Instituto de Pesquisas de Produtos Naturais, Universidade Federal do Rio de Janeiro - UFRJ, 21941-902 Rio de Janeiro, RJ, Brazil.
Laboratório de Biologia do Reconhecer, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense - UENF, 28013-602, Campos dos Goytacazes, RJ, Brazil; Laboratório de Produtos Bioativos, Curso de Farmácia, Universidade Federal do Rio de Janeiro - UFRJ, 27933-378 Macaé, RJ, Brazil.
J Ethnopharmacol. 2018 Jul 15;221:77-85. doi: 10.1016/j.jep.2018.04.007. Epub 2018 Apr 5.
Vochysia divergens Pohl (Vochysiaceae), popularly known as "Cambará", is a tree that is resistant to the seasonal floods in the Pantanal, and usually found in monodominant stands called "Cambarazal". The inhabitants of the Pantanal exploit this tree for medicinal uses. Infusions and decoctions of its leaves are taken as teas, particularly for the treatment of asthma, flu and diarrhea, according to the local tradition transmitted empirically through the generations.
To evaluate the beneficial health effects related to the ethnomedicinal uses of V. divergens (Vd) by using biomonitored fractionation of an aqueous leaf extract.
The aqueous leaf extract was obtained by decoction, and then the extract was fractionated by a combination of separation techniques including precipitation, organic partition and chromatography. Chromatographic analyses of the active samples were carried out using HPLC-DAD-MS. Flavonoid 1 was isolated from the n-BuOH fraction through classic chromatographic techniques. The inhibitory effects and cytotoxicity of the Vd extract, fractions and flavonoid 1 on NO and TNF-α production were assessed in LPS-stimulated RAW 264.7 macrophage cultures. Additionally, suppression on the proliferation of BALB/c lymphocytes was estimated by [H] thymidine incorporation. The antioxidant activity of the samples was verified by SNP and DPPH assays and the suppression of the iNOS protein expression was evaluated through Western blotting.
The HPLC-DAD-MS analysis of the Vd extract led to the identification of 5-methoxyluteolin-7-O-β-glucopyranoside (2), rutin (4) and the tannin galloyl-HHDP-glucopyranoside (3), besides the main flavonoid 3',5-dimethoxyluteolin-7-O-β-glucopyranoside (1), which was biologically evaluated in comparison with luteolin aglycone. The Vd extract, n-BuOH fraction and flavonoid 1 inhibited NO and TNF-α production by LPS-stimulated macrophages. The reduction of NO levels was mediated mainly by suppression of the iNOS expression. In addition, both the Vd extract (IC 13.6 µg/mL) and flavonoid 1 (IC 19.8 µg/mL; 41.6 µM) strongly inhibited stimulated lymphocyte proliferation when compared to the immunosuppressive agent cyclosporin A (IC 43.8 µg/mL; 36.4 µM). The Vd extract also showed a scavenging activity toward DPPH and NO free radicals. This is the first report describing the immunomodulatory potential of V. divergens and its major flavonoid (1).
Our findings showed that the aqueous leaf extract of V. divergens and its flavonoid reduced the production of excessive pro-inflammatory markers, collaborating with the Pantanal folk medicinal tradition that recommends the tea of cambará leaves for both asthma and flu. In addition, this study contributes to the knowledge of the pharmacological properties of 5-methoxy flavones, a poorly investigated subclass of flavonoids.
Vochysia divergens Pohl（卫矛科），俗称“Cambará”，是一种能抵抗潘塔纳尔季节性洪水的树木，通常生长在称为“Cambarazal”的单优势种群中。潘塔纳尔的居民利用这种树来治疗疾病。根据当地传统，人们会将其叶子煮沸或浸泡成茶，尤其用于治疗哮喘、流感和腹泻，这些经验是通过几代人的实践传承下来的。
通过对水提叶提取物进行生物监测分离，评估与 Vochysia divergens（Vd）的民族医学用途相关的有益健康影响。
通过煮沸获得水提叶提取物，然后通过包括沉淀、有机分配和色谱在内的分离技术组合对提取物进行分段。使用 HPLC-DAD-MS 对活性样品进行色谱分析。通过经典色谱技术从正丁醇（n-BuOH）部分中分离出黄酮 1。在 LPS 刺激的 RAW 264.7 巨噬细胞培养物中评估 Vd 提取物、馏分和黄酮 1 对 NO 和 TNF-α 产生的抑制作用和细胞毒性。通过[H]胸苷掺入来估计 BALB/c 淋巴细胞增殖的抑制作用。通过 SNP 和 DPPH 测定法验证样品的抗氧化活性，并通过 Western blot 评估 iNOS 蛋白表达的抑制作用。
通过 HPLC-DAD-MS 分析 Vd 提取物，鉴定出 5-甲氧基木犀草素-7-O-β-葡萄糖苷（2）、芦丁（4）和单宁没食子酰基 HHDP-葡萄糖苷（3），此外还鉴定出主要的黄酮 3',5-二甲氧基木犀草素-7-O-β-葡萄糖苷（1），与木犀草素糖苷相比，该化合物在生物评估中具有活性。Vd 提取物、正丁醇（n-BuOH）馏分和黄酮 1 抑制 LPS 刺激的巨噬细胞中 NO 和 TNF-α 的产生。NO 水平的降低主要是通过抑制 iNOS 表达来介导的。此外，与免疫抑制剂环孢素 A（IC 43.8 µg/mL；36.4 µM）相比，Vd 提取物（IC 13.6 µg/mL）和黄酮 1（IC 19.8 µg/mL；41.6 µM）均强烈抑制受刺激的淋巴细胞增殖。Vd 提取物还显示出对 DPPH 和 NO 自由基的清除活性。这是首次描述 Vochysia divergens 的免疫调节潜力及其主要黄酮（1）。
我们的研究结果表明，Vochysia divergens 的水提叶提取物及其类黄酮降低了过量促炎标志物的产生，这与潘塔纳尔民间医学传统相吻合，该传统建议饮用 cambará 叶茶来治疗哮喘和流感。此外，这项研究为 5-甲氧基黄酮这一研究较少的黄酮类化合物的药理学特性提供了更多的认识。
