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Genotoxicity of liquid products from pyrolysis of sewage sludge
Abstracts / Toxicology Letters 196S (2010) S37–S351
(HUVEC). For all cell types, 40 and 10 M concentrations were tested during 24 hours and developed by SRB method. F98226BM had stimulated IC50 > 40 M in all human cells tested. doi:10.1016/j.toxlet.2010.03.570
P202-035 Acute systemic toxicological and mutagenic evaluations of a proteolytic fraction from latex of Carica candamarcensis M. Villalba, A. Silva, C. Tagliati, G. Cassali, C. Salas, M. Lopes Universidade Federal de Minas Gerais, Brazil Our research group has been characterizing a fraction (P1G10) derived from latex of Carica candamarcensis L. (voucher #15063, Univ. La Serena, Chile). The fraction is rich in cysteine-proteases and displays wound healing activity in vivo on skin and gastric wounds. In preparation for clinical trials, this study aims to evaluate the pre-clinical toxicological and mutagenic effects induced by acute P1G10 administration. Methods and results: The acute toxicity assay (Fixed Dose Procedure) was carried out following the guidelines provided by the OECD 420, 2001. The lethal dose by p.o. route was determined at 2000 mg/kg, while i.p. or i.v. administration yielded a lethal dose of 50 mg/kg. Histopathological examination following necropsy showed diffuse bleeding on heart, kidneys, bowels and stomach, as probably cause of death. By giving 300 mg/kg (v.o.) or 5 mg/kg (i.p. or i.v.), P1G10 did not cause death or visible changes in body weight, water consumption or food intake, when compared to the control. The mutagenic evaluation was carried out in vitro by the Ames test, exposing different Salmonella typhimurium strains (TA97, TA98, TA100, TA102) to 0.1 or 1% P1G10, the corresponding positive control or sterile saline, during 24 h. The number of revertant colonies was significantly different when compared to the positive control (p < 0.001 ANOVA, Bonferroni post-test), but no significantly different to the negative control. Similarly, the in vivo micronucleous test, applied to Swiss mice treated with 5 or 10 mg/kg P1G10 did not alter the number of polychromatic erythrocytes with micronucleus levels at the bone marrow similar to the negative control. Conclusion: Considering these results, we can infer that P1G10 is well tolerated in the studied doses as determined by different routes of administration and has no mutagenic activity, clearing the way for future prospective studies. Financial support: CAPES, CNPq and FAPEMIG. doi:10.1016/j.toxlet.2010.03.571
P202-036 Genotoxic effect of 3-tesla magnetic resonance image (MRI) in human lymphocytes J.W. Lee 1 , Y.J. Kim 1 , M.S. Kim 2 , H.D. Woo 1 , Y.H. Lee 1 , H.W. Chung 1 1
The Graduate School of Public Health, Institute of Health and Environment, Seoul National University, Republic of Korea, 2 Department of Radiology, National Cancer Center, Republic of Korea Clinical use of magnetic resonance imaging (MRI) scanner with higher magnetic field strengths is increased in recent days. 3-Tesla (T) MRI has twice the field strength compared with conventional 1.5-T MRI units. In this study, the genotoxic potential of high magnetic field generated during a 3-T clinical MRI scan in human lymphocyte was investigated by analyzing chromosome aberration
S167
(CA), micronucleus (MN) and single cell gel electrophoresis (Comet assay) in vitro. Human lymphocytes were exposed to complex MRI field conditions (clinical routine brain protocol: 3 channel head coil in GE HDx 3.0-T; T2-FSE, T2-FLAIR, T1-SE, DW, DTI, T1-FLAIR sequence) for 22, 45, 67 and 89 min, respectively. To rule out the thermal effects, all sample temperatures were maintained at 25 ± 1 ◦ C by constant temperature and humidity equipment. DNA single-strand breaks were increased significantly after exposure to 3-T MRI. In the longest exposure time, DNA tail moment was on the average threefold more than that of control. Frequency of MN and CA were also increased in an exposure time dependent manner. The MN frequency in the lymphocytes exposed for 0, 22, 45, 67 and 89 min were 9, 11, 13.5, 16 and 18.5 per 1000 cells, respectively. Similarly, the frequency of CA in the lymphocytes exposed for 0, 67 and 89 min were 1.5, 3.5 and 5.5 per 200 cells. Obtained results suggest that exposure to 3-T MRI induce genotoxic effects in vitro in human lymphocytes. doi:10.1016/j.toxlet.2010.03.572
P202-037 Genotoxicity of liquid products from pyrolysis of sewage sludge ˜ 2 , M.J. Hazen 3 A. Pillco 1 , E. De La Pena 1
Biochemical Research Institute, University Major of San Andres (UMSA), Saavedra, 2224, La Paz, Bolivia, 2 Laboratory of Environmental Mutagenesis, Environmental Sciences Center, Spanish National Research Council (CSIC), Serrano 115, 28006 Madrid, Spain, 3 Department of Biology, Faculty of Sciences, Autonomous University of Madrid, Darwin 2, 28049 Madrid, Spain Sewage sludge is the waste produced in wastewater treatment plants. Various thermal treatments such as gasification and pyrolysis are currently being studied to valorize this waste. The aim of this study was to analyze the potential genotoxicity produced by a liquid product from pyrolysis (LPP) of sewage sludge. Two short-term bioessays were employed. First LPP was evaluated using the Ames Salmonella/microsome mutagenicity test. The protocol was followed, using tester strains TA98, TA100, TA102 and TA104 of Salmonella typhimurium. The results showed presence of mutagenic activity produced by LPP. TA98 and TA100 strains were the most sensitive. The second bioessay employed was the in vivo Wing Somatic Mutation and Recombination Test (SMART) in Drosophila melanogaster. The protocol followed used standard cross (ST) and high bioactivation cross (HB). 72-h-old larvae trans-heterozygous for two genetic markers “multiple wing hairs” (mwh) and “flare” (flr3), were treated at different concentrations of the LPP for a period of 48 h. The wings of the emerging adult flies were scored for the presence of spots of mutant cells. For ST, the result showed that none of three categories of mutant spots recorded (small, large and twin) increased significantly by these treatments, independently of the dose supplied. In contrast, using HB, LPP showed positive results only for small single spots and weak positive results for total spots. In conclusion, both alternative methods to animal testing (3Rs) employed demonstrate the existence of a genotoxic risk for LPP, and indicate the need for further research to delineate the exact mechanisms involved. Project OTT2007X1317, *MAEC-AECID. doi:10.1016/j.toxlet.2010.03.573
(HUVEC). For all cell types, 40 and 10 M concentrations were tested during 24 hours and developed by SRB method. F98226BM had stimulated IC50 > 40 M in all human cells tested. doi:10.1016/j.toxlet.2010.03.570
P202-035 Acute systemic toxicological and mutagenic evaluations of a proteolytic fraction from latex of Carica candamarcensis M. Villalba, A. Silva, C. Tagliati, G. Cassali, C. Salas, M. Lopes Universidade Federal de Minas Gerais, Brazil Our research group has been characterizing a fraction (P1G10) derived from latex of Carica candamarcensis L. (voucher #15063, Univ. La Serena, Chile). The fraction is rich in cysteine-proteases and displays wound healing activity in vivo on skin and gastric wounds. In preparation for clinical trials, this study aims to evaluate the pre-clinical toxicological and mutagenic effects induced by acute P1G10 administration. Methods and results: The acute toxicity assay (Fixed Dose Procedure) was carried out following the guidelines provided by the OECD 420, 2001. The lethal dose by p.o. route was determined at 2000 mg/kg, while i.p. or i.v. administration yielded a lethal dose of 50 mg/kg. Histopathological examination following necropsy showed diffuse bleeding on heart, kidneys, bowels and stomach, as probably cause of death. By giving 300 mg/kg (v.o.) or 5 mg/kg (i.p. or i.v.), P1G10 did not cause death or visible changes in body weight, water consumption or food intake, when compared to the control. The mutagenic evaluation was carried out in vitro by the Ames test, exposing different Salmonella typhimurium strains (TA97, TA98, TA100, TA102) to 0.1 or 1% P1G10, the corresponding positive control or sterile saline, during 24 h. The number of revertant colonies was significantly different when compared to the positive control (p < 0.001 ANOVA, Bonferroni post-test), but no significantly different to the negative control. Similarly, the in vivo micronucleous test, applied to Swiss mice treated with 5 or 10 mg/kg P1G10 did not alter the number of polychromatic erythrocytes with micronucleus levels at the bone marrow similar to the negative control. Conclusion: Considering these results, we can infer that P1G10 is well tolerated in the studied doses as determined by different routes of administration and has no mutagenic activity, clearing the way for future prospective studies. Financial support: CAPES, CNPq and FAPEMIG. doi:10.1016/j.toxlet.2010.03.571
P202-036 Genotoxic effect of 3-tesla magnetic resonance image (MRI) in human lymphocytes J.W. Lee 1 , Y.J. Kim 1 , M.S. Kim 2 , H.D. Woo 1 , Y.H. Lee 1 , H.W. Chung 1 1
The Graduate School of Public Health, Institute of Health and Environment, Seoul National University, Republic of Korea, 2 Department of Radiology, National Cancer Center, Republic of Korea Clinical use of magnetic resonance imaging (MRI) scanner with higher magnetic field strengths is increased in recent days. 3-Tesla (T) MRI has twice the field strength compared with conventional 1.5-T MRI units. In this study, the genotoxic potential of high magnetic field generated during a 3-T clinical MRI scan in human lymphocyte was investigated by analyzing chromosome aberration
S167
(CA), micronucleus (MN) and single cell gel electrophoresis (Comet assay) in vitro. Human lymphocytes were exposed to complex MRI field conditions (clinical routine brain protocol: 3 channel head coil in GE HDx 3.0-T; T2-FSE, T2-FLAIR, T1-SE, DW, DTI, T1-FLAIR sequence) for 22, 45, 67 and 89 min, respectively. To rule out the thermal effects, all sample temperatures were maintained at 25 ± 1 ◦ C by constant temperature and humidity equipment. DNA single-strand breaks were increased significantly after exposure to 3-T MRI. In the longest exposure time, DNA tail moment was on the average threefold more than that of control. Frequency of MN and CA were also increased in an exposure time dependent manner. The MN frequency in the lymphocytes exposed for 0, 22, 45, 67 and 89 min were 9, 11, 13.5, 16 and 18.5 per 1000 cells, respectively. Similarly, the frequency of CA in the lymphocytes exposed for 0, 67 and 89 min were 1.5, 3.5 and 5.5 per 200 cells. Obtained results suggest that exposure to 3-T MRI induce genotoxic effects in vitro in human lymphocytes. doi:10.1016/j.toxlet.2010.03.572
P202-037 Genotoxicity of liquid products from pyrolysis of sewage sludge ˜ 2 , M.J. Hazen 3 A. Pillco 1 , E. De La Pena 1
Biochemical Research Institute, University Major of San Andres (UMSA), Saavedra, 2224, La Paz, Bolivia, 2 Laboratory of Environmental Mutagenesis, Environmental Sciences Center, Spanish National Research Council (CSIC), Serrano 115, 28006 Madrid, Spain, 3 Department of Biology, Faculty of Sciences, Autonomous University of Madrid, Darwin 2, 28049 Madrid, Spain Sewage sludge is the waste produced in wastewater treatment plants. Various thermal treatments such as gasification and pyrolysis are currently being studied to valorize this waste. The aim of this study was to analyze the potential genotoxicity produced by a liquid product from pyrolysis (LPP) of sewage sludge. Two short-term bioessays were employed. First LPP was evaluated using the Ames Salmonella/microsome mutagenicity test. The protocol was followed, using tester strains TA98, TA100, TA102 and TA104 of Salmonella typhimurium. The results showed presence of mutagenic activity produced by LPP. TA98 and TA100 strains were the most sensitive. The second bioessay employed was the in vivo Wing Somatic Mutation and Recombination Test (SMART) in Drosophila melanogaster. The protocol followed used standard cross (ST) and high bioactivation cross (HB). 72-h-old larvae trans-heterozygous for two genetic markers “multiple wing hairs” (mwh) and “flare” (flr3), were treated at different concentrations of the LPP for a period of 48 h. The wings of the emerging adult flies were scored for the presence of spots of mutant cells. For ST, the result showed that none of three categories of mutant spots recorded (small, large and twin) increased significantly by these treatments, independently of the dose supplied. In contrast, using HB, LPP showed positive results only for small single spots and weak positive results for total spots. In conclusion, both alternative methods to animal testing (3Rs) employed demonstrate the existence of a genotoxic risk for LPP, and indicate the need for further research to delineate the exact mechanisms involved. Project OTT2007X1317, *MAEC-AECID. doi:10.1016/j.toxlet.2010.03.573