白及有效部位的吸收代谢机制及PK-PD研究

Bletilla striata is a well-known Chinese medicinal plant widely used by Guizhou local people for the treatment of hemorrhagic disease. As the genuine medicinal plant in Guizhou province , it is one of the traditional Chinese medicine industry chain to give priority to be cultivated and developed. Nowadays more than 20 preparations about Bletilla striata had been listed, which have huge development potential and predominance of regional characteristics. However, the pharmacodynamic substance base and the process in vivo of Bletilla striata is still not very clear. It has become the technical bottleneck for further exploitation and utilization. Based on early working foundation including chemical constituents, pharmacological action and the study of effective components of Bletilla striata, we will establish technological means to accomplish the study of fingerprint with multicomponent analysis and qualitative and quantitative analysis by UPLC-DAD-MS/MS and UPLC-QTOF/MS. Then we used absorption model of caco-2 cell and in situ intestine perfusion and metabolic model of intestinal flora, liver microsomes and zebra fish. It aims to study the dynamic variation of absorption, metabolism, transformation of main components in Bletilla striata. Meanwhile, PK-PD model of Bletilla striata effective substances will be builded. Pharmacokinetics characteristic of effective substances about Bletilla striata and the directly related material that have hemostatic effect in vivo also will be illuminated. It will provide scientific basis for new drug development and deep exploitation of medicine resources about Bletilla striata.

白及在贵州民间广泛用于治疗出血性疾病，为贵州地道药材和重点培育发展的中药材产业链之一，现有20多个白及相关制剂上市，具有较强地域特色资源优势和发展潜力。但目前白及药效物质基础及体内过程尚不清楚，已成为其深层次开发利用的技术瓶颈。课题组将在白及化学成分、药理作用、有效部位研究的前期工作基础上，以UPLC-DAD-MS/MS和UPLC-QTOF/MS建立基于紫外和质谱响应指纹图谱的多成分定性定量分析技术为基本手段，利用体内外吸收模型（Caco-2细胞、小肠灌流吸收模型）和代谢模型（肠道菌群、肝微粒体、斑马鱼代谢模型），探讨白及主要成分吸收、代谢、转化的动态变化规律，并建立PK-PD模型，通过药效学和体内成分经时过程的相关性分析，阐明白及的药代动力学特征及体内与止血作用直接相关的物质，为白及创新药物研制和药用资源深度开发奠定科学依据。

白及为兰科植物白及的干燥块茎，主产于贵州，为我省地道药材，现有20多个白及相关制剂上市，具有较强地域特色资源优势和发展潜力。本课题针对白及药效物质基础及体内过程尚不清楚的问题，应用在体肠灌流、离体肠外翻、Caco-2细胞模型，考察了白及提取物的可被吸收成分、吸收机制、吸收影响因素，提示不同模型的结果基本一致，白及有效部位中各成分的主要吸收部位在十二指肠，各成分吸收均受到胆汁的影响且均不是转运蛋白P-gp的底物；研究完成了白及有效部位在大鼠体内的代谢研究及白及代表性活性成分militarine在模式生物斑马鱼、大鼠及人肠道菌群、肝微粒体中的代谢特征，其代谢途径为以水解、脱糖、葡萄糖醛酸化、硫酸化为主的Ⅰ相和Ⅱ相反应；白及中丁二酸苄酯苷类化合物在体内的代谢转化关系主要通过糖苷键和葡萄糖氧苄基的断裂而实现的；研究获取了口服灌胃和静脉注射给药后白及多效应成分在大鼠体内的药代动力学特征和生物利用度；采用全身肝素化出血小鼠模型，利用WinNonLin软件建立了白及多效应成分的PK-PD模型，提示药效指标凝血酶时间（TT）、纤溶酶（PL）、血栓素B2（TXB2）及纤维蛋白原（Fbg）与白及有效部位中5种活性成分的浓度存在良好的相关性，说明白及可显著降低 TT并增加PL、TXB2和Fbg的含量，通过增强外源性凝血途径和共同凝血途径中的各凝血因子的活性，从而发挥止血作用。建立了基于白及药效物质的多指标质量控制新方法，研究分析了生长在贵州不同海拔高度的白及药材中有效成分含量变化规律，为白及药材的质量控制及道地性开发和栽培区域划分提供了参考。上述研究探讨了基于体内ADME过程的中药药效基础研究思路，并为白及创新药物研制及药材资源深度开发提供了科学依据。通过项目实施，发表科技论文11篇，申请发明专利2项，授权发明专利1项，以项目组为依托，经贵阳市科技局批准建设了“药代动力学研究科技创新人才团队”。