Antiproliferative effects of pinostrobin and 5,6-dehydrokavain isolated from leaves of Alpinia zerumbet

Abstract Natural products are a major source of drugs for the treatment of cancer. The species Alpinia zerumbet (Pers.) B.L. Burtt & R.M. Sm, Zingiberaceae, is widely distributed in Brazil where it is known as "colônia". The leaves are commonly used in the treatment of hypertension and dyspepsia, however, the effects of A. zerumbet extracts and isolated substances on human cancer cells remain to be elucidated. This study was designed to identify the chemical constituents of hydroalcoholic and dichloromethane extracts from A. zerumbet leaves and to investigate their in vitro antiproliferative activity. The isolated phytochemicals included kaempferol, dihydro-5,6-dehydrokavain, 5,6-dehydrokavain, and pinostrobin. The hydroalcoholic extract inhibited cellular proliferation only at high concentrations, while the dichloromethane extract showed a moderate antiproliferative effect against leukemia and lung tumor cell lines. 5,6-Dehydrokavain showed potent cytostatic activity against glioblastoma cells and a moderate effect on all other tumor cell lines. Pinostrobin showed potent activity against leukemia and breast tumor cell lines and moderate cytostatic effect against ovarian cell. Furthermore, this is the first report on the isolation of kaempferol and pinostrobin from A. zerumbet leaves. Moreover, the purification process described in this study was effective. These results suggest that A. zerumbet leaves are a promising source of anticancer compounds.

Comparative study of Passiflora taxa leaves: II. A chromatographic profile

ABSTRACT Popularly known as passion fruit, some species of the genus Passiflora are widely used in folk medicines, such as sedatives and tranquilizers in many countries. Although these plants are employed for the same purposes, research with different species of Passiflora has indicated their heterogeneous chemical compositions. Since different chemical compositions can result in varying degrees of therapeutic efficiency, quality control based on the chemical constituents of each species is essential. To that end, the aim of this study was to compare pharmacognostically species of Passiflora in order to establish a chromatographic profile for the quality control of drugs in herbal medicines containing passion fruit. The study was conducted by collecting samples of leaves from twelve Passiflora taxa (i.e., ten species and two forms of P. edulis) – P. actinia, P. alata, P. amethystina, P. capsularis, P. cincinnata, P. edulis f. flavicarpa, P. edulis f. edulis, P. incarnata, P. morifolia, P. urnifolia, P. coccinea, and P. setacea – from different locations and obtaining their chromatographic profiles via thin-layer chromatography and high-performance liquid chromatography. Both methods used the flavonoid C-glycosides isoorientin, orientin, vitexin, and isovitexin as reference compounds and could ultimately establish specific profiles for each species. The chromatographic analyses discussed here can be used to assist in determining the quality and authenticity of herbal drugs derived from Passiflora species.

Reprint of: Liquid chromatographic enzymatic studies with on-line Beta-secretase immobilized enzyme reactor and 4-(4-dimethylaminophenylazo) benzoic acid/5-[(2-aminoethyl) amino] naphthalene-1-sulfonic acid peptide as fluorogenic substrate.

High throughput screening (HTS) techniques are required for the fast hit inhibitors selection in the early discovery process. However, in Beta-secretase (BACE1) inhibitors screening campaign, the most frequently used methoxycoumarin based peptide substrate (M-2420) is not widely applicable when aromatic or heterocycle compounds of natural source show auto-fluorescence interferences. Here, in order to overcome these drawbacks, we propose the use of a highly selective 4-(4-dimethylaminophenylazo)benzoic acid/5-[(2-aminoethyl)amino]naphthalene-1-sulfonic acid (DABCYL/1,5-EDANS) based peptide substrate (Substrate IV), whose cleavage product is devoid of spectroscopic interference. HrBACE1-IMER was prepared and characterized in terms of units of immobilised hrBACE1. BACE1 catalyzed Substrate IV cleavage was on-line kinetically characterized in terms of KM and vmax, in a classical Michaelis and Menten study. The on-line kinetic constants were found consistent with those obtained with the in solution fluorescence resonance energy transfer (FRET) standard method. In order to further validate the use of Substrate IV for inhibition studies, the inhibitory potency of the well-known BACE1 peptide InhibitorIV (IC50: 0.19 ± 0.02 µM) and of the natural compound Uleine (IC50: 0.57 ± 0.05) were determined in the optimized on-line hrBACE1-IMER. The IC50 values on the hrBACE1-IMER system were found in agreement with that obtained by the conventional methods confirming the applicability of Substrate IV for on-line BACE1 kinetic and inhibition studies.

Liquid chromatographic enzymatic studies with on-line Beta-secretase immobilized enzyme reactor and 4-(4-dimethylaminophenylazo) benzoic acid/5-[(2-aminoethyl) amino] naphthalene-1-sulfonic acid peptide as fluorogenic substrate.

High throughput screening (HTS) techniques are required for the fast hit inhibitors selection in the early discovery process. However, in Beta-secretase (BACE1) inhibitors screening campaign, the most frequently used methoxycoumarin based peptide substrate (M-2420) is not widely applicable when aromatic or heterocycle compounds of natural source show auto-fluorescence interferences. Here, in order to overcome these drawbacks, we propose the use of a highly selective 4-(4-dimethylaminophenylazo)benzoic acid/5-[(2-aminoethyl)amino]naphthalene-1-sulfonic acid (DABCYL/1,5-EDANS) based peptide substrate (Substrate IV), whose cleavage product is devoid of spectroscopic interference. HrBACE1-IMER was prepared and characterized in terms of units of immobilised hrBACE1. BACE1 catalyzed Substrate IV cleavage was on-line kinetically characterized in terms of KM and vmax, in a classical Michaelis and Menten study. The on-line kinetic constants were found consistent with those obtained with the in solution fluorescence resonance energy transfer (FRET) standard method. In order to further validate the use of Substrate IV for inhibition studies, the inhibitory potency of the well-known BACE1 peptide InhibitorIV (IC50: 0.19±0.02µM) and of the natural compound Uleine (IC50: 0.57±0.05) were determined in the optimized on-line hrBACE1-IMER. The IC50 values on the hrBACE1-IMER system were found in agreement with that obtained by the conventional methods confirming the applicability of Substrate IV for on-line BACE1 kinetic and inhibition studies.

Estudo comparativo do uso de extrato de Pfaffia glomerata e do laser de baixa potência (Hélio-Neônio) na cicatrização de feridas em ratos / Comparative study of the use of Pfaffia glomerata and the low power laser (Helium-Neon) in the healing of the wounds in rats

RACIONAL: A Pfaffia glomerata é planta utilizada na medicina popular como tônico, antidiabético, para melhorar o processo de cicatrização, neutralizar distúrbios gástricos e antirreumático. OBJETIVOS: Comparar a contração da ferida e a neovascularização com o uso da Pfaffia glomerata e do laser de baixa potência em dorso de ratos. MÉTODOS: Foram utilizados 40 ratos da linhagem Wistar nos quais realizaram-se feridas cirúrgicas com punch de 3 mm de diâmetro na parte superior direita do dorso onde nenhuma substância foi aplicada e nesses mesmos animais foram também realizadas feridas cirúrgicas no lado esquerdo na região inferior, onde foi aplicado o extrato das raízes de Pfaffia glomerata (Spreng.) Pedersen, Amaranthaceae, obtendo-se desta maneira os grupos controle e planta nos mesmos animais. Em outros 40 animais, foi utilizado o laser de baixa potência. Nas datas programadas em 48 horas, uma, duas e três semanas foram tomadas as medidas da contração da ferida. Microscopicamente os resultados foram analisados utilizando-se imunoistoquímica como fator VIII para observar a densidade vascular. RESULTADOS: Macroscopicamente não houve diferença estatisticamente significante com relação à contração da ferida entre os grupos planta e laser, obtendo ambos resultados superiores ao grupo controle. Dentro desta variável, o resultado com diferença estatisticamente significante ocorreu dentro do grupo laser na comparação dos subgrupos: 48 horas vs. uma semana (p=0,008). A análise do Fator VIII mostrou significância estatística no subgrupo de uma semana da planta em relação ao laser(p=0,09). CONCLUSÕES: Não houve diferença estatisticamente significante com relação à contração da ferida entre os grupos planta e laser, obtendo ambos resultados superiores ao grupo controle no final do estudo; contudo, microscopicamente, o grupo planta obteve valor superior ao grupo laser quanto à neovascularização em uma semana, mas após esse período, houve equilíbrio com os outros grupos.

BACKGROUND: Pfaffia glomerata is a plant used in folk medicine as tonic, antidiabetic, to improve the healing process, neutralize stomach upset and antirheumatic. AIM: To compare the wound contraction and the neovascularization with the use of plant extract of Pfaffia glomerata (Spreng.) Pedersen, Amaranthaceae, and low power laser in rats. METHODS: It was used 40 Wistar rats in which surgical wounds with a punch of 3 mm in diameter at the top right of the back and no substance was applied at this site, and similar wounds in the lower left side had applicated extract of P. glomerata (Spreng.) Pedersen, Amaranthaceae, control groups and plants in the same animal. Another 40 animals had the low power laser application. On dates scheduled within 48 hours, one, two and three weeks measures were taken and wound contraction observed. Microscopic results were analyzed using immunohistochemistry with Factor VIII to observe the vascular density. RESULTS: It was possible to observe macroscopically no statistic significant difference with respect to the contraction of the wound between the groups plant and laser, both getting better results than the control group. This variable showed a statistical significance with the laser group when was compared the subgroups 48 hours X one week (p=0,008). Statistical analysis of Factor VIII showed an statistic significance in subgroup one week of the plant with the laser (p=0.09). CONCLUSIONS: There was no statistic significant difference between the plant and laser groups with respect to contraction of the wound. On microscopic analysis, the group plant earned more than the laser group with relation to neovascularization in subgroup one week, and after that occured a balance between them.

Medicinal mushroom Ganoderma lucidum (Leyss: Fr) Karst. triggers immunomodulatory effects and reduces nitric oxide synthesis in mice.

This study investigated the effect of Ganoderma lucidum supplementation on lymphocytes and peritoneal macrophages from mice. Our results show that G. lucidum in vivo was able to increase interferon-gamma (IFN-gamma) concentration but reduced CD3(+) and CD8(+) spleen lymphocytes. Ex vivo, IFN-gamma; and interleukin-10 levels were increased and the tumor necrosis factor-alpha (TNF-alpha) level was reduced by peritoneal macrophages from mice fed with G. lucidum. In the absence of stimuli nitric oxide production was reduced in mice fed with G. lucidum, and with lipopolysaccharide stimulation nitric oxide production was increased but was lower than control values (P < .05). G. lucidum was grown by solid-state culture in wheat grain, and a chow containing 10% G. lucidum mycelium was formulated (G10). Swiss male mice were divided into two groups, termed G10 and control groups according to the diet, and were fed for 3 months. Peritoneal macrophages were obtained and investigated with regard to phagocytosis, lysosomal volume, hydrogen peroxide, superoxide anion, and cytokines ex vivo. In the plasma we investigated concentrations of cytokines, and in the spleen we determined subsets of CD3(+), CD4(+), CD8(+), and CD19(+) lymphocytes.