Effect of Silymarin on oxidative damage in chicken liver cell membranes / Efecto de la silimarina sobre el daño oxidativo en membranas celulares de hígado de pollo

ABSTRACT In living beings, antioxidants are of vital importance for protection against oxidative damage caused by reactive oxygen species. Silymarin (SM), a plant-derived flavonoid present in the fruits and seeds of milk thistle Silybum marianum (L.) Gaertn., has a recognized hepatoprotective effect. In this work, the in vitro silymarin antioxidant effect on non-enzymatic peroxidation (NEP) in chicken liver mitochondria and microsomes was studied. Oxidative stress in the organelles was induced by subjecting the samples (1 mg of protein) to an ascorbate-Fe++-dependent prooxidant system at 37 °C. Oxidative damage was quantified by chemiluminescence (CL) using a Packard1900 TR liquid scintillation counter (Meriden CT, USA). CL expressed as cpm (counts per minute) was read every 10 minutes to establish the course of peroxidation as a function of time. Likewise, the total cpm value (sum of the readings) was used to compare the inhibitory effect of SM using different concentrations corresponding to 6.25; 12.5, and 25 µg of the active ingredient (silymarin phosphatide) per mg of mitochondrial and microsomal protein. Controls were run simultaneously without the addition of ascorbate. Peroxidation inhibition was dependent on the concentration of SM in the incubation mixture. The results show that a protective effect on induced oxidative damage was found for all concentrations tested.

RESUMEN En los seres vivos, los antioxidantes son de vital importancia para la protección contra el daño oxidativo causado por las especies reactivas del oxígeno. La silimarina (SM), un flavonoide de origen vegetal presente en los frutos y semillas del cardo mariano Silybum marianum (L.) Gaertn., tiene un reconocido efecto hepatoprotector. En este trabajo se estudió el efecto antioxidante in vitro de la silimarina sobre la peroxidación no enzimática (PNE) en mitocondrias y microsomas de hígado de pollo. El estrés oxidativo en los orgánulos se indujo sometiendo las muestras (1 mg de proteína) a un sistema prooxidante dependiente de ascorbato-Fe++ a 37 °C. El daño oxidativo se cuantificó por quimioluminiscencia (QL) utilizando un contador de centelleo líquido Packard 1900 TR (Meriden CT, EE. UU.). QL expresado como cpm (recuentos por minuto) se leyó cada 10 minutos para establecer el curso de la peroxidación en función del tiempo. Asimismo, se usó el valor total de cpm (suma de las lecturas) para comparar el efecto inhibitorio de SM mediante diferentes concentraciones correspondientes a 6,25; 12,5 y 25 µg del ingrediente activo (fosfátido de silimarina) por mg de proteína mitocondrial y microsomal. Los controles se realizaron simultáneamente sin la adición de ascorbato. La inhibición de la peroxidación dependía de la concentración de SM en la mezcla de incubación. Los resultados muestran que para todas las concentraciones probadas se encontró un efecto protector sobre el daño oxidativo inducido

Efecto del ácido alfa lipoico sobre la peroxidación no enzimática en mitocondrias de corazón y cerebro de rata / Effect of alpha-lipoic acid against non-enzymatic peroxidation of rat heart and brain mitochondria

RESUMEN El ácido alfa lipoico (AAL) ha sido caracterizado como un antioxidante eficiente. Se ha propuesto como un agente terapéutico potencial en el tratamiento o prevención de diferentes alteraciones que pueden estar relacionadas con un desequilibrio del estado celular oxidoreductor. El objetivo de este trabajo fue analizar la sensibilidad a la peroxidación no enzimática (PNE) (ascorbato-Fe++ dependiente) en mitocondrias de corazón y cerebro de ratas incubadas con una solución de AAL. La PNE fue evaluada por el método de quimioluminiscencia (QL). Cuando se compararon las muestras control (sin el agregado del ascorbato-Fe++) con las muestras ascorbato-Fe++ dependientes, se observó un incremento significativo en la emisión lumínica. Simultáneamente, se incubaron las mitocondrias de ambos órganos con diferentes concentraciones de AAL (0,05, 0,15 y 0,25 mg/ml) observándose una protección diferencial. Las mitocondrias de cerebro de rata incubadas con dosis de 0,15 y 0,25 mg/ml de AAL fueron protegidas de los efectos de la PNE, mientras que, en las mitocondrias cardíacas, solo se observó protección con la dosis más alta de AAL (0,25 mg/ml). El análisis de QL indicó que las mitocondrias de cerebro fueron protegidas de manera más eficiente que las mitocondrias de corazón de rata. En este último caso, será necesario probar nuevas dosis de AAL para demostrar los efectos en estas membranas. En conclusión, AAL actuó como un antioxidante protector de las membranas de ambos órganos contra el daño peroxidativo.

ABSTRACT Alphalipoc acid (ALA) has been characterized as an efficient antioxidant. It has been proposed as a potential therapeutic agent in the treatment or prevention of different pathologies that may be related to an imbalance of the oxido reductive cell state. The objective of this work was to analyze the sensitivity to non-enzymatic peroxidation (NEP) (ascorbate-Fe++ dependent) in heart and brain mitochondria of rats incubated with an ALA solution. NEP was evaluated by the chemiluminescence method (CL). When the control samples (without the addition of ascorbate-Fe++) were compared with the ascorbate-Fe++ dependent samples, a significant increase in the light emission. Simultaneously, the mitochondria of both organs were incubated with different concentrations of ALA (0.05, 0,15 and 0,25 mg/ml), observing a differential protection. Rat brain mitochondria incubated with doses of 0.15 and 0,25 mg/ml of ALA were protected from the effects of NEP, while in cardiac mitochondria, protection was only observed with the highest dose of ALA (0,25 mg/ml). The CL analysis indicated that rat brain mitochondria were protected more efficiently than rat heart mitochondria. In the latter case, it will be necessary to test new doses of ALA to demonstrate the effects on these membranes. In conclusion, ALA acted as a protective antioxidant of the membranes of both organs against peroxidative damage.

Dormência e germinação de sementes de Melaleuca alternifolia Cheel / Dormancy and germination of Melaleuca alternifolia Cheel. seeds

A Melaleuca alternifolia Cheel. é uma espécie aromática de expressivo interesse econômico, em função da presença de óleo essencial armazenado no tecido foliar. As sementes, entretanto, apresentam baixo poder germinativo, o que tem dificultado a obtenção de novos materiais genéticos e o avanço tecnológico para produção no Brasil. O presente trabalho teve por objetivo determinar uma metodologia adequada para avaliar o poder germinativo de sementes de melaleuca. Para tanto, foram realizados dois experimentos, um em condições de laboratório e outro em casa-de-vegetação, avaliando-se diferentes tratamentos para promover a germinação da semente. O delineamento experimental empregado foi o inteiramente casualizado, com quatro repetições, sendo nove tratamentos em laboratório e sete em casa de vegetação; a comparação de médias foi realizada pelo teste Tukey, a 5 por cento de probabilidade. Diante dos resultados obtidos, concluiu-se que a pré-embebição das sementes por 18h, em água, é o procedimento mais adequado, dentre os testados, para avaliar o poder germinativo da semente de melaleuca, sendo que em condições de casa de vegetação, o referido tratamento pode ser usado com os substratos areia ou Plantmax.

Melaleuca alternifolia Cheel. is an aromatic species of high economic interest due to the presence of essential oil in its leaf tissue. However, its seeds have low germinative potential, which has made difficult the obtaining of new genetic materials and the technological advance for production in Brazil. The present work aimed to determine a suitable methodology to evaluate the germinative potential of Narrow-leaved Paperbark seeds. Thus, two experiments were carried out, one under lab conditions and other in greenhouse, in order to evaluate different treatments to stimulate seed germination. Experimental design was completely randomized, with four replicates; nine treatments were used in the lab and other seven in greenhouse. Means were compared by the Tukey's test at 5 percent significance. The obtained results suggest that pre-imbibition of seeds for 18h in water is the most suitable procedure to evaluate the germinative potential of Narrow-leaved Paperbark seeds. Under greenhouse conditions, this treatment can be used with the substrates sand or Plantmax®.