Effects of prenatal exposure to deltamethrin on forced swimming behavior, motor activity, and striatal dopamine levels in male and female rats

The effects of prenatal exposure of rat pups to 0.08 mg/kg deltamethrin (DTM) on physical, reflex and behavioral developmental parameters, on forced swimming and open-field behaviors, and on striatal monoamine levels at 60 days of age were observed. Maternal and offspring body weight, physical and reflex development were unaffected by the exposure to the pesticide. At 21 days of age, open-field locomotion frequency and immobility duration of male and female offspring were not different between control and exposed animals. However, male rearing frequency was increased in experimental animals. A decreased immobility latency to float and in general activity after the swimming test in male offspring was observed at adult age; no interference was detected in the float duration during the swimming test. In addition, these animals presented higher striatal 3,4-dihydroxyphenylacetic acid (DOPAC) levels without modification in dopamine (DA) levels and an increased DOPAC/DA ratio. These data indicate a higher activity of the dopaminergic system in these animals. Noradrenaline (NA) levels were increased, while MHPG levels were not detectable in the system studied. Serotonin (5-HT) and 5-hydroxyindolacetic acid (5-HIAA) levels, as well as the homovanillic acid (HVA)/DA ratio, were not modified by the exposure to the pesticide. No changes were observed in swimming and open-field behaviors nor were there any changes in striatal monoamines or their metabolites in the female experimental group. In relation to the pesticide formula, the present data showing that prenatal exposure to DTM alters latency to float and the activity of striatal dopaminergic system might reflect a persistent effect of the pesticide on animal motor activity, mainly in males. on the other hand, the decrease in general activity observed in experimental male rats suggests higher levels of emotionality induced by previous exposure to the swimming behavior test in relation to control animals. Data gathered in the present study may be important for the assessment of the safety of pyrethroid insecticides. (C) 2001 Elsevier B.V. All rights reserved.

Chlorophyll fluorescence as a marker for herbicide mechanisms of action

Photosynthesis is the single most important source of 02 and organic chemical energy necessary to support all non-autotrophic life forms. Plants compartmentalize this elaborate biochemical process within chloroplasts in order to safely harness the power of solar energy and convert it into usable chemical units. Stresses (biotic or abiotic) that challenge the integrity of the plant cell are likely to affect photosynthesis and alter chlorophyll fluorescence. A simple three-step assay was developed to test selected herbicides representative of the known herbicide mechanisms of action and a number of natural phytotoxins to determine their effect on photosynthesis as measured by chlorophyll fluorescence. The most active compounds were those interacting directly with photosynthesis (inhibitors of photosystem I and II), those inhibiting carotenoid synthesis, and those with mechanisms of action generating reactive oxygen species and lipid peroxidation (uncouplers and inhibitors of protoporphyrinogen oxidase). Other active compounds targeted lipids (very-long-chain fatty acid synthase and removal of cuticular waxes). Therefore, induced chlorophyll fluorescence is a good biomarker to help identify certain herbicide modes of action and their dependence on light for bioactivity. Published by Elsevier B.V.

Characterization of glycerol kinase from baker's yeast: Response surface modeling of the enzymatic reaction

The present study describes a methodology of dosage of glycerol kinase (GK) from baker's yeast. The standardization of the activity of the glycerol kinase from baker's yeast was accomplished using the diluted enzymatic preparation containing glycerol phosphate oxidase (GPO) and glycerol kinase. The mixture was incubated at 60 degrees C by 15 min and the reaction was stopped by the SDS solution addition. A first set of experiments was carried out in order to investigate the individual effect of temperature (7), pH and substrate concentration (S), on GK activity and stability. The pH and temperature stability tests showed that the enzyme presented a high stability to pH 6.0-8.0 and the thermal stability were completely maintained up to 50 degrees C during 1 h. The K(m) of the enzyme for glycerol was calculated to be 2 mM and V(max) to be 1.15 U/mL. In addition, modeling and optimization of reaction conditions was attempted by response surface methodology (RSM). Higher activity values will be attained at temperatures between 52 and 56 degrees C, pH around 10.2-10.5 and substrate concentrations from 150 to 170 mM.This low cost method for glycerol kinase dosage in a sequence of reactions is of great importance for many industries, like food, sugar and alcohol. RSM showed to be an adequate approach for modeling the reaction and optimization of reaction conditions to maximize glycerol kinase activity. (C) 2007 Elsevier B.V. All rights reserved.

Crystal structure of a novel myotoxic Arg49 phospholipase A(2) homolog (zhaoermiatoxin) from Zhaoermia mangshanensis snake venom: Insights into Arg49 coordination and the role of Lys122 in the polarization of the C-terminus

The venom of Zhaoermia mangshanensis, encountered solely in Mt Mang in China's Hunan Province, exhibits coagulant, phosphodiesterase, L-amino acid oxidase, kallikrein, phospholipase A(2) and myotoxic activities. The catalytically inactive PLA(2) homolog referred to as zhaoermiatoxin is highly myotoxic and displays high myonecrotic and edema activities. Zhaoermiatoxin possesses a molecular weight of 13,972 Da, consists of 121 amino-acid residues crosslinked by seven disulfide bridges and shares high sequence homology with Lys49-PLA(2)s from the distantly related Asian pitvipers. However, zhaoermiatoxin possesses an arginine residue at position 49 instead of a lysine, thereby suggesting a secondary Lys49 -> Arg substitution which results in a catalytically inactive protein. We have determined the first crystal structure of zhaoermiatoxin, an Arg49-PLA(2), from Zhaoermia mangshanensis venom at 2.05 A resolution, which represents a novel member of phospholipase A(2) family. In this structure, unlike the Lys49 PLA(2)s, the C-terminus is well ordered and an unexpected non-polarized state of the putative calcium-binding loop due to the flip of Lys122 towards the bulk solvent is observed. The orientation of the Arg-49 side chain results in a similar binding mode to that observed in the Lys49 PLA(2)s; however, the guadinidium group is tri-coordinated by carbonyl oxygen atoms of the putative calcium-binding loop, whereas the N zeta atom of lysine is tetra-coordinated as a result of the different conformation adopted by the putative calcium-binding loop. (c) 2008 Elsevier Ltd. All rights reserved.

SISTEMAS QUIMILUMINESCENTES COM PEROXIDASE (EC:1.11.1.7) E SUAS APLICACOES EM ANALISES CLINICAS

The enzyme horseradish peroxidase HRP (EC:1.11.1.7), has both acid and basic isoenzymes, catalyses a wide range of reactions (acting as an oxiredutase or an oxidase) and is thought capable of one- or two-electrons oxidations depending on the substrate employed. Today, the methodology for these assay can be chemiluminescent reactions and enhanced chemiluminescent. The enhanced chemiluminescent assay with system HRP, luminol, peroxide and an enhancer has provided the basis for a convenient and sensitive assay for peroxidase and peroxidase conjugates, DNA probe and blotting assay. It is particularly more advantageous than the others, because is very rapid, more sensitive (attomoles), easy to do and technically simple, and is relatively specific for HRP (reduces the effect of the interference).

Use of sorghum seed tissue as a biocatalyst in a stirred reactor for oxalic acid determination

The enzyme oxalate oxidase, E.C. 1.2.3.4 from Sorghum vulgare seeds (variety BR303) was used to develop a new sensor for oxalate determination without any purification. The sorghum seeds were conditioned in a 0.10 mol I-1 KCl solution. Then, these seeds were put in a stirring bar type enzymic reactor and coupled with an electrode for CO2. This device was introduced into a cell containing 10.0 ml of a 0.10 mol I-1 KCl solution saturated with oxygen. This sensor showed a linear response between 1.0 and 4.0 × 10-3 mol I-1 with a slope of 30 mV per decade of oxalate concentration at 25.0°C. The sensor was stable for one month or 200 determinations. The response time was about 60 s. The Michaelis-Menten constant determined for this enzyme was 1.5 × 10-3 mol I-1.

Determination of Oxalate in Grass by FIA with a Spectrophotometric Detection System Using a Two Enzyme Reactor

A new methodology for soluble oxalic acid determination in grass samples was developed using a two enzyme reactor in an FIA system. The reactor consisted of 3 U of oxalate oxidase and 100 U of peroxidase immobilized on Sorghum vulgare seeds activated with glutaraldehyde. The carbon dioxide was monitored spectrophotometrically, after reacting with an acid-base indicator (Bromocresol Purple) after it permeated through a PTFE membrane. A linear response range was observed between 0.25 and 1.00mmol l-1 of oxalic acid; the data was fit by the equation A=-0.8(±1.5)+ 57.2(±2.5)[oxalate], with a correlation coefficient of 0.9971 and a relative standard deviation of 2% for n=5. The variance for a 0.25 mmol l-1 oxalic acid standard solution was lower than 4% for 11 measurements. The FIA system allows analysis of 20 samples per hour without prior treatment. The proposed method showed a good correlation with that of the Sigma Kit.

Oxalate determination in urine using an immobilized enzyme on Sorghum vulgare seeds in a flow injection conductimetric system

A flow-injection (FI) method was developed for the determination of oxalate in urine. It was based on the use of oxalate oxidase (E.C. 1.2.3.4) immobilized on ground seeds of the BR-303 Sorghum vulgare variety. A reactor was filled with this activated material, and the samples (200 μL) containing oxalate were passed through it, carried by a deionized water flow. The carbon dioxide produced by the enzyme reaction permeated through a microporous PTFE membrane, and was received in a water acceptor stream, promoting conductivity changes proportional to the oxalate concentration in the sample. The results obtained showed a useful linear range from 0.05 to 0.50 mmol dm-3. The proposed method, when compared with the Sigma enzymatic procedure, showed good correlation (Y = 0.006(±0.016) + 0.98(±0.019)X; r = 0.9995, Y = conductivity in μS, and X = concentration in mmol dm-3), selectivity, and sensitivity. The new immobilization approach promotes greater stability, allowing oxalate determination for 6 months. About 13 determinations can be performed per hour. The precision of the proposed method is about ± 3.2 % (r.s.d).

Enzymes in the hypopharyngeal gland extracts from workers of Scaptotrigona postica (Hymenoptera, Apinae, Meliponini) related to food storing in the colony

This study reports on research of enzymes produced by the hypopharyngeal glands, which are related to food storing in the colony, from gland extracts from nurse and forager workers of S. postica. Only the presence of the saccharase was detected in the extracts from the glands of forager workers. The results were compared to the enzymatic content of similar extracts of A. mellifera taking into account the behavioral differences among the two species.

The influence of diabetes mellitus and insulin therapy on biomechanical retention around dental implants: a study in rabbits.

The oral rehabilitation by dental implants in patients with diabetes remains a controversial issue. The aim of this study was to evaluate the influence of diabetes mellitus and insulin therapy on the bone healing around dental implants using torque removal. Twenty-seven rabbits were randomly divided into 3 groups with 9 animals each: control (C) group, induced diabetic (D) group, and insulin-treated diabetic (ITD) group (10 U/day). After 1 week, one implant was inserted at the tibial metaphysis of the animals. The glucose levels were periodically evaluated through the glucose-oxidase enzymatic method. The animals were killed at 4, 8, and 12 weeks after surgery and the biomechanical test was performed using a torque manometer. Statistically significant differences regarding the removal torque of the implant could not be found at 4 weeks (P = 0.2) among groups. Group C showed statistically higher values than groups D and ITD at the experimental periods of 8 (P = 0.0001 and P = 0.0002, respectively) and 12 weeks (P = 0.0053 and P = 0.001, respectively). There were no statistical differences between D and ITD groups in any of the experimental periods. Diabetes mellitus has negatively influenced the mechanical retention of implants placed at the tibial metaphysis of rabbits. Therapy with insulin did not induce any changes.

Use of carambola (Averrhoa carambola L. 'Fwang Tung') fruit at two stages of maturity for fresh-cut products

Carambola fruit ('Fwang Tung') were picked at two stages of maturity: mature-green (50% yellow) and mature (100% yellow). Fruit were washed with water, dipped in NaOCl solution (200 mg.L-1 for 5 minutes), and stored over night at 10°C. Fruit were sliced manually in to pieces of approximately 1 cm thickness. Slices were rinsed with NaOCl solution at 20 mg.L-1, drained for 3 minutes, and packaged in polyethylene tereftalate (PET) trays provided with a fit cover (Neoform® N94). Packages were stored at 6.5°C and 85% RH for 9 days, and samples taken every 3 days for physical, chemical and biochemical analysis, respiration, and internal atmosphere composition. Immediately after cutting, slices at both stages of maturity showed a wounding response with a 5-fold increase in respiration rate. Polygalacturonase (PG) and polyphenol oxidase (PPO) activity did not differ between stages of maturity. Despite the less mature stage being less preferred at the sensory evaluation owing to its greenish peel, the best stage of maturity for carambola fresh-cut production was mature-green, due to a higher resistance to cutting, and presenting a better colour and appearance maintenance for up to 9 days.

COX24 codes for a mitochondrial protein required for processing of the COX1 transcript

In most strains of Saccharomyces cerevisiae the mitochondrial gene COX1, for subunit 1 of cytochrome oxidase, contains multiple exons and introns. Processing of COX1 primary transcript requires accessory proteins factors, some of which are encoded by nuclear genes and others by reading frames residing in some of the introns of the COX1 and COB genes. Here we show that the low molecular weight protein product of open reading frame YLR204W, for which we propose the name COX24, is also involved in processing of COX1 RNA intermediates. The growth defect of cox24 mutants is partially rescued in strains harboring mitochondrial DNA lacking introns. Northern blot analyses of mitochondrial transcripts indicate cox24 null mutants to be blocked in processing of introns aI2 and aI3. The dependence of intron aI3 excision on Cox24p is also supported by the growth properties of the cox24 mutant harboring mitochondrial DNA with different intron compositions. The intermediate phenotype of the cox24 mutant in the background of intronless mitochondrial DNA, however, suggests that in addition to its role in splicing of the COX1 pre-mRNA, Cox24p still has another function. Based on the analysis of a cox14-cox24 double mutant, we propose that the other function of Cox24p is related to translation of the COX1 mRNA. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

Amphibian secretions for drug discovery studies: A search for new antiparasitic and antifungal compounds

The antiparasitic and antifungal activities of nine amphibian skin secretions were studied in different in vitro models. Seven secretions presented a considerable antiprotozoan activity and one showed promising results against Candida sp. These results can be the basis for the development of new drugs, especially for neglected parasitic diseases. © 2007 Bentham Science Publishers Ltd.

Nuclear mitochondrial-like sequences in ants: Evidence from Atta cephalotes (Formicidae: Attini)

Nuclear mitochondrial-like sequences (numts) are copies of mitochondrial DNA that have migrated to the genomic DNA. We present the first characterization of numts in ants, these numts being homologues to a mitochondrial DNA fragment containing loci the 3′ portion of the cytochrome oxidase I gene, an intergenic spacer, the tRNA leucine gene and the 5′ portion of the cytochrome oxidase II gene. All 67 specimens of Atta cephalotes (Hymenoptera: Formicidae: Attini) investigated had these homologues, which are within two monophyletic groups that we called numt1 and numt2. Numt1 and numt2 sequences are less variable than mitochondrial sequences and released from the severe purifying selection constraining the evolution of mitochondrial genes. Their formation probably involved bottlenecks related to two distinct transfer events of ancient and fast evolving mitochondrial DNA fragments to comparative slowly evolving nuclear DNA regions. © 2007 The Authors.

Transmission of Aggregatibacter actinomycetemcomitans between Brazilian women with severe chronic periodontitis and their children

This study evaluated the transmission of Aggregatibacter actinomycetemcomitans (Aa) in women with severe chronic periodontitis and their children. Thirty women (mean age = 36.1±6.0 years) who were mothers of at least one child aged 7 to 16 years were enrolled. In order to investigate mother-child transmission of Aa, the children were also evaluated when their mothers were colonized by the bacterium. Subgingival plaque samples of each woman were collected from 3 sites (mean probing depth of 7.3±1.2 mm and mean clinical attachment level of 7.9±1.5 mm) and pooled in reduced transport fluid (RTF). These samples were processed, inoculated onto TSBVagar selective medium and incubated at 37°C in microaerophilic atmosphere for 5 days. Aa was identified on the basis of colony morphology, Gram staining, catalase and oxidase reactions. Aa was found in 8 out of 30 women. Therefore, 8 children from these women (mean age= 12 ± 3.7 years) were evaluated, but Aa was found only in 2 of them. Aa strains of the two mother-child pairs were evaluated by arbitrarily-primed polymerase chain reaction (AP-PCR), although it was not found similarity between the amplitypes of each pair. No Aa transmission was found between Brazilian women with severe chronic periodontitis and their children.