Characterization of a new platelet aggregating factor from crotoxin Crotalus durissus cascavella venom

In this article we investigated the platelet aggregating activity of whole crotoxin and its subunits isolated from Crotalus durissus cascavella venom. During the purification protocols of the venom, using HPLC molecular exclusion, we detected the presence of two different serine protease activities in the gyroxin fraction, and another in the crotoxin fraction, which induced strong and irreversible platelet aggregation, in addition to blood coagulation. From crotoxin, we isolated PLA(2), crotapotin (both fractions corresponding approximately 85% of whole crotoxin) and another minor fraction (F20) that exhibited serine protease activity. After a new fractionation on reverse phase HPLC chromatography, we obtained three other fractions named as F201, F202 and F203. F202 was obtained with high degree of molecular homogeneity with molecular mass of approximately 28 kDa and a high content of acidic amino residues, such as aspartic acid and glutamic acid. Other important amino acids were histidine, cysteine and lysine. This protein exhibited a high specificity for BApNA, a Michaelis-Menten behavior with Vmax estimated in 5.64 mu M/min and a Km value of 0.58 mM for this substrate. In this work, we investigated the ability of F202 to degrade fibrinogen and observed alpha and beta chain cleavage. Enzymatic as well as the platelet aggregation activities were strongly inhibited when incubated with TLCK and PMSF, specific inhibitors of serine protease. Also, F202 induced platelet aggregation in washed and platelet-rich plasma, and in both cases, TLCK inhibited its activity. The N-terminal amino acid sequence of F202 presented a high amino acid sequence homology with other thrombin-like proteins, but it was significantly different from gyroxin. These results showed that crotoxin is a highly heterogeneous protein composed of PLA(2), thrombin-like and other fractions that might explain the diversity of physiological and pharmacological activities of this protein.

Flow-injection spectrophotometric determination of dipyrone in pharmaceutical formulations using ammonium molybdate as chromogenic reagent

A simple and rapid flow-injection spectrophotometric method is reported for the determination of dipyrone in pharmaceutical formulations. The method is based on the reaction of dipyrone with ammonium molybdate in acidic medium to produce blue molybdenum, which was detected spectrophotometrically at 620 nm. The analyte was determined in a single-line flow system. The calibration curve obtained was linear in the range of 5x10(-4) to 8x10(-3) mol L-1 for dipyrone concentration and the precision ( s r =1.7%) was satisfactory. The method proved to be selective and adequately sensitive. Application of the method to the analysis of pharmaceutical samples resulted in excellent accuracy; the percent mean recoveries were in the range 95.3%-101% and relative errors less than 5.0% for five pharmaceutical formulations were found.

The role of indomethacin and tezosentan on renal effects induced by Bothrops moojeni Lys49 myotoxin I

Renal changes determined by Lys49 myotoxin I (BmTx I), isolated from Bothrops moojeni are well known. The scope of the present study was to investigate the possible mechanisms involved in the production of these effects by using indomethacin (10 mu g/mL), a non-selective inhibitor of cyclooxygenase, and tezosentan (10 mu g/mL), an endothelin antagonist. By means of the method of mesenteric vascular bed, it has been observed that B. moojeni myotoxin (5 mu g/mL) affects neither basal perfusion pressure nor phenylephrine-preconstricted vessels. This fact suggests that the increase in renal perfusion pressure and in renal vascular resistance did not occur by a direct effect on renal vasculature. Isolated kidneys from Wistar rats, weighing 240-280 g, were perfused with Krebs-Henseleit solution. The infusion of BmTx-I increased perfusion pressure, renal vascular resistance, urinary flow and glomerular filtration rate. Sodium, potassium and chloride tubular transport was reduced after addition of BmTx-I. Indomethacin blocked the effects induced by BmTx-I on perfusion pressure and renal vascular resistance, however, it did not revert the effect on urinary flow and sodium, potassium and chloride tubular transport. The alterations of glomerular filtration rate were inhibited only at 90 min of perfusion. The partial blockade exerted by indomethacin treatment showed that prostaglandins could have been important mediators of BmTx-I renal effects, but the participation of other substances cannot be excluded.The blockage of all renal alterations observed after tezosentan treatment support the hypothesis that endothelin is the major substance involved in the renal pathophysiologic alterations promoted by the Lys49 PLA(2) myotoxin I, isolated from B. moojeni. In conclusion, the rather intense renal effects promoted by B. moojeni myotoxin-I were probably caused by the release of renal endothelin, interfering with the renal parameters studied. (c) 2006 Elsevier Ltd. All rights reserved.

A field study to describe diel, tidal and semilunar rhythms of larval release in an assemblage of tropical rocky shore crabs

Available information on the larval release rhythms of brachyurans is biased to temperate estuarine species and outcomes resulting from some sort of artificial manipulation of ovigerous females. In this study we applied field methods to describe the larval release rhythms of an assemblage of tropical rocky shore crabs. Sampling the broods of ovigerous females of Pachygrapsus transversus at two different shores indicated a spatially consistent semilunar pattern, with larval release maxima around the full and new moon. Yet, synchronism between populations varied considerably, with the pattern obtained at the site exposed to a lower wave action far more apparent. Breeding cohorts at one of the sampled shores apparently belonged to actual age groups composing the ovigerous population. The data suggest that these breeding groups release their larvae in alternate syzygy periods, responding to a lunar cycle instead of the semilunar pattern observed for the whole population. For the description of shorter-term rhythms, temporal series at hour intervals were obtained by sampling the plankton and confinement boxes where ovigerous females were held. Unexpectedly, diurnal release activity prevailed over nocturnal hatching. Yet, only grapsids living higher on the shore exhibited strong preferences over the diel cycle, with P. transversus releasing their larvae during the day and Geograpsus lividus during the night. The pea crab Dissodactylus crinitichelis, the spider crab Epialtus brasiliensis and a suite of xanthoids undertook considerable releasing activity in both periods. Apart from the commensal pea crab D. crinitichelis, all other taxa revealed tide-related rhythms of larval release, with average estimates of the time of maximum hatching always around the time of high tides; usually during the flooding and slack, rather than the ebbing tide. Data obtained for P. transversus females held in confinement boxes indicated that early larval release is mostly due to nocturnal hatching, while zoeal release in diurnal groups took place at the time of high tide. Since nocturnal high tides at the study area occurred late, sometimes close to dusk, early release would allow more time for offshore transport of larvae when the action of potential predators is reduced.

Population structure and growth of the hermit crab Pagurus brevidacylus (Anomura : Paguridae) from the northern coast of São Paulo, Brazil

The hermit crab Paguras brevidactylus (Crustacea: Anomura: Paguridea) from the infralittoral area of Anchieta Island, Ubatuba, was characterized by population Structure (size, sex ratio, reproduction and recruitment) and growth. Animals were collected monthly during 1999 by SCUBA diving. A total of 1525 individuals was collected (633 males and 892 females), 695 of them were ovigerous females. Overall sex ratio was 0.7:1 in favour of females. The crabs showed a unimodal distribution with males significantly larger than females. Ovigerous females were collected during all months and in high percentages from 1.0 mm of shield length, demonstrating intense and Continuous reproduction. The longevity was approximately 24 months for males and 18 for females, which showed larger growth rate and reached sexual maturity earlier (two months) than males. The low number of males in this Population may be due to the longer life span. Moreover, the sexual dimorphism favours males during the intra- and interspecific fights by shell, food, reproduction and territory. Females demonstrated a short life cycle and intense reproduction.

Whistle variability in South Atlantic spinner dolphins from the Fernando de Noronha Archipelago off Brazil

A series of quali- and quantitative analyses were conducted to evaluate the variability of spinner dolphin whistles from the Fernando de Noronha Archipelago off Brazil. Nine variables were extracted from each whistle contour, and the whistle contours shapes were classified into the seven categories described in Driscoll (1995). The analysis showed mean beginning and ending frequencies values of 10.78 and 12.74 kHz, respectively. on average, whistle duration was relatively short, with mean values around 0.495 s (N=702). Comparative analyses were also conducted to investigate the relationship between the obtained results and those presented in previous studies. When comparing averages, the results of the study of Oswald et al. (2003) in the Tropical Eastern Pacific (TEP) presented less significant differences in relation to this study; only whistle duration differed significantly between both works. The results of multivariate classification tests also pointed TEP population as the closest related to the population studied here. The similarities between such disjunct populations might be attributed to a more recent isolation event (the closing of the Panama Isthmus) than the divergence that has driven North and South Atlantic populations apart. (c) 2006 Acoustical Society of America.

Biological and structural characterization of a new PLA(2) from the Crotalus durissus collilineatus venom

In the present article we report on the biological characterization and amino acid sequence of a new basic Phospholipases A(2) (PLA(2)) isolated from the Crotalus durissus collilineatus venom (Cdcolli F6), which showed the presence of 122 amino acid residues with a pI value of 8.3, molecular mass of 14 kDa and revealed an amino acid sequence identity of 80% with crotalic PLA(2)s such as Mojave B, Cdt F15, and CROATOX. This homology, however, dropped to 50% if compared to other sources of PLA(2)s such as from the Bothrops snake venom. Also, this PLA(2) induced myonecrosis, although this effect was lower than that of BthTx-I or whole crotoxin and it was able to induce a strong blockage effect on the chick biventer neuromuscular preparation, independently of the presence of the acid subunid (crotapotin). The neurotoxic effect was strongly reduced by pre-incubation with heparin or with anhydrous acetic acid and rho-BPB showed a similar reduction. The rho-BPB did not reduce significantly the myotoxic activity induced by the PLA(2), but the anhydrous acetic acid treatment and the pre-incu-bation of PLA(2) with heparin reduced significantly its effects. This protein showed a strong antimicrobial activity against Xanthomonas axonopodis passiflorae (Gram-negative), which was drastically reduced by incubation of this PLA(2) with rho-BPB, but this effect was marginally reduced after treatment with anhydrous acetic acid. Our findings here allow to speculate that basic amino acid residues on the C-terminal and molecular regions near catalytic site regions such as Calcium binding loop or rho-wing region may be involved in the binding of this PLA(2) to the molecular receptor to induce the neurotoxic effect. The bactericidal effect, however, was completely dependent on the enzymatic activity of this protein.

Sensor for cysteine based on oxovanadium(IV) complex of Salen modified carbon paste electrode

The electrochemical behavior of a carbon paste electrode modified (CPEM) with N,N′-ethylenebis(salicylideneiminato)oxovanadium(IV) complex ([(VO)-O-IV(Salen)]) was investigated as a new sensor for cysteine. Cyclic voltammetry at the modified electrode in 0.1 mol L-1 KCl Solution (pH 5.0) showed a single-electron reduction/oxidation of the Couple VO3+/VO2+. The CPEM with [VO(Salen)] presented good electrochemical stability in a wide pH range (4.0-10.0) and an ability to electrooxidate cysteine at 0.65 V versus SCE. These results demonstrate the viability of the use of this modified electrode as an amperometric sensor for cysteine determination. © 2004 Elsevier B.V. All rights reserved.

Influence of a Brazilian sewage outfall on the toxicity and contamination of adjacent sediments

The submarine sewage outfall of Santos (SSOS) is situated in the Santos Bay (São Paulo, Brazil) and is potentially a significant source of contaminants to the adjacent marine ecosystem. The present study aimed to assess the influence of SSOS on the sediment toxicity and contamination at Santos Bay. At the disposal site, sediments tended to be finer, organically richer and exhibited higher levels of surfactants and metals, sometimes exceeding the Threshold Effect Level values. The SSOS influence was more evident toward the East, where the sediments exhibited higher levels of TOC, total S and metals during the summer 2000 sampling campaign. Sediment toxicity to amphipods was consistently detected in four of the five stations studied. Amphipod survival tended to correlate negatively to Hg, total N and % mud. This work provides evidence that the SSOS discharge affects the quality of sediments from Santos Bay, and that control procedures are warranted. (c) 2005 Elsevier Ltd. All rights reserved.

Longitudinal distribution and lateral pattern of megalopal settlement and juvenile recruitment of Carcinus maenas (L.) (Brachyura, Portunidae) in the Mira River Estuary, Portugal

Settlement is a critical process in the life history of crabs, and thus affecting the abundance, distribution and structure of estuarine communities. The spatial pattern of settlement of megalopae of the shore crab Carcinus maenas along a longitudinal estuarine gradient (Mira River Estuary, Portugal) was examined, as well as its effects on the juvenile population. To measure megalopal settlement, four replicate collectors were deployed in six equally spaced stations along the estuarine axis. Juveniles were collected on the same locations with a quadrat randomly deployed on the substrate. To assess fine-scale megalopal settlement within a curved region of the estuary, replicate collectors were deployed on both margins along Moinho da Asneira curve. Megalopae settled differently along the six longitudinal points, with a tendency to attenuate their settlement upstream. Within the curved region, megalopae preferentially settled on the left margin collectors, probably due to the weaker velocity speeds felt on this margin. Concerning the overall juvenile density, there were significant differences among the stations distributed along the estuary, but they did no reflect a longitudinal dispersion attenuation pattern. Size-frequency distribution of the juvenile population showed that the average size is higher on the left margin. Recruits (carapace length between 1.0 mm and 3.4 mm) were more abundant on the upstream stations. Density of early juveniles (3.4 mm-6.5 mm) and juveniles (6.5 mm-10 mm) was more stable throughout the estuary axis than that of recruits. This distribution pattern may result from tidal excursion processes or mechanisms to avoid biotic interactions, such as predation and competition. (c) 2006 Elsevier Ltd. All rights reserved.

Is the meander growth in the Brazil Current system off Southeast Brazil due to baroclinic instability?

Temporally-growing frontal meandering and occasional eddy-shedding is observed in the Brazil Current (BC) as it flows adjacent to the Brazilian Coast. No study of the dynamics of this phenomenon has been conducted to date in the region between 22 degrees S and 25 degrees S. Within this latitude range, the flow over the intermediate continental slope is marked by a current inversion at a depth that is associated with the Intermediate Western Boundary Current (IWBC). A time series analysis of 10-current-meter mooring data was used to describe a mean vertical profile for the BC-IWBC jet and a typical meander vertical structure. The latter was obtained by an empirical orthogonal function (EOF) analysis that showed a single mode explaining 82% of the total variance. This mode structure decayed sharply with depth, revealing that the meandering is much more vigorous within the BC domain than it is in the IWBC region. As the spectral analysis of the mode amplitude time series revealed no significant periods, we searched for dominant wavelengths. This search was done via a spatial EOF analysis on 51 thermal front patterns derived from digitized AVHRR images. Four modes were statistically significant at the 95% confidence level. Modes 3 and 4, which together explained 18% of the total variance, are associated with 266 and 338-km vorticity waves, respectively. With this new information derived from the data, the [Johns, W.E., 1988. One-dimensional baroclinically unstable waves on the Gulf Stream potential vorticity gradient near Cape Hatteras. Dyn. Atmos. Oceans 11, 323-350] one-dimensional quasi-geostrophic model was applied to the interpolated mean BC-IWBC jet. The results indicated that the BC system is indeed baroclinically unstable and that the wavelengths depicted in the thermal front analysis are associated with the most unstable waves produced by the model. Growth rates were about 0.06 (0.05) days(-1) for the 266-km (338-km) wave. Moreover, phase speeds for these waves were low compared to the surface BC velocity and may account for remarks in the literature about growing standing or stationary meanders off southeast Brazil. The theoretical vertical structure modes associated with these waves resembled very closely to the one obtained for the current-meter mooring EOF analysis. We interpret this agreement as a confirmation that baroclinic instability is an important mechanism in meander growth in the BC system. (C) 2008 Elsevier B.V. All rights reserved.

Checklist dos mamíferos do Estado de São Paulo, Brasil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Are deep-sea cephalopods really common preys for oceanic seabirds?

Uma análise de dados publicados sobre dietas de aves marinhas oceânicas mostra a predominância de cefalópodes musculares e de distribuição mais superficial nas camadas oceânicas, mas também são importantes as espécies gelatinosas e amoniacais restritas a camadas abaixo dos 300 m da superfície. A princípio, não deveria se esperar que cefalópodes de profundidade fossem considerados presas comuns de aves marinhas oceânicas como reportados por muitos autores. É proposto neste estudo que uma fonte indireta, importante e de fácil obtenção, surgiu com o início das atividades dos barcos atuneiros que operam com espinhel. O hábito de ingerir restos de vísceras de peixes capturados em barcos espinheleiros pode explicar as prováveis conclusões equivocadas de que cefalópodes de profundidade são presas naturais de aves marinhas oceânicas.

Anti-Helicobacter pylori activity and oxidative burst inhibition by the naphthoquinone 5-methoxy-3,4-dehydroxanthomegnin from Paepalanthus latipes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Play behaviour in nonhuman animals and the animal welfare issue

The mission of defining animal welfare indicators is methodologically difficult, limited, and possibly impossible. A promising alternative, however, to evaluate suitable environmental conditions is the assessment of play behaviour. In the present review, we summarise the general aspects of play behaviour in nonhuman animals and propose its use as a potential indicator of animal welfare. Play behaviour probably occurs in most vertebrates and some invertebrates, but predominately in mammals. It is also more frequent in young males and is associated with the environmental context in which animals find themselves. Animals play if they are healthy and well-fed, but not if they are under stressful conditions or if they are in a stressful state. We can therefore use the prevalence of play behaviour as an indicator of suitable environmental conditions, considering the specificity associated with the above-mentioned modifying factors.