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Toxicity of echinoid gonadal extracts
Abstracts/Toxlcon 38 (2000) 487-595
593
samples, rising to only 56% after 48 h. Conclusions: BeV clearance occurs via urinary and gastrointestinal tracts.
Comparative toxiciO, of two puffelfishes .ft'om the brazilian coast. J. Stolarz de Oliveira, J.C. Freitas (Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, S~o Paulo, Brazil). Pufferfishes are known to possess tetrodotoxin, a neurotoxin which blocks the voltage sensitive sodium channels in excitable membranes. In other countries such as Japan, Singapore, China, and Korea, several pufferfish poisonings have been reported; in Brazil, however, human consumption of these fishes is sporadic because all pufferfishes are thought to be lethal. Here we present a comparative study of two species of pufferfish from the Brazilian coast. Specimens of Sphoeroides spengleri (baiacu pintado) and Lagocephalus laevigatus (baiacu arara or pachaco) were caught in S~o Sebasti~o Channel (North coast of S~o Paulo State, Brazil) between 1996 and 1997. Skins + viscera, and muscles of 39 specimens of S. spengleri were excised and extracted in 80% ethanol containing 1% acetic acid. After removal of ethanol by evaporation, the extract was partitioned in methylene chloride and dried again by evaporation. The residue was dissolved in water and injected intraperitonially into mice using the methods of Kawabata, 1978 (Japan. Food Hygienic Association, Tokyo, pp.232-240). We found that the skin plus viscera extract of S. spengleri contained 422.0_+ 25.5 MU/g up to 488.9 _+ 54.2 MU/g and muscle extract contained (up to 385.1 _+ 14.6 MU/g). These levels are always toxic with respect to human consumption. Specimens of S. spengleri from Silo Sebasti~o Island (in the same Channel), had a higher toxicity reaching 946.4 _+ 60.9 MU/g. The extracts of skin+viscera and muscle of 83 specimens of L. laevigatus never exceeded 1.6 MU/g. This is under 10 MU/g, which is consadered to be non toxic. Some fishermen are known to eat the ovary and liver of this species. As it is known that toxicity may vary geographically, we can not be certain that all L. laevigatus from the northern coast of Brazil are edible. Acknowledgements: Supported by CNPq and FAPESP, Brazil. Toxici O, of echinoid gonadal extracts. A.M. Gomes, J.C. Freitas (Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, SP, Brazil). Objectives: Lytechinus variegatus and Echinometra lucunter are the most abundant sea urchins species, in the Brazilian coast. People from north coast of S~o Paulo State occasionally consume E. lucunter gonads, but they do not eat L. variegatus because it is known to be poisonous. In this work, we studied acute toxicity in mice through intraperitonial and oral administrations of gonadal extracts of both species. Methods and results: Gonads were separated by species and sex, homogeneized in methanol (1:10 w/v), filtered, concentrated in a vacuum evaporator and partitioned with methylene chloride in order to obtain the polar and apolar fractions. The EDso method was performed according to IPSEN and FEIGL, 1970 (Brancofts Introduction to Bioestatistcs, NY pp.163 170). We
594
Abstracts/Toxicon 38 (2000) 487 595
used 6 males mice (lineage: Balb-C) from 15 to 20 g for each fraction of extracts to be tested. The apolar fractions were diluted in propileneglicol + saline solution for mammals (1:9) and the polar fractions just in the saline solution in the following concentrations: 108.0, 129.6, 155.5 and 186.6 mg/ml. In the control animals, we used the same vehicle used in these dilutions. The symptoms were observed during 24 h and in the case of lethality the necropsy was realized. Only the polar fractions of L. variegatus intraperitonially administrated, produced motor disturbs, respiratory difficulty and the death occuring in _+10 rain. The DLs0 and the 95% confidence intervals obtained were: 7.88 (7.18-8.45) - male extract and 8.06 (7.36-8.64)g/kg - female extract. Both fractions of E. lucunter administrated orally and intraperitonially didn't cause any symptoms or death. Conclusions: We found in this work that the polar fractions of L. variegatus are more active than the other fractions of E. lucunter, confirming the ethnopharmacologycal data. Acknowledgements: Supported by FAPESP.
Hemolytic activity in the extract of the marine sponge Geodia corticostylifera. M. Rangel a, R.G.S. Berlinck a, J.C. Freitas a (~Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, 05508900, S~o Paulo, Brazil; bChemistry Institute, University of S~o Paulo, S~o Carlos, Brazil). Objectives" Marine sponges are a rich source of bioactive compounds, some of them are precursors for new pharmacological tools and medicines. The aim of this work is to study the hemolytic activity of the extract of the marine sponge Geodia corticostylifera (Porifera, Demospongiae). Materials and methods: Specimens of G. corticostylifera were collected in S~o Paulo State coast, homogeneized in methanol (I:3)(W:V) and filtered. The filtered material was evaporated and partitioned with water/methylene chloride (1:1) (V:V). The apolar fraction volume was reduced in a vacuum evaporator and partitionated in methanol-water(9:l)/n-hexane (1:2) (V:V). The methanol-water fraction was freezedried and tested in the following experiments. After one hour incubation at room temperature (25 + 2°C) with a 0.5% mouse erythrocytes suspension (ES), it was centrifuged and the hemolytic activity was measured by the absorbance at 540 nm in the supernatant. The ECs0 of hemolytic activity was 168.0 ~tg/ml ES with 95% confidence interval from 153.0 to 184.4 (n=5). The membrane phospholipids sphingomyelin, phosphatidylcholine and cholesterol were tested as acceptors for the hemolysin of the extract in three different concentrations: 75, 225 and 675 ~g/ ml ES. The EC50 and 95% confidence intervals values were, respectively (in ~tg/ml ES) to sphingomyelin: 244.7 (225.1 266.0), 279.3 (226.3-344.7) and 378.6 (206.4 694.3); phosphatidylcoline: 269.6 (223.8 324.7), 330.2 (230.2-473.7) and 516.9 (122.1 2189.0); and to cholesterol: 209.1 (197.5 221.4); 231.6 (222.9-240.6) and 201.0 (175.0-230.8) (n=4). The logarithms of EC50 of the tests performed with the lipids were statistically different from the control (with the extract only), according with analysis of variance and Student-Newman-Keuls post test (p < 0.05). Conclusions: The methanol-water fraction of the extract of G. corticostylifera showed hemolytic activity, and the lipids sphingomyelin,
593
samples, rising to only 56% after 48 h. Conclusions: BeV clearance occurs via urinary and gastrointestinal tracts.
Comparative toxiciO, of two puffelfishes .ft'om the brazilian coast. J. Stolarz de Oliveira, J.C. Freitas (Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, S~o Paulo, Brazil). Pufferfishes are known to possess tetrodotoxin, a neurotoxin which blocks the voltage sensitive sodium channels in excitable membranes. In other countries such as Japan, Singapore, China, and Korea, several pufferfish poisonings have been reported; in Brazil, however, human consumption of these fishes is sporadic because all pufferfishes are thought to be lethal. Here we present a comparative study of two species of pufferfish from the Brazilian coast. Specimens of Sphoeroides spengleri (baiacu pintado) and Lagocephalus laevigatus (baiacu arara or pachaco) were caught in S~o Sebasti~o Channel (North coast of S~o Paulo State, Brazil) between 1996 and 1997. Skins + viscera, and muscles of 39 specimens of S. spengleri were excised and extracted in 80% ethanol containing 1% acetic acid. After removal of ethanol by evaporation, the extract was partitioned in methylene chloride and dried again by evaporation. The residue was dissolved in water and injected intraperitonially into mice using the methods of Kawabata, 1978 (Japan. Food Hygienic Association, Tokyo, pp.232-240). We found that the skin plus viscera extract of S. spengleri contained 422.0_+ 25.5 MU/g up to 488.9 _+ 54.2 MU/g and muscle extract contained (up to 385.1 _+ 14.6 MU/g). These levels are always toxic with respect to human consumption. Specimens of S. spengleri from Silo Sebasti~o Island (in the same Channel), had a higher toxicity reaching 946.4 _+ 60.9 MU/g. The extracts of skin+viscera and muscle of 83 specimens of L. laevigatus never exceeded 1.6 MU/g. This is under 10 MU/g, which is consadered to be non toxic. Some fishermen are known to eat the ovary and liver of this species. As it is known that toxicity may vary geographically, we can not be certain that all L. laevigatus from the northern coast of Brazil are edible. Acknowledgements: Supported by CNPq and FAPESP, Brazil. Toxici O, of echinoid gonadal extracts. A.M. Gomes, J.C. Freitas (Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, SP, Brazil). Objectives: Lytechinus variegatus and Echinometra lucunter are the most abundant sea urchins species, in the Brazilian coast. People from north coast of S~o Paulo State occasionally consume E. lucunter gonads, but they do not eat L. variegatus because it is known to be poisonous. In this work, we studied acute toxicity in mice through intraperitonial and oral administrations of gonadal extracts of both species. Methods and results: Gonads were separated by species and sex, homogeneized in methanol (1:10 w/v), filtered, concentrated in a vacuum evaporator and partitioned with methylene chloride in order to obtain the polar and apolar fractions. The EDso method was performed according to IPSEN and FEIGL, 1970 (Brancofts Introduction to Bioestatistcs, NY pp.163 170). We
594
Abstracts/Toxicon 38 (2000) 487 595
used 6 males mice (lineage: Balb-C) from 15 to 20 g for each fraction of extracts to be tested. The apolar fractions were diluted in propileneglicol + saline solution for mammals (1:9) and the polar fractions just in the saline solution in the following concentrations: 108.0, 129.6, 155.5 and 186.6 mg/ml. In the control animals, we used the same vehicle used in these dilutions. The symptoms were observed during 24 h and in the case of lethality the necropsy was realized. Only the polar fractions of L. variegatus intraperitonially administrated, produced motor disturbs, respiratory difficulty and the death occuring in _+10 rain. The DLs0 and the 95% confidence intervals obtained were: 7.88 (7.18-8.45) - male extract and 8.06 (7.36-8.64)g/kg - female extract. Both fractions of E. lucunter administrated orally and intraperitonially didn't cause any symptoms or death. Conclusions: We found in this work that the polar fractions of L. variegatus are more active than the other fractions of E. lucunter, confirming the ethnopharmacologycal data. Acknowledgements: Supported by FAPESP.
Hemolytic activity in the extract of the marine sponge Geodia corticostylifera. M. Rangel a, R.G.S. Berlinck a, J.C. Freitas a (~Department of Physiology, Biosciences Institute and Marine Biology Center, University of S~o Paulo, 05508900, S~o Paulo, Brazil; bChemistry Institute, University of S~o Paulo, S~o Carlos, Brazil). Objectives" Marine sponges are a rich source of bioactive compounds, some of them are precursors for new pharmacological tools and medicines. The aim of this work is to study the hemolytic activity of the extract of the marine sponge Geodia corticostylifera (Porifera, Demospongiae). Materials and methods: Specimens of G. corticostylifera were collected in S~o Paulo State coast, homogeneized in methanol (I:3)(W:V) and filtered. The filtered material was evaporated and partitioned with water/methylene chloride (1:1) (V:V). The apolar fraction volume was reduced in a vacuum evaporator and partitionated in methanol-water(9:l)/n-hexane (1:2) (V:V). The methanol-water fraction was freezedried and tested in the following experiments. After one hour incubation at room temperature (25 + 2°C) with a 0.5% mouse erythrocytes suspension (ES), it was centrifuged and the hemolytic activity was measured by the absorbance at 540 nm in the supernatant. The ECs0 of hemolytic activity was 168.0 ~tg/ml ES with 95% confidence interval from 153.0 to 184.4 (n=5). The membrane phospholipids sphingomyelin, phosphatidylcholine and cholesterol were tested as acceptors for the hemolysin of the extract in three different concentrations: 75, 225 and 675 ~g/ ml ES. The EC50 and 95% confidence intervals values were, respectively (in ~tg/ml ES) to sphingomyelin: 244.7 (225.1 266.0), 279.3 (226.3-344.7) and 378.6 (206.4 694.3); phosphatidylcoline: 269.6 (223.8 324.7), 330.2 (230.2-473.7) and 516.9 (122.1 2189.0); and to cholesterol: 209.1 (197.5 221.4); 231.6 (222.9-240.6) and 201.0 (175.0-230.8) (n=4). The logarithms of EC50 of the tests performed with the lipids were statistically different from the control (with the extract only), according with analysis of variance and Student-Newman-Keuls post test (p < 0.05). Conclusions: The methanol-water fraction of the extract of G. corticostylifera showed hemolytic activity, and the lipids sphingomyelin,