Department of Tropical Medicine, Casilla de Correo 2511, Instituto de Investigaciones en Ciencias de la Salud Asunción, Universidad Nacional de Asunción, Paraguay.
J Ethnopharmacol. 2011 Feb 16;133(3):986-93. doi: 10.1016/j.jep.2010.11.032. Epub 2010 Dec 4.
Zanthoxylum chiloperone var. angustifolium Engl. (Rutaceae) stem bark is used traditionally in Paraguay for its antiparasitic properties. Canthin-6-one is main compound isolated from Zanthoxylum chiloperone var angustifolium with broad spectrum antifungal, leishmanicidal and trypanocidal activities.
The qualitative and quantitative characterization and the isolation of main alkaloidal components of different organs of Zanthoxylum chiloperone are investigated by HPLC-UV-MS. The in vitro biological activity of each extract against trypomastigote and amastigote forms of Trypanosoma cruzi parasites were evaluated, then comparison the in vivo efficacy of the ethanolic leaves extract of Zanthoxylum chiloperone with reference drug, benznidazole, in acute Trypanosoma cruzi infected mice when administered by oral route. We have also evaluated the mutagenic and cytotoxic activity of the main component of Zanthoxylum chiloperone, i.e. canthin-6-one, by mouse bone marrow micronucleus test.
The compositions of the ethanol extracts obtained after the maceration process were studied by HPLC-UV-MS methods. The quantitation analysis was performed by external standard method, using a calibration curve constructed utilizing solutions containing different concentrations of the reference samples. The anti-trypomastigote activity was evaluated by the lysis effect on mouse blood trypomastigotes (Y strain Trypanosoma cruzi). The anti-amastigote Trypanosoma cruzi activity was evaluated by a modified colorimetric method with chlorophenol red-β-d-galactopyranoside (CPRG). The cytotoxicity of extracts and compounds was performed on NCTC 929 cells. The in vivo efficacy of the ethanolic leaves extract of Zanthoxylum chiloperone and benznidazole, in acute Trypanosoma cruzi (two different strains) was evaluated in Trypanosoma cruzi infected mice; the drugs were administered by oral route. The mortality rates were recorded and parasitaemias in control and treated mice were determined once weekly for 70 days. The mutagenic and cytotoxic activity of the main component of Zanthoxylum chiloperone, canthin-6-one, by mouse bone marrow micronucleus test.
Canthin-6-one was the main compound of stem and root bark and 5-methoxy-canthin-6-one in leaves and fruits. The ethanolic leaves extract, canthin-6-one and benznidazole presented, approximately, the same level of in vitro activity against trypomastigote and amastigote forms of Trypanosoma cruzi. We have also evaluated the mutagenic and cytotoxic effects of canthin-6-one by micronucleus test in mice. This test showed any mutagenic and cytotoxic damages. The effects of oral or subcutaneous treatments at 10 mg/kg daily for 2 weeks with the ethanolic extract of leaves of Zanthoxylum chiloperone were examined in Balb/c mice infected acutely with Trypanosoma cruzi (CL or Y strain) and compared with benznidazole at 50 mg/kg for 2 weeks. In these experiments, 70 days after infection, parasitaemia and serological response were significantly reduced with the oral ethanolic extract treatment compared with reference drug.
This study have shown the efficacy of the leaves extract of Zanthoxylum chiloperone in reducing Trypanosoma cruzi parasitaemia in vivo assays and could be welcomed by scientific and rural communities of Paraguay because it could help them towards the use of local resources to treat an endemic infection, Chagas disease, affecting 20% of the population of this country.
Zanthoxylum chiloperone var. angustifolium Engl.（芸香科）树皮在巴拉圭传统上用于驱虫。Canthin-6-one 是从 Zanthoxylum chiloperone var angustifolium 中分离出的主要化合物，具有广谱抗真菌、杀利什曼原虫和杀锥虫活性。
通过 HPLC-UV-MS 对 Zanthoxylum chiloperone 的不同器官的主要生物碱成分进行定性和定量特征描述和分离。评估各提取物对锥虫Trypanosoma cruzi 滋养体和包囊体形式的体外生物活性，然后比较 Zanthoxylum chiloperone 叶乙醇提取物与参考药物苯并咪唑在急性 Trypanosoma cruzi 感染小鼠中的体内疗效，通过口服途径给药。我们还通过小鼠骨髓微核试验评估了 Zanthoxylum chiloperone 的主要成分 Canthin-6-one 的致突变和细胞毒性活性。
通过 HPLC-UV-MS 方法研究了醇提物的组成。通过外标法进行定量分析，使用构建的校准曲线，其中包含不同浓度参考样品的溶液。抗锥虫滋养体活性通过对鼠血锥虫（Y 株 Trypanosoma cruzi）的裂解作用进行评估。抗锥虫滋养体 Trypanosoma cruzi 活性通过改良的比色法用氯苯酚红-β-d-半乳糖吡喃糖苷（CPRG）进行评估。在 NCTC 929 细胞上进行提取物和化合物的细胞毒性测定。在急性 Trypanosoma cruzi（两种不同株系）感染的小鼠中，通过口服途径评价 Zanthoxylum chiloperone 叶乙醇提取物和苯并咪唑的体内疗效；记录死亡率，并每周确定对照和治疗小鼠中的寄生虫血症 70 天。通过小鼠骨髓微核试验评估了 Zanthoxylum chiloperone 的主要成分 Canthin-6-one 的致突变和细胞毒性活性。
Canthin-6-one 是茎和根皮的主要化合物，5-甲氧基-canthin-6-one 是叶和果实的主要化合物。叶乙醇提取物、Canthin-6-one 和苯并咪唑对锥虫滋养体和包囊体形式的 Trypanosoma cruzi 具有大致相同的体外活性。我们还通过小鼠微核试验评估了 Canthin-6-one 的致突变和细胞毒性作用。该试验显示无致突变和细胞毒性损伤。在感染急性 Trypanosoma cruzi（CL 或 Y 株）的 Balb/c 小鼠中，每日口服或皮下给予 10mg/kg 叶乙醇提取物治疗 2 周，并与 50mg/kg 的苯并咪唑进行比较。在这些实验中，感染后 70 天，与参考药物相比，口服乙醇提取物治疗可显著降低锥虫血症和血清学反应。
这项研究表明，Zanthoxylum chiloperone 叶提取物在体内试验中降低锥虫寄生虫血症的疗效，并可能受到巴拉圭科学和农村社区的欢迎，因为它可以帮助他们利用当地资源治疗该国 20%人口的地方性感染——恰加斯病。
