue by dialysis and by addition of dialysis fluid (DF), pronase treated DF, amino acid mixture, bivalent ions: Ca++, Zn++, Mg++, Co++ DF containing EDTA or DF heated to 600 degrees C suggest the reactivating factor includes both peptides and ions.