Journal of natural toxins最新文献

A simple, rapid, and sensitive diagnostic kit for detecting Thai cobra (Naja kaouthia) venom was developed using latex particles sensitized with venom specific immunoglobulin. The kit is capable of detecting 25-50 ng/ml of Thai cobra venom. The capability was not affected by human plasma. Specificity of the kit was proven using snake venoms from Vipera russelli, Calloselasma rhodostoma, Trimeresurus albolabris, Naja siamensis, Ophiophagus hannah, and Bungarus fasciatus. The diagnostic kit does not lose its capability under refrigeration for two months and by lyophilization.

Research on natural products provides not only knowledge and understanding of living organisms but chemical entities obtained from bioresources can also be used as structural models for further development in the various fields, notably in the agricultural and pharmaceutical sciences. Selected examples on the scientific aspects of traditional usage of bioresources are described.

Acetylcholine receptor (AChR) binding activity was detected in Duvernoy's secretions from B. blandingi and B. dendrophila as they competitively inhibited formation of 3[H]bungarotoxin-acetylcholine receptor complexes (3[H]Bgt-AChR) in a concentration-dependent manner. Secretions contained two types of toxin: low affinity and high affinity. Reversed-phase HPLC of B. blandingi and B. dendrophila secretions afforded 20 and 14 peaks, respectively. AChR binding components, as revealed by SDS-PAGE, had apparent molecular weights of 10 and 11 kDa (B. blandingi) or 11 and 12 kDa (B. dendrophila). Periodic acid-Schiff staining indicated these were not glycoproteins. Alkylation with 4-vinylpyridine significantly decreased their ability to inhibit 3[H]Bgt-AChR binding, indicating disulphide bridges were necessary for receptor-binding activity. Attempts to sequence the B. blandingi peptides were negative as these components seemed to be N-terminally blocked.

The effect of bolus intravenous injection of sub-LD50 (35 micrograms Kg-1) of the neurotoxic fraction (F3) of the Egyptian cobra Naja haje on the plasma level of ACTH and serum levels of cortisol, insulin, glucose, total lipids, triacylglycerols, free fatty acids, total cholesterol, HDL and LDL-cholesterol, and glycogen content of liver and kidneys were studied in rabbit pretreated with cyproteron acetate (CA) or saline solution and propylene glycol (PG) to elucidate the possible role of the hypothalamo-pituitary adrenal (HPA) axis in the venom fraction-induced hyperglycemia. F3 increased cortisol and insulin level in both groups, whereas ACTH was found to decrease subsequent to the treatment. Serum glucose level was elevated by F3 treatment and this effect was substantiated in CA-treated rabbits. This hyperglycemia was concomitant with a decline in glycogen content of the liver and kidneys. A decline in serum total lipids, triacylglycerols, and free fatty acids was observed following F3 treatment, and this effect was intensified by CA-pretreatment. These data suggest that F3 stimulates glucocorticoid release from adrenocortical cells which, in turn, may modulate both insulin and glucose turnover to maintain hyperglycemia during stress period. The possible underlying mechanisms were discussed.

This paper describes the involvement of sex hormones in the edematogenic response produced by staphylococcal enterotoxin B (SEB) in the mouse hindpaw. Both the paw weight variation and the protein exudate produced by the intraplantar administration of SEB (12.5 micrograms/paw) to intact, randomly cycling female (IRCF) mice were significantly attenuated when the animals were ovariectomized (OVX). The attenuation of SEB-induced paw swelling produced by OVX was not reversed by estradiol (OE2) reposition. Thus, 4 h after the injection of SEB the increase in paw weight in OVX-mice treated with OE2 (10 micrograms/kg in corn oil) was 15.0 +/- 0.9 mg, while the exudation corresponded to 2.1 +/- 0.3 micrograms of Evans blue dye/g of tissue. Neither of these values differed significantly from those obtained 4 h after the intraplantar injection of SEB (12.5 micrograms/paw) in non-treated OVX-mice (paw swelling, 14.0 +/- 0.8 mg; dye exudation, 2.0 +/- 0.3 micrograms/g, N = 6). Pretreating IRCF mice once a day for three days with human chorionic gonadotrophin (40 IU/kg, i.m.) reduced the paw edema produced by the toxin, thus indicating an involvement of gonadotrophins in this event. A pronounced decrease in paw weight variation (about 45%) and dye exudation (61%) was detected when IRCF mice were previously treated every 72 h with three injections of OE2 (10 micrograms/kg in corn oil, i.m.). Similar situations were also seen when the animals were pretreated at 72 h intervals with three injections of testosterone (10 mg/kg in corn oil, i.m.). We conclude that the paw edema induced by SEB in female mice is hormonally regulated. Our results also indicate that the HPA-immune axis is involved in this phenomenon.

A hemorrhagic toxin, designated Elegatoxin, was isolated from the venom of Trimeresurus elegans using HW-55, DEAE-Sephacel, CM-Cellulose and Mono S column chromatographies. The purified toxin was shown to be homogeneous by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, isoelectric electrophoresis, and Ouchterlony immunodiffusion. Elegatoxin has a molecular weight of 26,000 with an isoelectric point of 8.6. The toxin demonstrated both hemorrhagic and proteolytic activities. Hemorrhagic activity was inhibited by ethylenediaminetetraacetic acid (EDTA), ethyleneglycol-bis-(2-amino-ethylether)N,N'-tetraacetic acid (EGTA), o-phenanthroline, and N-bromosuccinimide, but not by amidinophenylmethanesulfonyl fluoride hydrochloride (APMSF). The minimum hemorrhagic dose was found to be 0.8 microgram/mouse. Elegatoxin possesses proteolytic activity as evidenced by hydrolyzing type IV collagen, actin and the A alpha, B beta, and gamma chains of bovine fibrinogen. This purified toxin contains 1 mol of zinc and 2 mols of calcium per mol of protein and a partial amino acid sequence was determined. The pathological and biochemical properties of Elegatoxin were investigated, and these results are reported in this paper.

There is a broad spectrum of renal involvement following snake envenomation. At the clinical level the renal manifestation may be absent or minimal. Mild proteinuria with abnormal urinary sediment may be observed. Significant proteinuria is uncommon. Hematuria and hemoglobinuria are seen in envenomation by vipers or crotalids, while myoglobinuria follows envenomation of sea snakes or elapids. Acute renal failure can occur in these snake bites. All renal structures can be involved. Mesangial proliferative glomerulonephritis is common. Tubular necrosis is the important pathological counterpart of acute renal failure. Three mechanisms including hemodynamic alterations, immunologic reactions, and direct nephrotoxicity are incriminated in the pathogenesis of renal lesions.

Elucidation of naturally occurring toxins is very important from various points of view. Of these toxin studies, search for cytotoxins which show selective toxicity against tumor cells is a challenging subject of study. In search of new bioactive substances from marine organisms, we have been investigating cytotoxic constituents in marine sponges. Through bioassay-guided separation of marine spongean extracts by use of L1210 and KB cell lines, we isolated several potent cytotoxins. This article reviews our recent investigations of three spongean cytotoxins: 1) altohyrtin A and its allied compounds (macrolides, IC50 0.01-0.4 ng/ml (KB cells)) from Hyrtios altum, 2) arenastatin A (a depsipeptide, IC50 5 pg/ml (KB)) from Dysidea arenaria, and 3) callystatin A (a polyketide, IC50 10 pg/ml (KB)) from Callyspongia truncata.

Acetylcholinesterase (AChE) plays a key role in cholinergic transmission. At the neuromuscular junction of vertebrates, for example, it allows a fine temporal control of muscle contraction. The presence of AChE in tissues devoid of cholinergic function is also well known and raises the question of its role. In particular, AChE is abundant in the venoms of Elapid snakes, except mambas. AChE purified from snake venom consists of soluble, hydrophilic monomers. Cloning of cDNA of the AChE from Bungarus fasciatus venom showed that its C-terminal peptide is very different from those of other AChEs. The partial sequence of the Bungarus fasciatus AChE gene shows that this peptide is encoded by a new alternative exon, called S for soluble and snake. It is a short very basic peptide of 15 residues. Analysis of the venom enzyme and in vitro expression experiments showed that the last eight residues are removed in the mature protein. AChEs from snake venoms vary in their sensitivity to peripheral site inhibitors, notably to fasciculins from mamba venoms. While Ophiophagus AChE is as sensitive as Torpedo enzyme (IC50 around 10(-10) M), Naja and Heamacatus AChEs are insensitive to the toxin up to a concentration of 10(-6) M Bungarus AChE has an intermediate IC50 of 10(-8) M. Analysis of its sequence reveals two major differences in the peripheral site region, compared to Torpedo or mammalian AChEs: at position 70 it contains a methionine instead of a tyrosine, and at position 285 it contains a lysine instead of an acidic residue (glutamic or aspartic acid). Modification of these residues by site-directed mutagenesis, and enzymatic analysis of modified recombinant enzymes, confirmed that these two residues are implicated in the properties of the Bungarus AChE peripheral site. The presence of an alternative exon, which generates a soluble form of AChE in venoms, raises interesting evolutionary questions: Does it exist in snakes whose venom does not contain AChE, e.g., mambas? Did this exon pre-exist, for expression in other contexts? Snake venom AChEs offer an exceptional system for analyzing the mechanism of peripheral site inhibition, because of their wide range of activities.

Studies on structures, biological activities, chemical properties, taxonomic distribution, biosynthesis, and evolution of toxins from sea cucumbers (the phylum Echinodermata, the class Holothurioidea) were reviewed with special emphasis on recent results from our laboratory. These toxins are triterpene oligoglycosides having very often one or several sulfate groups in carbohydrate moieties. Their aglycones belong to lanostane derivatives and sometimes contain shortened side chains. Many aglycones are labile in the acid medium. There is a relationship between structures of the glycosides and taxonomic positions of corresponding animals, producers of these toxins. Toxins from sea cucumbers act on delta 5-sterol-containing biological membranes with the formation of glycoside-sterol complexes followed by the disturbance of membrane permeability and inhibition of activities of some membrane enzymes. The presence of the toxins causes the alterations in membrane sterol compositions of toxic sea cucumbers in comparison with non-toxic species. These alterations include the change of delta 5-sterols for those having 7(8)- and 9(11)-double bonds as well as biotransformation of a part of free sterol fractions into sterol sulfates and sterol xylosides.