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Tese IPEN-doc 30061 Potencial genotóxico in vivo de 2-Alcilciclobutanonas provenientes de manteiga de cacau irradiada, em células hepáticas de ratos Wistar2020 - MARTINS, REGIANEA irradiação de alimentos mostra-se uma técnica eficaz para reduzir alguns patógenos dos alimentos e isso ganhou atenção significativa ao seu potencial de segurança do alimento. As 2-Alcilciclobutanonas são produtos radiolíticos únicos gerados pela quebra induzida, por radiação, de trigicerídeos nos alimentos que contêm gordura. 2-Dodecilciclobutanona (2-dDCB) e 2-Tetradecilciclobutanona (2- tDCB) são os compostos mais abundantes detectados em alimentos irradiados, e sabe-se que parte desses compostos ingeridos são excretados pelas fezes e uma pequena parte é depositada no tecido adiposo. Trabalhos realizados anteriormente sugeriram efeitos genotóxicos e citotóxicos em células de cólon. Apesar de estudos mais recentes mostrarem a não genotoxicidade de 2-ACBs, os resultados são conflitantes e, portanto, continuamos os estudos para confirmar a segurança dos compostos para a saúde humana. As células hepáticas foram escolhidas para análise de genotoxicidade por até o momento não terem sido avaliadas in vivo e pela importância do fígado na metabolização de compostos e acúmulo de gordura. O teste de micronúcleo in vivo foi realizado com células específicas extraídas do tecido hepático, por meio de Citometria de fluxo. A análise histológica do tecido hepático foi realizada como análise complementar. Utilizamos manteiga de cacau irradiada com doses de 10, 20 e 30 kGy (consideradas altas doses) para suplementação diária dos animais, a detecção e quantificação de 2-ACBs nas amostras foram obtidas por GC/MS. Os resultados confirmam a segurança do processo de irradiação de alimentos, que mesmo em altas doses de radiação e maior produção de 2-ACBs não indicaram o potencial genotóxico das amostras em células hepáticas.Artigo IPEN-doc 27057 Evaluation of cytotoxicity through MTS test of 2-ACBs (2-dDCB AND tDCB) after exposure to hepatic cells2019 - BARBEZAN, A.B.; CARVALHO, L.R.; VIEIRA, D.P.; MACHADO-SANTELLI, G.M.; MARTINS, R.; VILLAVICENCIO, A.L.C.H.Food irradiation has been approved in more than 60 countries for many applications in a wide variety of foods. 2-Alkylcyclobutanones (2-ACBs) are the only known radiolytic products formed when foods containing fatty acids are irradiated. Despite the importance of food irradiation, the toxicological potential of 2-ACBs in irradiated food is still no fully understood. In this study we investigated the cytotoxic effects of irradiated palmitic and stearic fatty acids byproducts, 2-dDCB and 2-tDCB, in hepatic cell lines (HepG2, BRL3A and HTC). The cytotoxic effects of 2-dDCB and 2-tDCB were evaluated at 100, 300 and 500 μM for 24 and 48 hours and the cell viability was measured using the MTS assay. While no toxicity was observed for 2-tDCB in all cells for all tested conditions, 2-dDCB was found to be toxic to BRL3A cells (at 100 μM after 48 hours) and HTC cells (at 24 hours in all tested concentrations). HepG2 cells on the other hand, were found to be resistant to 2-dDCB-induced toxicity. Overall our data shows that the byproduct 2-tDCB is not toxic for hepatic cells while 2-dDCB can be used has a potential marker for food irradiation-induced toxicity.Artigo IPEN-doc 27906 In vivo genotoxicity of 2-alkylcyclobutanones in liver cells from rats fed with irradiated cocoa butter using flow cytometry2021 - MARTINS, R.; VIEIRA, D.P.; CARVALHO, L.R.; BARBEZAN, A.B.; VILLAVICENCIO, A.L.C.H.Food irradiation proves to be an effective technique of eliminating some pathogens from food and this has gained significant attention to its potential for food safety. Since 1990, studies on the toxicological safety of 2-Alkylcyclobutanones have been conducted extensively. 2-Alkylcyclobutanones are unique radiolytic products generated by the radiation-induced breakage of triglycerides in food, 2-dodecylcyclobutanone (2-dDCB) and 2-tetradecylcyclobutanone (2-tDCB) are the predominant compounds detected . Despite studies showing non-genotoxicity of 2-ACBs (2-Alkylcyclobutanones), the results are conflicting and therefore we continue the studies in order to confirm the compounds are safety for human health. In vivo micronucleus test were performed to verify the 2-ACBs genotoxic effects in hepatic cells using flow cytometry. We used cocoa butter irradiated with 20 kGy. A group with animals was treated with daily intake of irradiated cocoa butter, synthesized 2-dDCB and 2-tDCB for one month. Hepatic cells were selected for genotoxicity analysis due to the liver importance in the compounds metabolization. The improvement of the analytical techniques is important for the research future since the irradiation process is already consolidated. The results confirmed the safety of the food irradiation process, as they did not indicate the genotoxic potential of the samples.Resumo IPEN-doc 26930 Avaliação do potencial citotóxico da 2-tetradecilciclobutana em células hepáticas linhagem HepG22016 - BARBEZAN, A.B.; SALES, B.R.; MARTINS, R.; BUENO, J.B.; SANTELLI, G.M.M.; VILLAVICENCIO, A.L.C.H.Introdução: A irradiação de alimentos é um método eficaz e seguro para a preservação e armazenamento em longo prazo, é aprovado para utilização em mais de 60 países para diversas aplicações em uma ampla variedade de produtos alimentares (Agric. Food Chem. 51; 927, 2003. Food Chem. 201; 52-58, 2016). Este processo é realizado através da utilização de feixes de elétrons acelerados, raios-X ou radiação gama (60Co ou 137Cs). As 2-Alcilciclobutanonas (2-ACBs) são os únicos produtos radiolíticos conhecidos gerados a partir de alimentos que possuem ácidos graxos (Triglicérides) e são submetidos à irradiação (J. Food Protc. 67; 142, 2004. T. Food Scie.Tech. 44; 66-78, 2015). O ácido analisado neste estudo é o esteárico que quando irradiado forma 2-Tetradecilciclobutanona (2-tDCB). Desde a década de 1990 estudos toxicológicos de segurança das 2-ACBs tem sido conduzido extensivamente através de compostos sintéticos. Testes de mutagenicidade das 2-ACBs realizados indicam claramente que nenhuma evidência foi observada, enquanto estudos de viabilidade apresentaram citotoxicidade notada através da morte celular (Food Scie. Tech. 44; 66-78, 2015). Parte das 2-ACBs ingeridas é excretada através das fezes e parte ficam depositadas em tecidos adiposos. Estudos realizados até o momento foram somente em células de cólon. A linhagem escolhida para este trabalho é derivada de células hepáticas uma vez que o acumulo de gordura neste órgão é bastante comum. Objetivo: Avaliar possíveis danos citotóxicos, através do teste de viabilidade celular MTT observando a influência de diversas concentrações da 2-tDCB em diferentes tempos de incubação em células hepáticas da linhagem HepG2. Métodos: O composto 2-tDCB foi solubilizado em etanol a 2%. A linhagem celular escolhida é derivada de hepatocarcinoma humano (HepG2) e foi cultivada em meio de cultura suplementado com 10% de soro fetal bovino. As células foram plaqueadas na densidade de 5x103 cél/poço em uma placa de 96 poços. O efeito citotóxico da 2-tDCB foi avaliado nas concentrações de 100, 300 e 500μM, durante 24 e 48 horas. Os testes foram realizados de acordo com instruções do kit CellTiter96 Aqueus Non-radioactive Cell Proliferation Assay, em triplicatas (biológica e experimental) e os resultados foram analisados pelo programa Prisma GraphPad. Resultados: A linhagem tratada com 2-tDCB em 24 e 48h não apresentou citotoxicidade em nenhuma das concentrações avaliadas. Conclusão: Não houve inviabilidade causada pelo composto 2-tDCB na linhagem de células hepática estudadas, nenhum dano foi observado em nenhuma das variações pesquisadas. Estudos mais aprofundados são necessários para identificar os mecanismos moleculares pela qual o composto em questão atua.Artigo IPEN-doc 26253 In vivo genotoxicity of 2-Alkylcyclobutanones in liver cells from rats fed with irradiated cocoa butter using flow citometry2019 - MARTINS, REGIANE; VIEIRA, DANIEL P.; CARVALHO, LUMA R. de; BARBEZAN, ANGELICA B.; VILLAVICENCIO, ANNA L.C.H.Food irradiation proves to be an effective technique of eliminating some pathogens from food and this has gained significant attention to its potential for food safety. Since 1990, studies on the toxicological safety of 2-Alkylcyclobutanones have been conducted extensively. 2- Alkylcyclobutanones are unique radiolytic products generated by the radiation-induced breakage of triglycerides in food, are exclusively found in irradiated lipid containing foods. 2-dodecylcyclobutanone (2-dDCB) and 2-tetradecylcyclobutanone (2-tDCB) are the predominant compounds detected in irradiated food. Despite studies showing non- genotoxicity of 2-ACBs (2-Alkylcyclobutanones), the results are conflicting and therefore we continue the studies in order to confirm the compounds safety for human health. In vivo micronucleus test were performed to verify the 2-ACBs genotoxic effects in hepatic cells using flow citometry. We used cocoa butter irradiated with 20 kGy at IPEN GAMACELL. A group with animals (IPEN Ethical Animal Experimentation Committee, process number 148/14) was treated with daily intake of irradiated cocoa butter, synthesized 2- Dodecylcyclobutanone and 2-Tetradecylcyclobutanone for two months. Hepatic cells were selected for genotoxicity analysis due to the liver importance in the compounds metabolization. Analyzes were made by micronucleus test with specific cells extracted from hepatic tissue using flow cytometry, which is an alternative to conventional techniques, allowing faster analysis and reduction in the animals number that is a subject much approached in research today. The improvement of the analytical techniques is important for the research future since the irradiation process is already consolidated. The results confirmed the safety of the food irradiation process, as they did not indicate the genotoxic potential of the samples.Artigo IPEN-doc 24270 In vivo evaluation of genotoxic potential of liver cells from rats fed with irradiated diet using flow cytometry2017 - MARTINS, R.; CARVALHO, L.R. de; VIEIRA, D.P.; BARBEZAN, A.B.; VILLAVICENCIO, A.L.C.H.Radiation sources became widely available since 1960’s, and between its main uses are the applications in food irradiation and research of effects of ionizing radiation on food products. Despite some public concern, the process is safe, free from chemical residues and presents advantages for preservation and storage. Nevertheless, safety dose parameters must be adopted in irradiation procedures to inhibit formation of undesirable and/or toxic products, for example, 2-ACB’s (2-alkylcyclobutanones) that are cyclic compounds containing four carbon rings that can be formed in food when its fat content is irradiated through breakdown of fatty acids. 2-ACBs are considered a unique class of compounds due to divergences between results of its mutagenicity potential collected from different studies. In this study, a cell population collected from rat livers were chosen for in vivo genotoxicity analysis because the importance of the liver in the metabolization of compounds. Analysis was performed using the micronuclei test using flow cytometry, allowing faster analysis, use of few materials and reduction in the number of animals, what is a subject much addressed currently in research. Irradiated rat diet did not show any genotoxic effect on liver cell populations. The improvement of the techniques is important for the future of the research since the irradiation process is already consolidated.Resumo IPEN-doc 23834 Potential evaluation of cytotoxic 2-tetradecylcyclobutanone lineage in cellular BRL 3A - studies in vitro2016 - BARBEZAN, A.B.; SALES, B.R.; MARTINS, R.; BUENO, J.B.; SANTELLI, G.M.M.; VILLAVICENCIO, A.L.C.H.Artigo IPEN-doc 23831 Ames test to detect mutagenicity of 2-alkylcyclobutanones: a review2017 - BARBEZAN, ANGELICA B.; MARTINS, REGIANE; BUENO, JENNIFER B.; VILLAVICENCIO, ANNA L.C.H.Food irradiation is an effective and safe method for preservation and long-term storage, and it is approved for use in over 60 countries for various applications in a wide variety of food products. This process is performed by use of accelerated electron beams, X-rays, or gamma radiation (60Co or 137Cs). 2-Alkylcyclobutanones (2-ACBs) are the only known radiolytic products generated from foods that have fatty acids (triglycerides) and are subjected to irradiation. Since the 1990s toxicological safety studies of 2-ACBs have been conducted extensively through synthetic compounds, then and tests to determine if the compounds have any mutagenic activity are strictly necessary. The Ames test was chosen by many researchers to assess the mutagenicity of 2-ACBs. The test uses distinct bacterial cell lines Salmonella typhimurium to detect point mutations at sites guanine–cytosine (G–C) and Escherichia coli to detect point mutations at sites adenine–thymine (A–T). This bibliographic research aims to bring together all the results obtained and a comparison and cell lines used, type of plates, and solvents. This research showed that no mutagenic activity was observed in any of the cell lines and concentrations evaluated by the works of authors, so the 2-ACBs compounds showed no mutagenic substance in concentrations detectable by the Ames test.Resumo IPEN-doc 23759 Potential evaluation of mutagenic compounds 2-dodecylcyclobutanone and 2-tetradecylcyclobutanone through the Ames test2016 - BARBEZAN, ANGELICA B.; BISPO, ANA C.A.; MENDES, BRUNO M.; MARTINS, REGIANE; VILLAVICENCIO, ANNA L.C.H.Food irradiation is an effective and safe method for preservation and long-term storage for various applications in a wide variety of food products. This process is performed by the use of accelerated electron beams, X-rays or gamma radiation. The 2-Alkylcyclobutanones (2-ACBs) are the only known radiolytic products generated in foods that have fatty acids and were subjected to irradiation. In this study, the stearic and palmitic acid was analyzed when irradiated medium 2-Tetradecylcyclobutanone (2-tDCB) and 2-Dodecylcyclobutanone (2-dDCB). Since the 1990s toxicological safety studies of 2-ACBs have been conducted extensively through synthetic compounds. Additionally, tests to determine if the compounds have any mutagenic activity are strictly necessary. The Ames test was the test choosen for assessing the genotoxicity of both 2- dDCB and 2tDCB compounds. It was used five different bacterial strains TA-1535, TA-1537, TA 98 and TA 100 Salmonella typhimurium, to detect mutations in specific sites Guanine-Cytosine (GC) and WP2 uvrA, Escherichia coli was used to detect point mutations at sites of Adenine- Thymine (AT). This research, unlike those undertaken by other authors up to now, presents some particularities in their factors for a more complete assessment, such as the use of all necessary cell lines to identify possible mutations in specific sites, changes in solvent commonly used, appropriate plates and the investigation of possible mutagenic effects on two important compounds 2-dDCB and 2-tDCB simultaneously. The study revealed no mutagenic activity in any of the cell lines and concentrations evaluated. In conclusion, the compounds 2-dDCB and 2tDCB showed no mutagenic effect in concentrations detectable by the AMES test.Resumo IPEN-doc 23504 Avaliação do potencial citotóxico da 2-tetradecilciclobutana em células hepáticas linhagem HepG2– estudos in vitro2016 - BARBEZAN, A.B.; SALES, B.R.; MARTINS, R.; BUENO, J.B.; SANTELLI, G.M.M.; VILLAVICENCIO, A.L.C.H.