Turbo-extraction of glycosides from Stevia rebaudiana using a fractional factorial design

ABSTRACT Stevia rebaudiana (Bertoni) Bertoni, Asteraceae, leaf extract has recently called the attention of food industry as a proposal for natural sweetener. The sweet flavor is attributed to the glycosides, in especial stevioside and rebaudioside A, which are the plant main chemical markers. The aim of the work reported here was to optimize the turbo-extraction of stevia leaves using water, ethanol 70% and 90% (w/w) as green solvents. A 25-2 factorial design was applied to study the linear effects of the drug size, solvent to drug ratio, temperature, time and also the turbolysis speed on the extraction of glycosides. The glycosides exhaustive extraction showed that ethanol 70% gave better results and was used for turbo-extraction. The stevioside and rebaudioside A contents were quantified by a validated method by high performance liquid chromatographic with photodiode array detector. The contents of stevioside and rebaudioside A in fluid extract increased with the drug size, but decreased at high shearing speeds and solvent to drug ratio, while their yields decreased at higher temperature and were not affected by turbo speed. An increase in solvent to drug ratio reduced significantly the glycosides percent in dried extract. Optimal solution for S. rebaudiana leaves turbo-extraction was determined by desirability functions. The optimal extraction condition corresponded to drug size of 780 µm, solvent to drug ratio of 10, extraction time of 18 min; temperature of 23 ºC and turbo speed of 20,000 rpm, resulting in yields of 4.98% and 2.70%, for stevioside and rebaudioside A, respectively. These yields are comparable to the ones recently published for dynamic maceration, but with the advantage of shorter extraction times. This work demonstrates that turbolysis is promising for S. rebaudiana glycosides extraction and stimulate new research on the purification of these extracts, which may become an interesting source of income for developing countries such as India and Brazil.

Effect of Poria cocos hydroethanolic extract on treating adriamycin-induced rat model of nephrotic syndrome / 中国结合医学杂志

<p><b>OBJECTIVE</b>To evaluate the effect of Poria cocos (Schw.) Wolf hydroethanolic extract (PHE) against nephrotic syndrome (NS) in rats and to identify the potential active components from PHE.</p><p><b>METHODS</b>The high content compounds were isolated and purified by using column chromatography followed by preparative highperformance liquid chromatography (p-HPLC). Forty male Wistar rats with adriamycin (ADR)-induced NS were randomly divided into 5 groups, 8 in each group: model control group, positive control group (with prednisone treatment), PHE low-dose group, PHE middle-dose group and PHE high-dose group. Another 8 rats were recruited as vehicle control group. All rats received the intragastric administration of corresponding drugs or saline for 30 days. During the experimental period, rats' behavior and appearance were observed and recorded daily, and their body weights were recorded weekly. After treatment, 24-h urine samples were collected to evaluate the urine protein and urine creatinine (Ucr); then the rats were sacrificed to collect carotid blood and to determine the levels of serum total protein (TP), albumin (Alb), globulin (Glo), total cholesterol (TC) and cytokine interlukin-4 (IL-4).</p><p><b>RESULTS</b>Six acidic components were isolated and identified from the PHE section: pachymic acid, 15α-hydroxydehydrotumulosic acid, trametenolic acid, dehydropachymic acid, 3β-hydroxy-lanosta-7,9(11), 24-trien-21-oic-acid and dehydroeburicoic acid. Compared with the model control group, the urine protein content were significantly decreased in the PHE treatment groups and positive control group (P<0.05), especially PHE middle-dose group (P<0.01). The Ucr values and serum levels of TP, Glo, TC and IL-4 in PHE low- and middle-dose groups were also presented obvious recover tendency as compared with the model control group (P<0.05 or P<0.01). However, positive control group and all PHE groups indicated no significant therapeutic effect on raising Alb value, although PHE low- and middle-dose treatment groups showed better outcomes than positive control group (P>0.05).</p><p><b>CONCLUSIONS</b>PHE showed an encouraging therapeutic effect against ADR-induced NS in a rat model. PHE might be a group of effective substances for the treatment of NS.</p>

Development of hydroethanolic extract of Ipomoea pes-caprae using factorial design followed by antinociceptive and anti-inflammatory evaluation

Ipomoea pes-caprae (L.) R. Br., Convolvulaceae, is a medicinal plant that grows abundantly as a pan-tropical stand plant. The 3² (two factors and three levels) factorial design, was applied to determine the best time and drug/solvent proportion to maximize the flavonoid content in the hydroethanolic extract by maceration process. The antinociceptive and anti-inflammatory effects were studied at 5-20 mg/kg, i.p., using the writhing test and carrageenan-induced pleurisy models in mice. The optimized extract was able to inhibit more than 50% of abdominal writhing at 20 mg/kg, with 55.88%±2.4 of maximum inhibition. Indomethacin, used as positive control, inhibited 64.86% at 10 mg/kg. In the pleurisy model, the extract produced dose-dependent inhibition of the first phase of inflammation (4 h) in the pleural cavity induced by injection of carrageenan (1%) in mice. It inhibited 50%±0.82 (p<0.01) of exudation induced by carrageenan, and 60.88%±0.14 (p<0.01) of leukocyte migration to the pleural cavity. In conclusion, the results validate the technological conditions of the maceration process to produce an optimized bioactive herb extract for the development of analgesic and anti-inflammatory phytopharmaceuticals using 70 ºGL ethanol, a plant to solvent ratio of 12.5% (w/v), and ten days of maceration.

Gingko biloba leaves extract on growth and morphology of trypanosomatids / Efecto del extracto de las hojas de Gingko biloba en el crecimiento y la morfología de tripanosomátidos

Gingko biloba has been one of the most used medicinal plants all over the world in the past years. In this study, our group has studied the effect of a hydroethanolic extract from the aerial parts of this plant on the growth and morphological differentiation of trypanosomatids. Herpetomonas samuelpessoai and Herpetomonas sp were used in this study. The extract was obtained in a Soxhlet apparatus (50 oC, 2 hours). This extract was aseptically added to Roitman’s medium in different concentrations (4, 20, 40, 60, 80 and 100 mg/ml). The growth rate was determined using a Newbauer chamber to count numbers of cells after the extract inoculation (24 and 72 hours later). Smears stained by the Panotic method was used to determine the percentages of pro, para and opisthomastigote forms. The extract inhibited Herpetomonas sp growth in concentrations higher than 20 mg/ml. H. samuelpessoai has been inhibited in doses higher than 40 mg/ml. No morphological differentiation was observed in Herpetomonas sp cell. However, morphological differentiations could be noticed in H. samuelpessoai cell using doses higher than 40 mg/ml. These alterations are probably related to the cell division process, since cells with 3 or 4 nucleus were observed. Also, cytoplasmatic expansions, representing unsuccessful process of cell division were frequently found out. Further ultrastructural analysis using a transmission electron microscope showed cells with homogeneous nucleus or the absence of it. Protozoan protein profile was also analyzed. It was possible to notice changes in both trypanosomatids used in this study. H. samuelpessoai has shown over expression and accumulation of proteins which its degradation is essential to continue the cell differentiation. Also, it is possible to suggest that this extract acts through the modulation of the genetic expression and may be harmful to human cells if not purified.

Gingko biloba es una de las plantas medicinales más utilizadas en todo el mundo en los últimos años. En este estudio, nuestro grupo ha estudiado el efecto de un extracto hidroetanólico de la parte aérea de esta planta sobre el crecimiento y la diferenciación morfológica de tripanosomátidos. Herpetomonas samuelpessoai y Herpetomonas sp se utilizaron en este estudio. El extracto se obtuvo en un aparato Soxhlet (50° C/2 horas). Este extracto se agregó asépticamente a medio Roitman en diferentes concentraciones (4, 20, 40, 60, 80 y 100 mg /ml). La tasa de crecimiento se determinó utilizando una cámara de Newbauer para contar el número de células después de la inoculación de extracto (24 y 72 horas más tarde). Frotis teñidos por el método Panotic se utilizó para determinar los porcentajes de pro, para y las formas opistomastigota. El extracto inhibió el crecimiento Herpetomonas sp en concentraciones superiores a 20 mg /ml. H. samuelpessoai se ha inhibido en dosis superiores a 40 mg /ml. No se observó diferenciación morfológica en la celda Herpetomonas sp. Sin embargo, las diferenciaciones morfológicas se pudo observar en la celda H. samuelpessoai con dosis superiores a 40 mg /ml. Estas alteraciones son probablemente relacionado con el proceso de división celular, ya que las células con 3 o 4 núcleos se observaron. Además, las expansiones citoplasmáticas, lo que representa el proceso fallido de la división celular se encontraron con frecuencia hacia fuera. Un análisis más detallado ultraestructural usando microscopio electrónico de transmisión mostró células con núcleo homogéneo o la ausencia de ella. El perfil de proteínas por Protozoarios también se ha analizado. Fue posible notar cambios tanto en tripanosomátidos utilizados en este estudio. H. samuelpessoai ha demostrado a lo largo de expresión y la acumulación de proteínas que su degradación es esencial para continuar con la diferenciación celular. Además, es posible sugerir que este extracto...

Antifungal activity of extracts from Brazilian Cerrado plants on Colletotrichum gloeosporioides and Corynespora cassiicola / Atividade antifúngica de extratos de plantas do Cerrado brasileiro sobre Colletotrichum gloeosporioides e Corynespora cassiicola

This study aimed to determine the antifungal activity of leaf aqueous and hydroethanolic extracts of 10 plants from the Brazilian Cerrado on Colletotrichum gloeosporioides and Corynespora cassiicola. Antifungal activity was measured through the incorporation of each extract in a culture media or spore suspension, at 50 percent concentration relative to the volume, determining respectively the mycelial growth and the spore germination. Then, the percentages of mycelial growth inhibition and spore germination inhibition were obtained based on the comparison with the control. The extracts had a variable action on the phytopathogens, from mycelial growth stimulation for Aristolochia esperanzae and Byrsonima verbascifolia extracts to complete inhibition of mycelial growth and spore germination for Myracrodruon urundeuva and Lafoensia pacari extracts. M. urundeuva, L. pacari and Caryocar brasiliense leaf extracts had antifungal activity against Colletotrichum gloeosporioides and Corynespora cassiicola; the hydroethanolic extracts presented more antifungal activity than the aqueous extracts, and spore germination of both phytopathogens was more affected than their mycelial growth.

O objetivo deste trabalho foi determinar a atividade antifúngica de extratos aquosos e extratos hidroetanólicos de folhas de 10 plantas do Cerrado brasileiro sobre Colletotrichum gloeosporioides e Corynespora cassiicola. A determinação da atividade antifúngica foi realizada pela incorporação do extrato em meio de cultura ou na suspensão de esporos, na concentração de 50 por cento em relação ao volume, determinando-se, respectivamente, o crescimento micelial e a germinação de esporos. Em seguida, pela comparação com a testemunha, foram obtidas as percentagens de inibição do crescimento micelial e da germinação dos esporos. Foi constatado comportamento variável dos extratos sobre os fitopatógenos, desde o estímulo no crescimento micelial para os extratos de Aristolochia esperanzae e Byrsonima verbascifolia, até a inibição completa do crescimento micelial e dagerminação dos esporos para os extratos de Myracrodruon urundeuva e Lafoensia pacari. Extratos de folhas de L. pacari, de M. urundeuva e de Caryocar brasiliense apresentaram atividade antifúngica sobre Colletotrichum gloeosporioides e Corynespora cassiicola; os extratos hidroetanólicos proporcionaram mais atividade antifúngica que os extratos aquosos, e a germinação de esporos de ambos os fitopatógenos foi mais afetada que o crescimento micelial.