Avaliação hematológica e do potencial genotóxico, antigenotóxico e antioxidante do óleo e dos extratos etanólico, hidroetanólico e aquoso da polpa do pequi (Caryocar brasiliense Camb.) em camundongos Swiss

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Saúde, Programa de Pós-Graduação em Patologia Molecular, 2013.Os radicais livres são espécies reativas produzidas sob condições fisiológicas e patológicas nos organismos aeróbios, que podem causar danos às macromoléculas biológicas e participar do desenvolvimento de doenças degenerativas e do processo carcinogênico. O consumo de antioxidantes naturais, a partir da dieta ou da suplementação com nutracêuticos e fitoterápicos, tem-se mostrado eficaz contra os efeitos deletérios das espécies reativas. O pequi (Caryocar brasiliense Camb.) é um fruto típico do bioma cerrado utilizado pela população nativa para fins gastronômicos, nutricionais e medicinais. A polpa do pequi contém diversos antioxidantes naturais como as vitaminas C e E, compostos fenólicos, carotenoides e óleos essenciais que podem atuar na proteção do DNA, lipídeos e proteínas contra os danos causados por espécies reativas. Baseando-se nas características da polpa do pequi, o objetivo do estudo foi avaliar o potencial antioxidante, genotóxico, antigenotóxico e as alterações hematológicas promovidas pelo tratamento com o óleo e os extratos etanólico, hidroetanólico e aquoso da polpa de pequi em camundongos Swiss de ambos os sexos. O quimioterápico doxorrubicina (DX), conhecido por seu potencial em induzir danos oxidativos, foi utilizado para verificar a capacidade protetora do óleo e dos extratos da polpa de pequi contra as lesões promovidas pelos radicais livres (RL). A caracterização fitoquímica das amostras da polpa de pequi confirmou a presença de carotenoides no óleo e no extrato etanólico e de compostos fenólicos nos extratos etanólico, hidroetanólico e aquoso. Os resultados da determinação de macro e micronutrientes demonstraram a presença de minerais nas amostras, porém em diferentes quantidades. O óleo e os extratos apresentaram atividade antioxidante in vitro no ensaio DPPH, sendo a do óleo a menos eficiente entre as amostras da polpa do pequi analisadas. Entre os extratos etanólico, hidroetanólico e aquoso não houve diferenças significativas em relação à atividade antioxidante in vitro. A atividade antioxidante in vivo foi analisada no plasma dos camundongos tratados pelo ensaio TBARS, porém os resultados obtidos sofreram grande interferência da hemoglobina presente no plasma devido à hemólise durante o processo de obtenção das amostras a serem analisadas. Em relação às alterações hematológicas, apenas o tratamento com o extrato aquoso nos machos induziu ao decréscimo de parâmetros do eritrograma, provavelmente devido às saponinas hemolíticas no extrato. O tratamento associado a DX promoveu alterações nos parâmetros do hemograma e do leucograma dos camundongos tratados devido à peroxidação lipídica e degradação de membranas induzidas pela DX e incapacidade dos extratos em preveni-las. Os efeitos genotóxicos e mutagênicos dos tratamentos foram avaliados por meio do teste do cometa e do teste do micronúcleo nas células de medula óssea, respectivamente. Apenas o tratamento com o extrato etanólico foi genotóxico nas fêmeas, porém não foi mutagênico. O tratamento associado a DX foi mutagênico e citotóxico, evidenciado que óleo e os extratos não foram capazes de prevenir os efeitos tóxicos da DX na dose administrada. O possível mecanismo pelo qual as amostras da polpa de pequi foram incapazes de proteger o organismo contra os efeitos da DX, deve- se, possivelmente, ao conteúdo de metais de transição no óleo e nos extratos, principalmente o ferro, que reagem com a molécula de DX e através de reações de oxirredução formam o potente radical livre semiquinona, capaz de interagir com o DNA e outras macromoléculas biológicas. ______________________________________________________________________________ ABSTRACTFree radicals are reactive species, produced under physiological and pathological conditions in aerobic organisms, which can cause damage to biological macromolecules and participate in the development of degenerative diseases and the carcinogenic process. Consumption of natural antioxidants from diet or supplementation with herbal drugs and nutraceuticals has been shown to be effective against the deleterious effects of reactive species. Pequi (Caryocar brasiliense Camb.) is a typical fruit of the Cerrado and is used by the native population for culinary, nutritional and medicinal purposes. Pequi pulp contains many natural antioxidants such as vitamins C and E, phenolic compounds, carotenoids and essential oils that can act in protecting DNA, lipids and proteins against the damage caused by reactive species. Based on pequi pulp characteristics, the aim of the study was to evaluate the antioxidant, genotoxic, antigenotoxic potential and hematological alterations promoted by oil or ethanolic, hydroethanolic and aqueous extracts of pequi pulp treatment in Swiss mice of both sexes. The chemotherapeutic doxorubicin (DX), known for its potential to induce oxidative damage, was used to verify the protective capacity of the oil and extracts against injury promoted by free radicals. Phytochemical characterization of the samples confirmed the presence of carotenoids in oil and ethanol extract and phenolic compounds in ethanolic, hydroethanolic and aqueous extracts. Determination of macro and micronutrients showed the presence of minerals in the samples, but in different quantities. Pequi pulp oil and extracts exhibited antioxidant activity in the DPPH in vitro assay, but the oil was less efficient than the extracts. Among the ethanolic, hydroethanolic and aqueous extracts there were no significant differences. The in vivo antioxidant activity was analyzed in plasma of treated mice by TBARS assay, but the results underwent significant interference of hemoglobin present in the plasma due to hemolysis during the process of obtaining the samples to be analyzed. Regarding hematological values, only the treatment with the aqueous extract in males induced a decrease in erythrocyte parameters, probably due to hemolytic saponins in the extract. The treatment associated with DX promoted changes in the parameters of the CBC and leukocyte counts of treated mice, due to lipid peroxidation and degradation of membranes induced by DX and inability of extracts to prevent them. The genotoxic and mutagenic effects of the treatments were evaluated by the comet assay and micronucleus test in bone marrow cells, respectively. Only treatment with the ethanol extract was genotoxic in females, but was non-mutagenic. The treatment associated with DX was mutagenic and cytotoxic and showed that the pequi pulp oil and extracts were not able to prevent the toxic effects of the high dose of DX. The possible mechanism by which Pequi pulp samples were unable to protect against the effects of DX is due to the content of transition metals in oil and extracts, mainly iron, which react with the DX molecule by redox reactions that form the potent free radical semiquinone capable of interacting with DNA and other biological macromolecules

The pequi pulp oil (Caryocar brasiliense Camb.) provides protection against aging-related anemia, inflammation and oxidative stress in Swiss mice, especially in females

Continued exposure to reactive oxygen species and inflammation are the rationale behind aging theories and associated diseases. Scientific evidence corroborates the ethnomedicinal use of the oil of pequi (Caryocar brasiliense Camb.), a typical Brazilian Cerrado fruit, against oxidative damage to biomolecules and inflammation. We aimed to investigate in vivo the antioxidant and anti-inflammatory effects of pequi oil on hemogram and DNA damage in healthy young adult and older middle-aged Swiss mice of both genders. Animals, aged 6-7 and 11-12 months, were orally treated for 15 days with pequi oil at 30 mg/day. Blood samples were used for hemogram and comet assay, and bone marrow for micronucleus test. Female controls of 11-12 months had significantly lower haemoglobin and hematocrit than those of 6-7 months. Treatment with pequi oil improved this state, removing the differences. Pequi oil had no genotoxic or clastogenic effects and significantly increased lymphocytes and decreased neutrophils+monocytes in females of 11-12 months, removing the significant differences observed between controls of 6-7 and 11-12 months. The results suggest that dietary supplementation with pequi oil could protect against anemia, inflammation and oxidative stress related to aging, helping to prevent aging-related chronic degenerative diseases, mainly for females

The lipidome, genotoxicity, hematotoxicity and antioxidant properties of andiroba oil from the Brazilian Amazon

Andirobeira is an Amazonian tree, the seeds of which produce a commercially valuable oil that is used in folk medicine and in the cosmetic industry. Andiroba oil contains components with anti-inflammatory, cicatrizing and insect-repellant actions. However, virtually nothing is known of the safety of this oil for humans. The aim of this work was therefore to investigate the hematotoxicity, genotoxicity and mutagenicity of andiroba oil using the comet and micronucleus assays, and to assess its antioxidant properties and lipidome as a means of addressing safety issues. For the experiments, andiroba oil was administered by gavage for 14 consecutive days in nulliparous female Swiss mice randomly distributed in four groups: negative control and three doses of oil (500, 1000 and 2000 mg/kg/day). These doses were chosen based on recommendations of the OECD guideline no. 474 (1997). GC/MS was used to investigate the free fatty acid, cholesterol and triterpene content of andiroba oil in a lipidomic analysis. No clinical or behavioral alterations were observed throughout the period of treatment, and exposure to andiroba oil at the doses and conditions used here did not result in hematotoxic, genotoxic or mutagenic effects. Tests in vitro showed that oil sample 3 from southwestern of Brazilian Amazon had a high antioxidant capacity that may protect biological systems from oxidative stress, although this activity remains to be demonstrated in vivo