Protección del ácido clorogénico frente a lesiones inducidas por venzo(a)pireno en Saccharomyces cerevisiae / Protection of chlorogenic acid against benzo(a)pyrene-induced lesions in Saccharomyces cerevisiae / Proteção do ácido clorogênico contra lesões induzidas por benzo(a)pireno em Saccharomyces cerevisiae

El objetivo del presente estudio fue analizar el efecto del ácido clorogénico, uno de los compuestos polifenólicos con mayor concentración en la infusión de Ilex paraguariensis, sobre el daño celular y molecular inducido por el benzo(a)pireno. La infusión de Ilex paraguariensis ("mate") es bebida por la mayoría de los habitantes de Argentina, Paraguay, sur de Brasil y Uruguay. La levadura Saccharomyces cerevisiae (cepas SC7K lys2-3; SX46A y SX46Arad14() se utilizó como modelo eucariota. Las células en crecimiento exponencial se expusieron a concentraciones crecientes de benzo(a)pireno y a tratamientos combinados con una concentración de 250 ng/mL de benzo(a)pireno y ácido clorogénico a una concentración igual a la encontrada en la infusión de yerba mate. Luego de los tratamientos se determinaron fracciones de sobrevida, frecuencia mutagénica y roturas de doble cadena de ADN así como la modulación en la expresión de la proteína Rad14 a través de un análisis de Western Blot. Se observó un aumento significativo en las fracciones de sobrevida así como una disminución en la frecuencia mutagénica después de la exposición combinada con benzo(a)pireno y ácido clorogénico en comparación con los tratamientos con benzo(a)pireno como único agente. En la cepa mutante deficiente en la proteína Rad14 se observó un aumento significativo en la sensibilidad a benzo(a)pireno en comparación con la cepa SC7K lys2-3. En los tratamientos combinados de benzo(a)pireno y ácido clorogénico se observó una importante disminución de la letalidad. Con respecto a la determinación de roturas de doble cadena de ADN no se observó fraccionamiento cromosómico a la concentración de benzo(a)pireno utilizada en los experimentos. Los análisis de Western Blot mostraron un aumento en la expresión de la proteína Rad14 en las muestras tratadas con benzo(a)pireno como único agente en comparación con la muestra control. Adicionalmente se observó una disminución en la expresión de la proteína cuando en los tratamientos se utilizaron benzo(a)pireno y ácido clorogénico combinados. Los resultados indican que el ácido clorogénico disminuye significativamente la actividad mutagénica producida por el benzo(a)pireno, la cual no se encuentra relacionada con un incremento en la remoción de las lesiones inducidas por el sistema de reparación por escisión de nucleótidos.

The aim of this study was to analyze the effect of chlorogenic acid, a polyphenolic compound found at high concentrations in Ilex paraguariensis infusions, on cellular and molecular damage induced by benzo(a)pyrene. Ilex paraguariensis infusions ("mate") are consumed by most of the population in Argentina, Paraguay, southern Brazil and Uruguay. Saccharomyces cerevisiae yeast (SC7K lys2-3; SX46A and SX46Arad14( strains) were used as eukaryotic model organisms. Cells in an exponential growth phase were exposed to increasing concentrations of benzo(a)pyrene, as well as combined treatments of benzo(a)pyrene at a concentration of 250 ng/mL and chlorogenic acid at a concentration matching that which is commonly found in mate. Determinations of surviving fraction, mutagenic frequency and double strand DNA breaks induction were performed, along with the assessment of the modulation of the expression of protein Rad14 by Western Blot. A significant increase of surviving fractions and a decrease in mutagenic frequency were observed after exposure to benzo(a)pyrene plus chlorogenic acid, contrary to benzo(a)pyrene alone. A substantial increase in sensitivity to benzo(a)pyrene was observed for the Rad14 protein-deficient mutating strain when compared to the SC7K lys2-3 strain. An important decrease in lethality was observed when combined benzo(a)pyrene and chlorogenic acid treatments were applied. As for the determination of DSBs, no chromosomic fractionation was observed at the benzo(a)pyrene concentration tested in the experiments. Western Blot analysis showed an increase in the expression of protein Rad14 for samples treated with benzo(a)pyrene as a single agent when compared against the control sample. Additionally, the expression of this protein was observed to diminish when combined treatments with benzo(a)pyrene and chlorogenic acid were used. Results obtained indicate that chlorogenic acid significantly decreases the mutagenic activity of benzo(a)pyrene, which is not related to an increase in the removal of lesions induced by nucleotide excision repair system.

O objetivo deste estudo foi analisar o efeito do ácido clorogênico, um dos compostos polifenólicos com maior concentração na infusão de Ilex paraguariensis, sobre o dano celular e molecular induzido pelo benzopireno. A infusão de Ilex paraguariensis ("mate") é consumida pela maioria dos habitantes da Argentina, Paraguai, sul do Brasil e Uruguai. A levedura Saccharomyces cerevisiae (cepas SC7K lys2-3; SX46A e SX46Arad14() foi utilizada como modelo eucariótico. Células em crescimento exponencial foram expostas a concentrações crescentes de benzopireno e tratamentos combinados foram realizados com uma concentração de 250 ng/mL de benzo(a)pireno e ácido clorogênico, igual à encontrada na infusão de erva-mate. Após os tratamentos, foram determinadas as frações de sobrevivência, frequência mutagênica e quebras de fita dupla do DNA, bem como a modulação na expressão da proteína Rad14 por meio de análise de Western Blot. Um aumento significativo nas frações de sobrevivência, bem como uma diminuição na frequência mutagênica foram observados após a exposição combinada de benzo(a)pireno e ácido clorogênico em comparação com tratamentos de agente único de benzo(a)pireno. Um aumento significativo na sensibilidade ao benzo(a)pireno foi observado na cepa mutante deficiente em proteína Rad14 em comparação com a cepa SC7K lys2-3. Nos tratamentos combinados de benzo(a)pireno e ácido clorogênico, observou-se uma diminuição significativa na letalidade. Com relação à determinação das quebras de fita dupla de DNA, não foi observado fracionamento cromossômico na concentração de benzo(a)pireno utilizada nos experimentos. A partir da análise de Western Blot, observou-se um aumento na expressão da proteína Rad14 nas amostras tratadas com benzo(a)pireno como agente único em relação à amostra controle. Além disso, uma diminuição na expressão da proteína foi observada quando combinados de benzo(a)pireno e ácido clorogênico foram usados ​​nos tratamentos. Os resultados obtidos neste trabalho indicam que o ácido clorogênico diminui significativamente a atividade mutagênica produzida pelo benzo(a)pireno, a qual não está relacionada a um aumento na remoção de lesões induzidas pelo sistema de reparo por excisão de nucleotídeos.

Mutagenicity and antimutagenicity of Salacia crassifolia (mart. Ex. Schult.) G. Don. evaluated by Ames test / Mutagenicitade e antimutagenicidade de Salacia crassifolia (mart. Ex. Schult.) G. Don. avaliadas pelo teste de Ames

Abstract Salacia crassifolia (Mart. Ex. Schult.) G. Don. is a bush which belongs to Celastraceae family and occurs specially in Brazilian Cerrado. Its leaves, stem, seeds and fruits are popularly used for several medicinal purposes, such as antitumoral, antirheumatic, anti-inflammatory and antimicrobial. In this study, the mutagenic and antimutagenic activities of S. crassifolia stem bark fractions (hexane, ethyl acetate and hydroalcoholic) were evaluated by the Ames mutagenicity assay in Salmonella typhimurium TA98 and TA100 strains. By the obtained results, all S. crassifolia fractions did not significantly increase the number of prototrophic revertants for histidine (His+) in both S. typhimurium strains tested (p > 0.05), suggesting absence of mutagenicity. Regarding antimutagenicity, the fractions ethyl acetate and hydroalcoholic significantly decreased the number of His+ revertants colonies induced by positive control for strain TA98 (p < 0.05), demonstrating protection against mutagenicity induced by 4-nitroquinolile1-oxide, whereas the hexane fraction did not show antimutagenic effect in this strain. In the TA100 strain, all fractions of S. crassifolia protected DNA against the harmful action of sodium azide, and the hexane fraction exhibited the greatest protection in this work. Thus, it's possible conclude that the fractions of S. crassifolia tested in this study could be used in chemoprevention.

Resumo Salacia crassifolia (Mart. Ex. Schult.) G. Don. é uma árvore que pertence à família Celastraceae e ocorre especialmente no Cerrado Brasileiro. Suas folhas, caule, sementes e frutos são popularmente utilizados para vários fins medicinais, tais como antitumoral, antirreumático, anti-inflamatório e antimicrobiano. Neste estudo, nós avaliamos as atividades mutagênica e antimutagênica de frações da casca do caule de S. crassifolia (hexânica, acetato de etila e hidroalcoólica) pelo ensaio de mutagenicidade de Ames em Salmonella typhimurium, cepas TA98 e TA100. Pelos resultados obtidos todas as frações de S. crassifolia não aumentaram significativamente o número de revertentes prototróficas para histidina (His+) em ambas as cepas de S. typhimurium testadas (p > 0.05), sugerindo ausência de mutagenicidade. Em relação à antimutagenicidade, as frações acetate de etila e hidroalcoólica reduziram significativamente o número de colônias revertentes His+ induzidas pelo controle positive para a cepa TA98 (p < 0.05), demonstrando sua ação protetora contra a mutagenicidade induzida por 4-nitroquinolile1-oxide, enquanto a fração hexânica não demonstrou efeito antimutagênico nesta cepa. Na cepa TA100, todas as frações de S. crassifolia protegeram o DNA contra a ação lesiva de azida sódica, e a fração hexânica exibiu a maior proteção desse trabalho. Assim, concluímos que as frações de S. crassifolia testadas neste estudo poderiam ser utilizadas em quimioprevenção.

Analysis of radioprotection and antimutagenic effects of Ilex paraguariensis infusion and its component rutin

DNA repair pathways, cell cycle checkpoints, and redox protection systems are essential factors for securing genomic stability. The aim of the present study was to analyze the effect of Ilex paraguariensis (Ip) infusion and one of its polyphenolic components rutin on cellular and molecular damage induced by ionizing radiation. Ip is a beverage drank by most inhabitants of Argentina, Paraguay, Southern Brazil, and Uruguay. The yeast Saccharomyces cerevisiae (SC7Klys 2-3) was used as the eukaryotic model. Exponentially growing cells were exposed to gamma rays (γ) in the presence or absence of Ip or rutin. The concentrations used simulated those found in the habitual infusion. Surviving fractions, mutation frequency, and DNA double-strand breaks (DSB) were determined after treatments. A significant increase in surviving fractions after gamma irradiation was observed following combined exposure to γ+R, or γ+Ip. Upon these concomitant treatments, mutation and DSB frequency decreased significantly. In the mutant strain deficient in MEC1, a significant increase in γ sensitivity and a low effect of rutin on γ-induced chromosomal fragmentation was observed. Results were interpreted in the framework of a model of interaction between radiation-induced free radicals, DNA repair pathways, and checkpoint controls, where the DNA damage that induced activation of MEC1 nodal point of the network could be modulated by Ip components including rutin. Furthermore, ionizing radiation-induced redox cascades can be interrupted by rutin potential and other protectors contained in Ip.

Antigenotoxicity protection of Carapa guianensis oil against mitomycin C and cyclophosphamide in mouse bone marrow

ABSTRACT The aim of this study was to evaluate the possible protective of C. guianensis oil against MMC and CP, which are direct- and indirect-acting chemical mutagens, using the micronucleus test. Three experiments were performed. First the C. guianensis oil was co-administered to mice at doses of 250, 500 and 1000 mg/kg bw with 4 mg/kg bw MMC or 50 mg/kg bw CP. Second, the mutagenic drug (CP) was administered ip 50 mg/kg bw and after 6 and 12 hours 250 and 500 mg/kg bw of C. guianensis oil were administered. In the last, C. guianensis oil was administrated (250 and 500 mg/kg bw) during five days and after it was administered ip 50 mg/kg bw CP. The results obtained showed that the C. guianensis oil is not cytotoxic neither genotoxic to mouse bone marrow. Regarding the antimutagenic effect, all doses of C. guianensis oil were significantly (p < 0.05) effective in reducing the frequency of micronucleated polychromatic erythrocytes, when compared with MMC or CP alone. Based on these results, our results suggest that the C. guianensis oil shows medicinal potential as an antimutagenic agent, modulating the mutagenicity caused by both direct- and indirect-acting chemical mutagens, in a mammalian model.

Efeito antimutagênico, antigenotóxico, antiobesidade e antioxidante da Ilex paraguariensis (erva-mate) / Antimutagenic, antigenotoxic, antiobesity and antioxidante effects of Ilex paraguariensis (yerba mate)

O extrato aquoso de erva-mate, obtido a partir de folhas secas de Ilex paraguariensis, é uma bebida amplamente consumida na América do Sul. Inicialmente, nosso objetivo foi caracterizar os compostos presentes nas amostras de erva-mate disponíveis no mercado brasileiro (CH: chimarrão; T: chá mate torrado; G: chá mate torrado, comercialmente acondicionado em garrafas ou C: em copos; TS: chá mate torrado solúvel A mutagenicidade, citotoxicidade e antimutagenicidade de todas as amostras também foram avaliadas atavés do Teste de Ames na presença e na ausência de ativação metabólica. Em seguida, analisamos a amostra TS (2,5, 5,0 e 10 mg/mL) quanto a sua atividade antioxidante e antigenotóxica. Além disso, avaliamos também os efeitos da amostra TS sobre a sinalização da leptina e da insulina no hipotálamo e o estresse oxidativo hepático de ratos adultos obesos programados pela superalimentação neonatal (S). Para induzir S, o tamanho da ninhada foi reduzido a três filhotes por lactante e as ninhadas com número padrão de filhotes (dez/lactante) foram utilizadas como controle. Aos 150 dias de vida, as proles S foram subdivididas em: TS - tratados com extrato aquoso de erva-mate (1g/kg de peso corporal/dia, por gavagem) e S - recebendo água por gavagem durante 30 dias. A prole controle (C) também recebeu água nas mesmas condições do grupo S. Em nossos resultados, verificamos a presença de ácido clorogênico, cafeína e teobromina em todas as amostras analisadas. O conteúdo de compostos fenólicos nas infusões estudadas foram CH: 5,14±0,23; T: 4,33±0,01; G: 0,93±0,25; C: 0,80±0,3 e TS: 8,35±0,5 mg/ml. Não observamos efeito mutagênico ou citotóxico nas amostras analisadas. Um efeito antimutagênico significativo foi observado para a cepa TA97 (pré-, co- e pós-tratamento), na presença de ativação metabólica, em todas as amostras testadas. A amostra TS também apresentou um efeito antimutagênico significativo para a TA102 (pré-, co-e e pós-tratamento)...

Yerba mate extract, made from dried leaves of Ilex paraguariensis, is a tea-like beverage consumed in South America. Here, we aim, firstly, to characterize the compounds present in yerba mate beverages available in the Brazilian market (chimarrao –CH; mate tea –T; mate tea commercially packed in bottles –B and cups ready-to-drink –C; and roasted yerba mate soluble tea –TS). We also evaluated its mutagenic, cytotoxic and antimutagenic properties through Ames test in the presence and absence of metabolic activation. Then, we analyzed exclusively TS sample (2.5, 5.0 and 10 mg/mL) as its antioxidant activity, on plasmid DNA cleavages and upon epithelial esophagus primary culture of Wistar rats treated with a genotoxic agent. Futhermore, we evaluated the effects of mate tea (TS) upon the hypothalamic changes of leptin and insulin signaling and hepatic oxidative stress in postnatal early overfeeding (S) rats. To induce S, litter size was reduced to three pups per dam and normal litters (ten pups/dam) were used as control. In postnatal day (PN) 150, S offspring were subdivided into: S and TS groups treated, respectively, with water or mate tea (1g/kg BW/day, by gavage) during 30 days. C offspring received water. Our results showed the presence of chlorogenic acid, caffeine and theobromine in all analyzed samples. The contents of phenolic compounds in the studied infusions were CH:5,14±0,23; T:4,33±0,01; B:0,93±0,25; C:0,80±0,3 and TS:8,35±0,5 mg/mL. No mutagenic or cytotoxic effect was observed for all analyzed samples. A significant antimutagenic effect was observed for S.typhimurium TA97 (CoT, PreT and PosT), in the presence of metabolic activation, for all tested samples. TS sample also exhibited a significant antimutagenic effect for S.typhimurium TA102 (CoT, PreT and PosT) in the presence of metabolic activation. We did not observed an antimutagenic effect using TA98, TA100 and TA102 for CH, T, C and B samples; and no effect for TA98 and TA100 for TS sample...

The mutagenic and antimutagenic effects of the traditional phytoestrogen-rich herbs, Pueraria mirifica and Pueraria lobata

Pueraria mirifica is a Thai phytoestrogen-rich herb traditionally used for the treatment of menopausal symptoms. Pueraria lobata is also a phytoestrogen-rich herb traditionally used in Japan, Korea and China for the treatment of hypertension and alcoholism. We evaluated the mutagenic and antimutagenic activity of the two plant extracts using the Ames test preincubation method plus or minus the rat liver mixture S9 for metabolic activation using Salmonella typhimurium strains TA98 and TA100 as indicator strains. The cytotoxicity of the two extracts to the two S. typhimurium indicators was evaluated before the mutagenic and antimutagenic tests. Both extracts at a final concentration of 2.5, 5, 10, or 20 mg/plate exhibited only mild cytotoxic effects. The plant extracts at the concentrations of 2.5, 5 and 10 mg/plate in the presence and absence of the S9 mixture were negative in the mutagenic Ames test. In contrast, both extracts were positive in the antimutagenic Ames test towards either one or both of the tested mutagens 2-(2-furyl)-3-(5-nitro-2-furyl)-acrylamide and benzo(a)pyrene. The absence of mutagenic and the presence of anti-mutagenic activities of the two plant extracts were confirmed in rec-assays and further supported by a micronucleus test where both plant extracts at doses up to 300 mg/kg body weight (equivalent to 16 g/kg body weight plant tuberous powder) failed to exhibit significant micronucleus formation in rats. The tests confirmed the non-mutagenic but reasonably antimutagenic activities of the two plant extracts, supporting their current use as safe dietary supplements and cosmetics.

Dietary spices in health and diseases: I.

Spices are heterogeneous collections of a wide variety of volatile and non-volatile staple dietary additives. India with its wide climatic conditions and topographical features naturally possesses wide variety of medicinal flora. Spices have a diverse array of natural phytochemicals that have complementary and overlapping actions, including antioxidant effects, modulation of detoxification enzymes, stimulation of immune system, reduction of inflammation, modulation of steroid metabolism and antibacterial and antiviral effects. In the present essay, various studies on effects of different well characterized spices on molecular, cellular, autocrine, paracrine and endocrine mechanisms and their role in neuromodulation, immunoinodulation, anti inflammatory, antioxidant, anti-carcinogenic, antimutagenic and psychoactive phenomena have been reviewed.

Assessment of the mutagenic, antimutagenic and cytotoxic activities of ethanolic extract of araticum (Annona crassiflora Mart. 1841) by micronucleus test in mice

A typical Brazilian plant, araticum (Annona crassiflora Mart.), is widely used in humans as therapeutic medicine to treat several diseases such as diarrhea, rheumatism and syphilis. It contains acetogenins which present cytotoxic, antitumogenic, and antiparasitic properties. In this study, mutagenic, antimutagenic and cytotoxic effects of araticum leaves ethanolic extract were evaluated by micronucleus test in mice. To evaluate the mutagenic activity, animals were treated with ethanolic extract of araticum (EEA) using 10, 20, 50, 100 and 160 mg.kg-1. For all doses, micronucleated polychromatic erythrocytes (MNPCE) frequency was evaluated at 24, 48 and 72 hours after treatment. To evaluate the antimutagenic activity, animals were treated with 10, 20, 50 and 100 mg.kg-1 of EEA and 4 mg.kg-1 of MMC simultaneously. The frequency of MNPCE was evaluated 36 hours after exposure. Cytotoxicity was evaluated by the polychromatic and normochromatic erythrocytes ratio (PCE/NCE). In the mutagenicity assessment, all doses of EEA resulted in no significant increase of MNPCE (P > 0.05), compared to solvent- control group. Regarding administration time, no significant difference among three evaluation periods was observed (P > 0.05). Such results indicate that EEA did not exert mutagenic activity. Cytotoxicity was evident in doses of 50, 100 and 160 mg.kg-1 at 24 and 48 hours after exposure. Concerning antimutagenicity, except the 10 mg.kg-1 co-administered with 4 mg/kg of MMC, all doses reduced significantly the frequency of MNPCE compared to the positive control group (P < 0.05). These results, therefore, indicate an antimutagenic activity of the EEA. Cytotoxicity was significantly increased (P < 0.01) at 100 mg.kg-1 EEA doses co-administered with 4 mg.kg-1 of MMC.

O araticum (Annona crassiflora Mart.) é uma planta tipicamente brasileira, largamente utilizada em humanos como remédio para o tratamento de diversas doenças como diarréia, reumatismo e sífilis. Esta planta contém acetogeninas que apresentam propriedades citotóxica, antitumorigênica e antiparasitária. Neste estudo, foram avaliados os possíveis efeitos mutagênico, antimutagênico e citotóxico do extrato etanólico de folhas de araticum, pelo teste de micronúcleos em camundongos. Para a investigação da atividade mutagênica, os animais foram tratados com o extrato etanólico de araticum (EEA) utilizando 10, 20, 50, 100 e 160 mg.kg-1. Para todas as doses, as freqüências de eritrócidos policromáticos micronucleados (MNPCE) foram avaliadas em 24, 48 e 72 horas após o tratamento. Para a investigação da atividade antimutagênica, os animais foram tratados com 10, 20, 50 e 100 mg.kg-1 de EEA simultaneamente com 4 mg.kg-1 de MMC. A freqüência de MNPCE foi avaliada após 36 horas de exposição. A citotoxicidade foi avaliada pela razão de eritrócitos policromáticos e normocromáticos (PCE/NCE). Na avaliação da mutagenicidade, todas as doses de EEA não aumentaram significativamente o número de MNPCE (P > 0,05), comparativamente as do grupo solvente-controle. Em relação ao tempo de administração, não foi constatada diferença significativa entre os 3 períodos avaliados (P > 0,05). Esses resultados indicam que o EEA não exerceu atividade mutagênica.A citotoxicidade foi evidente nas doses de 50, 100 e 160 mg.kg-1 em 24 e 48 horas depois da exposição. Em relação à antimutagenicidade, exceto para a dose de 10 mg.kg-1 co-administrada com 4 mg.kg-1 de MMC, todas reduziram significativamente a freqüência de MNPCE, comparativamente as do grupo controle positivo (P < 0,05). Esses resultados, portanto, indicam uma atividade antimutagênica do EEA. A citotoxicidade foi significativamente aumentada (P < 0,01) na dose de 100 mg.kg-1 de EEA co-administrada com 4 mg.kg-1 de MMC.

Absence of antimutagenicity of Cochlospermum regium (Mart. and Schr.) Pilger 1924 by micronucleus test in mice

Cochlospermum regium (Mart. and Schr.) Pilger, popularly known as "algodãozinho do campo", is a medicinal plant that grows in the Cerrado of Brazil. This plant has been used in traditional medicine against various diseases such as leucorrhoea, gastritis and ulcers. It has also been effective in treating skin problems like pimples, boils and blotches. In the present study, the in vivo antimutagenicity of aqueous extract of C. regium was evaluated. The Micronucleus Test was performed in polychromatic erythrocytes from Swiss male mice treated with one of the four doses of extract of the plant (19, 38, 76 and 114 mg.kg-1 body weight), administered by intraperitonial injection (i.p.) simultaneously with cyclophosphamide (24 mg.kg-1 b.w.) or mitomycin C (4 mg.kg-1 b.w.). The cytotoxicity was evaluated by polychromatic and normochromatic erythrocytes ratio (PCE/NCE). The results showed no significant reduction of the micronucleated polychromatic erythrocytes frequency (P > 0.05). In conclusion, the data indicate that C. regium roots aqueous extract, for the conditions used, did not exhibit the antimutagenic effect.

Cochlospermum regium (Mart. & Schr.) Pilger, conhecido popularmente como "algodãozinho-do-campo", é uma planta medicinal que cresce no Cerrado brasileiro. Esta planta tem sido utilizada na medicina tradicional contra várias doenças como leucorréia, gastrites e úlceras. Esta também tem se mostrado efetiva no tratamento de doenças de pele como espinhas, furúnculos e manchas. No presente estudo, foi avaliada a antimutagenicidade do extrato aquoso de C. regium pelo Teste do Micronúcleo in vivo. Este ensaio foi realizado em eritrócitos policromáticos de camundongos machos Swiss tratados com quatro concentrações diferentes do extrato da planta (19, 38, 76 e 114 mg.kg-1 por peso corpóreo), administrado por injeção intraperitonial (i.p.) simultaneamente com ciclofosfamida (24 mg.kg-1 p.c.) ou mitomicina C (4 mg.kg-1 p.c.). A citotoxicidade foi avaliada pela razão eritrócitos policromáticos e normocromáticos (PCE/NCE). Os resultados obtidos não mostraram redução significativa da freqüência de eritrócitos policromáticos micronucleados (P > 0,05). Em conclusão, os resultados indicam que o extrato aquoso de raiz de C. regium, para as condições utilizadas, não exibiu efeito antimutagênico.

Cytotoxic, mutagenic and antimutagenic screening of Arenosclera brasiliensis acetone and ethanol extracts

The marine environment is a rich source of biologically active compounds with pharmacological properties. Marine organisms often produce secondary metabolites with structural features different from those produced by terrestrial ones, and the Phylum Porifera seems to be one of the most productive in this sense. This study was undertaken to provide data on mutagenic and antimutagenic activities from an acetone (Areac) and an ethanol (Areet) extract obtained from Arenosclera brasiliensis, an endemic Brazilian sponge. A qualitative Salmonella reverse mutation test was performed with the TA97, TA98, TA100, and TA102 strains by incubating cells with Areac and Areet in the presence and absence of a known mutagen. A cytotoxic evaluation of the extracts was also performed. A. brasiliensis did not display any mutagenic activity, but Areac showed significant toxicity against test strains. In the antimutagenic assay, a reduction in the number of his+ revertants was observed for the TA97, TA100 and TA102 strains treated with Areac when compared to the positive controls. Areet treatment showed protective activity against DNA lesions only for the TA100. These results are in agreement with those obtained previously with other A. brasiliensis extracts, suggesting an antimutagenic activity.

Antimutagenic effects of piperine on cyclophosphamide-induced chromosome aberrations in rat bone marrow cells.

Piperine is a major pungent substance and active component of black pepper (Piper nigrum Linn.) and long pepper (Piper longum Linn.). Both plants are used worldwide as household spices and condiments. They are also used as important ingredients in folklore medicine in many Asian countries. Therefore, it is of interest to study antimutagenic effects of piperine. In this study, its influence on chromosomes was investigated in rat bone marrow cells. Male Wistar rats were orally administered piperine at the doses of 100, 400 and 800 mg/kg body weight for 24 hours then challenged with cyclophosphamide at a dose of 50 mg/kg body weight by intraperitoneal injection. Twenty-four hours thereafter, all animals were sacrificed and bone marrow samples were collected for chromosomal analysis. The results demonstrated that piperine at a dose of 100 mg/kg body weight gave a statistically significant reduction in cyclophosphamide-induced chromosomal aberrations. In conclusion, piperine may have antimutagenic potential. The underlying molecular mechanisms now require attention.

Ginseng extract exhibits antimutagenic activity against induced mutagenesis in various strains of Salmonella typhimurium.

Ginseng has been reported to exhibit antioxidant and antimutagenic activity. The present study was undertaken with a view to confirm whether the antioxidant activity of Ginseng is responsible for its antimutagenic action. The concentrated root extract of Panax ginseng (Ginseng extract I) and its lyophilized powder (Ginseng extract II) obtained from two different manufacturing houses, were tested against mutagenesis using the well-standardized Ames microsomal test system. The extracts exhibited antimutagenic effect against hydrogen peroxide induced mutagenesis in TA100 strain, and against mutagenesis produced by 4-nitroquinoline-N-oxide in both TA98 and TA100 strains of Salmonella typhimurium. Both the extracts failed to show any antimutagenic potential against tert-butyl hydroperoxide (an oxidative mutagen) in TA102 strain, a strain highly sensitive to active oxygen species. The extracts also indicated a weak antioxidant activity in a series of in vitro test systems viz., 1,1-diphenyl picryl hydrazyl (DPPH) assay, hydrogen peroxide scavenging and superoxide anion scavenging. The results indicate that the protective effects shown by ginseng extract(s) against 4-nitroquinoline-n-oxide and hydrogen peroxide induced mutagenesis in TA98 and TA100 could mainly be due to its property to initiate and promote DNA repair rather than free radical scavenging action.

Mutagenicity study of weeds and common plants used in traditional medicine and for animal feed.

Mutagenicity and antimutagenicity potentials were tested using Ames' test in crude distilled water and absolute ethanol extracts from the stems and leaves of Peperomia pellucida (Linn.) Kunth, Eichhornia crassipes Solms, Colocasia esculenta Schott and Brachiaria mutica (Forssk.) Stapf, and the stems of Musa sapientum Linn. No mutagenic effect was found in any of the 10 mg/plate crude extracts of these plants for either TA98 or TA100 of Salmonella typhimurium, in a direct test and a mutagenic induced test by S-9 mix. Both distilled water and absolute ethanol extract of 0.5-10 mg/plate B. mutica showed strong antimutagenicity to AFB1, B(a)P and 4NQO in two tester strains. Ethanol extract of 0.1-0.5 mg/plate C. esculenta also showed antimutagenicity to AFB1, B(a)P and 4NQO in two tester strains, but the 0.5-10 mg/plate water extract had an antimutagenic effect only for B(a)P in TA98. The ethanol extracts of 5 mg/plate B. mutica and 0.5 mg/plate C. esculenta are cytotoxic, as indicated by their partial killing effect.

Antimutagenic activity of Sesbania javanica Miq. flower DMSO extract and its major flavonoid glycoside.

The antimutagenic activity of Sesbania javanica Miq. or Sano, an edible vegetable flower DMSO extract against aflatoxin B1 (AFB1) benzo (a) pyrene [B(a)P], was evaluated by means of the Ames' test. The Sesbania javanica Miq. flower DMSO extract showed a strong inhibitory effect against AFB1 and B(a)P mutagens. A search to isolate the major flavonoid in Sesbania javanica Miq. flower extract found the flavonol glycoside, Quercetin 3-2(G)-rhamnosylrutinoside, which was confirmed by its physicochemical properties as a major constituent of the flower. Quercetin 3-2(G)-rhamnosylrutinoside (207 microg/plate) also showeda strong inhibitory effect against AFB1 and B(a)P with a more than 70% inhibition rate.

Antigenotoxic and antimutagenic potential of an annatto pigment (norbixin) against oxidative stress

Carotenoids are 40-carbon molecules with conjugated double bonds, making them particularly effective for quenching free radicals. They have always been believed to possess anticancer properties, which could be due to their antioxidant potential. Norbixin is an unusual dicarboxylic water-soluble carotenoid present as a component in the pericarp of the seeds of Bixa orellana L. (from the Bixaceae family), a tropical shrub commonly found in Brazil. The main carotenoids present in these seeds, bixin and norbixin, form a coloring material, known as annatto, which is mainly used in the food industry. As annatto is only used as a coloring material, most studies of annatto pigments have focused on the determination of annatto levels in food. However, little attention has been given to the biological properties of bixin and norbixin. We evaluated the effect of norbixin on the response of Escherichia coli cells to DNA damage induced by UV radiation, hydrogen peroxide (H2O2) and superoxide anions (O2*-)) and found that norbixin protects the cells against these agents. Norbixin enhanced survival at least 10 times. The SOS induction by UVC was inhibited 2.3 times more when cells were grown in the presence of norbixin. We also found that norbixin has antimutagenic properties, with a maximum inhibition of H2O2-induced mutagenic activity of 87%, based on the Salmonella mutagenicity test

Antimutagenic and antioxidant/prooxidant activity of quercetin.

The present study has been performed to evaluate the antimutagenic activity of quercetin, ascorbic acid and their combination against an oxidative mutagen. An effort was also made to correlate this activity to the in vitro antioxidant activity of these agents. Antimutagenicity testing was done in Ames Salmonella Assay system using Salmonella typhimurium TA102 against t-butylhydroperoxide as an oxidative mutagen. In vitro antioxidant scavenging activity was tested for DPPH free radical, superoxide anion, hydrogen peroxide and hydroxyl radical in their specific test systems. Quercetin (0.5-8 nmole/plate) and ascorbic acid (0.1-100 micromole/plate) showed significant effect. Quercetin (4 and 8 nmole/plate) when combined with ascorbic acid (500 nmole/plate) showed an increase in the antimutagenic activity. In vitro antioxidant activity of quercetin was better than ascorbic acid in all the test systems used. The study indicated that the antimutagenic activity of quercetin was not solely accountable by its antioxidant nature. However, in vitro free radical scavenging activity of quercetin correlated well with the antimutagenic activity.

Antimutagenic effects of black tea in the Salmonella typhimurium reverse mutation assay.

Black tea (Camellia sinensis) is one of the most widely consumed beverages worldwide. Its chemopreventive effects are well documented in the literature. In the present set of investigations antimutagenic effects of aqueous black tea extract (ATE) and black tea polyphenols (BTP) were evaluated in the Ames test using Salmonella typhimurium tester strains TA 98 and TA 100. Addition of benzo(a)pyrene (BaP) and cyclophosphamide (CP), two well known mutagens, at the concentrations of 20 and 15 microg/plate, respectively, in an S-9 metabolically activated system resulted in significant induction of his+ revertant colonies. However, addition of 500 microl 1, 2 and 4% ATE to the BaP and CP treated plates resulted in a dose dependent inhibition in the number of his+ revertant colonies. Furthermore in another set of experiments, supplementation with BTP at the concentrations of 100, 200 and 400 microg/plate also led to a significant inhibition in BaP and CP induced colony formation. The antimutagenic activity of BTP was found to be higher than that of ATE, which may be attributable to the higher amount of polyphenolic ingredients. Hence the study revealed that black tea has a protective efficacy in suppressing BaP and CP induced mutagenicity in a microbial test system.

Inhibitory effect of curcumin and its natural analogues on genotoxicity of heterocyclic amines from cooked food.

Curcumin (C) and its natural analogues demethoxycurcumin (dmC) and bisdemethoxycurcumin (bdmC), known for their potent anti-inflammatory, antioxidant, antimutagenic and anticarcinogenic effects, were tested for their possible inhibitory effects against seven cooked food mutagens (heterocyclic amines): 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1), in both TA98 and TA100 strains of Salmonella typhimurium using Ames Salmonella/reversion assay in the presence of Aroclor induced rat liver S9 homogenate. In the present investigations, curcumin as well as its two natural analogues i.e., dmC and bdmC were found to be highly effective in suppressing genotoxicity of all the tested cooked food mutagens in a dose-dependent manner, in both the frame shift (TA98) as well as base pair mutation sensitive (TA100) strains of S. typhimurium. However, bdmC appeared to be a relatively less active antimutagen compared to C and dmC. More than 80% inhibition of mutagenicity was observed at 200 microg/plate in case of C and dmC in both TA98 and TA100 against all tested cooked food mutagens. Where as, bdmC showed 39-79% inhibition in TA100 and 60-80% inhibition in TA98, at a dose of 200 microg/plate. These findings warrant further biochemical, enzymatic and in vivo investigations in animal models as well as in humans to establish the chemoprotective effect of these agents against mutagenic heterocyclic amines found in cooked food.

An anticlastogenic in vivo micronucleus assay for tea.

Common use of antimutagens and anticarcinogens in everyday life is an effective measure for preventing human cancer and genetic diseases. Antioxidant properties of tea have vast potential as protective agents against diverse toxic effects. The present study was aimed to evaluate the role of aqueous clonal tea extracts (green tea, oolong tea and black tea) in modulating the genotoxic damage induced by cyclophosphamide (CP), a commonly used chemotherapeutic drug and a well-known mutagen and clastogen. All the three tea extracts at 1 and 2% concentration did not increase the frequency of micronucleated polychromatic erythrocytes (MPE) in bone marrow cells of mice when administered individually. The tea extracts decreased the micronuclei (MN) induced by CP. Therefore, regular intake of tea may improve the antioxidant status in in vivo and thereby reduce the risk of cancer and coronary heart disease.

Efficiency of Coleus aromaticus extract in modifying cyclophosphamide and mitomycin-C induced clastogenicity in mouse bone marrow cells.

The anticlastogenic potency of the ethanolic extract of a medicinal plant, C. aromaticus was investigated by taking bone marrow chromosomal aberration assay and micronucleus (MN) test as the test parameters. Swiss albino mice were fed orally with different doses (10,15, 25, 50 and 100 mg/kg body weight) of ethanolic extract for 7 days and on the 7th day, two doses each of anticancer drugs cyclophosphamide (CP; 25 and 50 mg/kg body weight) and mitomycin-C (MMC; 4 and 8 mg/kg body weight) were injected, ip, to different groups of animals. Bone marrow MN preparations were made at 24 and 48 hr time intervals. Coleus extract reduced CP and MMC induced MN and lower doses of the extract were found to be more effective than higher doses. The effective doses of extract in MN test were selected to study the anticlastogenic effects against CP (25 and 50 mg/kg body weight) and MMC (2 and 4 mg/kg body weight) induced chromosomal aberrations. The results indicate the protective effect of C. aromaticus against CP and MMC induced cytogenetic damage.