荨麻属植物中5-α还原酶抑制剂-裂木脂素类成分的定向快速发现与优化

It is a hotspot to discover non steroidal 5α-reductase inhibitors as new agents for the treatment of benign prostatic hyperplasia, alopecia and acne. During our previous studies, extracts of Urtica species were screened and were found to exhibit strong inhibitory activities on 5α-reductase. Fifteen secolignans were isolated from U. fissa E. Pritz, U. triangularis Hand.-Mazz. and U. mairei Levl. and all of them were assayed to show potent inhibitory effects on 5α-reductase. .On the basis of the studies mentioned above, further research will be conducted in 3 aspects as follows: (1) the MS and NMR spectra of the isolated secolignans will be studied and the regular MS and NMR pattern will be concluded. UPLC/Q-TOF MS and NMR will be used to trace the secolignans guided by this pattern in other Urtica extracts and fractions. The secolignans will be isolated by means of modern chromatography and their structures will be determined by spectral analyses. （2）The content of secolignans will be detected by HPLC-UV-MS and NMR in different Urtica species and the dynamic variation pattern of secolignans during growing process of Urtica sepcies will also be studied. (3) The inhibitory activities of these secolignans and their modifed derivatives will be assayed and the 3D-SAR will be concluded by comparative molecular field analysis. The secolignans will be optimized according the 3D-QSAR and the activity of the optimized product will be assayed in vitro and in vivo..This project not only provides new ideas, methods and theoretical evidences for the rapid detection of active lead compounds from Urtica species for the full and rational utilization of Urtica resources, but also establishes the basis for the further research and development of secolignans as non-steroidal 5-α reductase inhibitors.

前期研究工作表明6种荨麻属植物根对5-α还原酶具有较强的抑制活性，通过对3种荨麻根中活性成分进行追踪，获得了15个具有很强活性的裂木脂素类成分。.在此基础上，本项目拟进一步开展以下三方面的研究：(1)分析已鉴定的裂木脂素类的MS、NMR规律，结合此规律，并利用UPLC/Q-TOF MS及NMR技术，快速追踪其它荨麻属植物中该类成分，采用现代色谱、波谱技术分离鉴定该属其它植物中的裂木脂素类；（2）采用LC-UV-MS技术测定该类成分在不同产地各种荨麻中的含量及该类成分随季节的动态变化规律；(3)测定该类成分的5-α还原酶抑制活性，采用比较分子力场分析法探讨三维定量构效关系，据此进行结构优化并对优化结构的体内外活性进行测定。.本项目为充分利用荨麻植物资源，从中快速发现活性先导化合物并对其优化提供新的思路、方法和理论依据，为裂木脂素类非甾体5-ARI先导物的研究开发奠定基础。

本项目在四川、贵州、云南、吉林、新疆等地采集了9种荨麻属植物，1种艾麻属植物、4种草胡椒属植物和5种卷柏属植物。分别对其进行了乙醇提取、溶剂萃取和大孔吸附树脂吸附得到总提物和各组分。在总结文献中裂木脂素类成分质谱裂解规律的基础上采用HPLC-Q/TOF-MS对上述总提物和各组分中该类成分进行了追踪。进一步对目标组分采用硅胶、ODS、Sephadex LH20柱色谱法粗分。再对粗分后组分中裂木脂素类成分进行HPLC-Q/TOF-MS追踪，然后对目标组分采用硅胶、ODS、Sephadex LH20柱色谱、HPLC等法纯化，采用NMR、HR-MS、CD等波谱学方法鉴定结构。共分离得到133种化合物，其中裂木脂素类41种，新化合物35个，50余种为首次从所在属中发现，80余种为首次从所在种中发现。. 采用HPLC-MS 技术测定了3个产地、3个种的荨麻属植物中裂木脂素类成分的含量，通过含量的比较发现四川马尔康产的三角叶荨麻、裂叶荨麻和四川泸州产的滇藏荨麻中裂木脂素含量较高。对6-11月份每个月中旬采集的荨麻根中裂木脂素含量进行了测定，发现9月份含量最高，确定9月份最佳采收期。. 以3-CMF为模板利用同源模建构建了5α-还原酶（5-AR）的三维结构、并与裂木质素类抑制剂进行对接，找到5-AR模拟蛋白与5-ARI的潜在结合位点。并从41种裂木质素类化合物中筛选出最佳发挥药理作用的化合物。. 综上，通过本项目研究为充分合理利用我国荨麻属、艾麻属、草胡椒属、卷柏属植物资源，从中快速发现活性先导化合物提供新的思路、方法和理论依据，为确定最佳品种和最佳采收期提供科学依据，为裂木脂素类作为5α-还原酶抑制作用先导化合物的研究开发奠定基础。