Toxicity evaluation of water extract of tissue-cultured Taraxacum formosanum by acute, subacute administration, and Ames test

BACKGROUND: Taraxacum species (commonly known as dandelion) used as herbal medicine have been reported to exhibit an antiproliferative effect on hepatoma cells and antitumor activity in non-small-cell lung cancer cells. Although several investigations have demonstrated the safety of Taraxacum officinale, the safety of tissue-cultured plants of T. formosanum has not been assessed so far. Therefore, the present study examines the safety of the water extract of the entire plant of tissue cultured T. formosanum based on acute and subacute toxicity tests in rats, as well as the Ames tests. RESULTS: No death or toxicity symptoms were observed in the acute and subacute tests. The results of the acute test revealed that the LD50 (50% of lethal dose) value of the T. formosanum water extract for rats exceeded 5 g/kg bw. No abnormal changes in the body weight, weekly food consumption, organ weight, or hematological, biochemical, and morphological parameters were observed in the subacute toxicity test. Thus, the no observed adverse effect level (NOAEL) of T. formosanum water extract was estimated to be higher than 2.0 g/kg. Finally, the results of the Ames test revealed that T. formosanum water extract was not genotoxic at any tested concentration to any of five Salmonella strains. CONCLUSIONS: The water extract of tissue-cultured T. formosanum was non-toxic to rats in acute and subacute tests and exhibited no genotoxicity to five Salmonella strains.

Concentraciones de alcaloides, glucósidos cianogénicos, polifenoles y saponinas en plantas medicinales seleccionadas en Ecuador y su relación con la toxicidad aguda contra Artemia salina / Concentrations of alkaloids, cyanogenic glycosides, polyphenols and saponins in selected medicinal plants from Ecuador and their relationship with acute toxicity against Artemia salina

ResumenEntre los principales compuestos químicos sintetizados por las plantas, pero considerados no esenciales para su metabolismo básico, están los alcaloides, los polifenoles, los glucósidos cianogénicos y las saponinas que tienen diversas funciones en las plantas y reconocidas propiedades medicinales y farmacológicas. En esta investigación se determinaron las concentraciones de los mencionados metabolitos secundarios en los extractos de las hojas de las plantas medicinales Taraxacum officinale, Parthenium hysterophorus, Artemisia absinthium, Cnidoscolus aconitifolius y Piper carpunya y se relacionaron con la toxicidad aguda contra Artemia salina. En cada bioensayo con A. salina se usaron los extractos alcohólicos de las hojas de las plantas seleccionadas a diferentes concentraciones, calculándose la proporción de organismos muertos y los CL50. Las concentraciones de alcaloides, fenoles totales, taninos, glucósidos cianogénicos y saponinas fueron determinadas mediante métodos espectrofotométricos. Este es el primer reporte de cuantificación de metabolitos secundarios en las plantas analizadas provenientes de Ecuador. T. officinale presentó las mayores concentraciones de fenoles (22.30 ± 0.23 mg/g) y taninos (11.70 ± 0.10 mg/g), C aconitifolius de glucósidos cianogénicos (5.02 ± 0.37 µg/g) y P. hysterophorus de saponinas (6.12 ± 0.02 mg/g). Las plantas evaluadas presentaron actividades hemolíticas dependiendo de las concentraciones de saponinas. Los valores de taninos determinados estuvieron entre 0.20 ± 0.01 y 11.70 ± 0.10 mg/g, por lo que no son adversos para su consumo. Aunque los valores de glucósidos cianogénicos son permisibles, es necesario monitorear la presencia de estos compuestos químicos en las plantas para minimizar problemas de salud. Los CL50 obtenidos oscilaron entre los valores 3.37 µg/mL, extremadamente letal o tóxica, para P. carpunya y 274.34 µg/mL, altamente tóxica, para T. officinale. De los análisis de correlaciones realizados a los resultados, se observó que los alcaloides favorecen de manera significativa (p<0.001) a la toxicidad aguda contra A. salina, mientras que a mayor contenido de polifenoles dicha toxicidad disminuye significativamente (p<0.001) el nivel de toxicidad de las plantas. Del análisis de componentes principales, se demuestra que las saponinas están en sinergia con los polifenoles para disminuir la toxicidad, pero tienen un efecto antagónico con los alcaloides y los glucósidos cianogénicos, lo cual evidencia que estos metabolitos secundarios presentan variabilidades en los mecanismos de acción contra A. salina, como compuestos citotóxicos. Estos resultados demuestran que las saponinas y los polifenoles pueden ser letales para A. salina a bajas concentraciones, evidenciando que este bioensayo permite evaluar extractos vegetales que contengan bajas concentraciones de compuestos con altas polaridades. La correspondencia significativamente positiva entre citoxicidad y concentración de los alcaloides, confirmada con el bioensayo de Artemia salina, puede ser útil para hallar fuentes promisorias de compuestos antitumorales y para evaluar los límites tolerables que no afecten otras células benignas. El contenido de metabolitos secundarios hallados en las plantas analizadas les atribuye un gran valor farmacológico.

Abstract:Alkaloids, polyphenols, cyanogenic glycosides and saponins are among the main chemical compounds synthesized by plants but not considered essential for their basic metabolism. These compounds have different functions in plants, and have been recognized with medicinal and pharmacological properties. In this research, concentrations of the mentioned secondary metabolites were determined in the medicinal plants Artemisia absinthium, Cnidoscolus aconitifolius, Parthenium hysterophorus, Piper carpunya and Taraxacum officinale, from Ecuador, and related with cytotoxic effects against Artemia salina. Alcoholic and aqueous extracts from leaves of these selected plants were prepared at different concentrations. To assess cytotoxicity of these extracts, different bioassays with A. salina were undertaken, and the mortality rates and LC50 were obtained. Besides, concentrations of alkaloids, cyanogenic glycosides, phenols, tannins and saponins were determined by spectrophotometric methods; this constituted the first report of quantification of secondary metabolites in the selected plants from Ecuador. T. officinale had the highest concentration of total phenols (22.30 ± 0.23 mg/g) and tannins (11.70 ± 0.10 mg/g), C. aconitifolius of cyanogenic glycosides (5.02 ± 0.37 µg/g) and P. hysterophorus of saponins (6.12 ± 0.02 mg/g). Tannins values obtained were not adverse to their consumption. Alcoholic and aqueous extracts of selected plants had hemolytic activity depending on the concentration of saponins. Although the values of cyanogenic glycosides were permissible, it was necessary to monitor the presence of this metabolite in plants to minimize health problems. LC50 values ranged from extremely toxic (3.37 µg/mL) to highly toxic (274.34 μg/mL), in P. carpunya and T. officinale, respectively. From correlation analysis, it was observed that increase values of alkaloids concentrations had highly significant (p<0.001) acute toxicity against A. salina, while at a higher polyphenol concentration the level of plants cytotoxicity decreased significantly (p<0.001). The results of principal component analysis showed that saponins apparently were in synergy with polyphenols to decrease cytotoxicity, but antagonize with alkaloids and cyanogenic glycosides, indicating that these secondary metabolites present variability in the mechanisms of action against A. salina, as cytotoxic compounds. These results also demonstrate that polyphenols and saponins can be lethal at low concentrations, demonstrating the potential of brine shrimp bioassay as a model to evaluate plant extracts containing low concentrations of chemical compounds with high polarities. The significant positive correlation between cytotoxicity and concentration of alkaloids confirmed by the bioassay of brine shrimp can be useful to identify promising sources of antitumor compounds, and to evaluate tolerable limits not affecting other benign cells. Contents of secondary metabolites found in the selected plants confer them great pharmacologic values. Rev. Biol. Trop. 64 (3): 1171-1184. Epub 2016 September 01.

Actividad antiviral contra el virus de la fiebre amarilla, cepa vacunal 17D, de extractos de hojas de Taraxacum officinale GH Weber ex Wiggers / Phytochemicaland antiviral activity against yellow fever virus, 17D vaccine strain, of extracts from leaves of Taraxacum officinale GH Weber ex Wiggers

Taraxacum officinale leaves were collected at two and 5 months of growth, for antiviral activity against flavivirus, using the 17D vaccine strain of yellow fever virus as a model. Using spectroscopy technique, a total of twelve (12) compounds were identified in the chloroform (C) and hexane (H) extracts of two and five months (2M and 5M) of recollection., The antiviral activity against the yellow fever 17D virus was evaluated with the plaque assay and the concentrations used (50 - 1,5 ug/mL) were no cytotoxic to Vero cells as determined using the MTT(3-(4,5-Dimetiltiazol-2yl)-2,4-difenilbromuro de tetrazolium) assay. The phytochemical composition of leaves growing for 5 months is different and more complex than leaves growing for 2 months. From the four extracts, only C5M inhibited the viral replication in a dose depend manner, with 100 percent viral inhibition at 50 ug/mL (p=0,0124) and the effective doses 50 (ED50: 10,2 +/- 8,7 ug/mL), meanwhile, ED50 of C2M extract was 93,5 +/- 23,5 ug/mL, thus, the extract C5M is 8 times more effective than extract C2M. The identified compounds in extract C5M were: Psi taraxasteryl acetate, cafeic acid, taraxasteryl acetate, taraxerol, taraxerilo acetate and Psi-taraxasterol. One of these compounds or the combinations of them is responsible for the reported high antiviral activity.

Las hojas de Taraxacum officinale fueron colectadas a dos y cinco meses de crecimiento, para determinar actividad antiviral contraflavivirus, utilizando como modelo el virus de fiebre amarilla cepa vacunal 17D. Se identificaron por métodos espectroscópicos, un total de doce (12) compuestos provenientes de los extractos de hexano (H) y cloroformo (C) a dos y cinco meses (2M y 5M) de recolección La actividad antiviral se determinó mediante un ensayo de placa y las concentraciones de extractos utilizadas (50-1,5 ug/mL) fueron no citotóxica en células Vero, determinadas por el método colorimétrico del MTT (3-(4,5-Dimetiltiazol-2yl)-2,4-difenilbromuro de tetrazolio). La composición fitoquímica de los extractos de 5 meses es distinta y más compleja que la de dos meses de crecimiento. De los cuatro extractos sólo el C5M inhibió la replicación del virus en una manera dosis dependiente, con una inhibición del 100 por ciento a 50 ug/mL (p=0,0124) y una dosis efectiva 50 (DE50) de 10,2 +/- 8,7 ug/mL, mientras que el DE50 del extracto C2M es de 93,5 +/- 23,5 ug/mL, lo que hace al extracto clorofórmico de 5 meses aproximadamente 8 veces más efectivo que el C2M. Los compuestos presentes en el extracto C5M son Psi taraxasterilo, ácido cafeíco, acetato de taraxasterilo, taraxerol, acetato de taraxerilo y Psi-taraxasterol. Uno o más de estos compuestos son responsables de alta actividad antiviral reportada.