多基源藏药圆柏的清热药效物质基础与品质评价

Yuanbai is a traditonal Tibetan medicine with multiple origins. Both the leaves with twigs and the fruits of no less than ten Sabina plants are used as Yuanbai on Tibet Plateau. Yuanbai can clear heat of lung, kidney, liver and gallbladder, and is used for the treatment of pneumonia, arthritis, cholecystitis and nephritis. It is worth research and development on distinctive drugs to treat severe pneumonia and rheumatoid arthritis, which are more popular on plateau. However, the medicinal material foundation for clearing heat of Yuanbai have not been clarified. It is almost impossible to evaluate the effect or quality of the multiple origins of Yuanbai at present. On the basis of our previous research on heat-clearing Chinese medicine and the Tibetan medicinal theory of Yuanbai, we put forward and proved preliminarily the deduction that the inhibition on the excessive activation of complement system would be an important mechanism of Yuanbai’s heat-clearing effect. .In order to clarify the medicinal material foundation for clearing heat and the authentic medicinal resources with high quality of Yuanbai, the anticomplement components of the leaves with twigs of Sabina pingii var. wilsonii will be studied firstly, guided by anticomplement test in vitro. The relationships of structures and activities of the anticomplement compounds will be discussced. And the anticomplement activities in vitro of the important components will be comfirmed by their therapeutic effects on acute lung injury in vivo. Based on these results, the anticomplement components of the leaves with twigs and fruits of the other original plants of Yuanbai will be fast recognized on line by UPLC-MSn. The main and specific anticomplement compounds will be directionally prepared by HPLC and HSCCC. Not only the different species but also the different parts of the Sabina plants will be compared and evaluated on their activities and active components. This research will greatly improve the reasonable exploitation and utilization of Yuanbai and its compound medicines.

藏药圆柏，来源于青藏高原柏科圆柏属10多种植物的枝叶和果实，能清肺、肝胆、肾热，治肺炎、关节炎、胆囊炎、肾炎等，有望开发成为防治高原多发重症肺炎、类风湿性关节炎的特色藏药。但药效物质基础不明，用药混乱，有效性和稳定性难以保证。申请人结合前期清热类中药的研究成果及圆柏藏药理论，提出“抑制补体过度激活是圆柏发挥清热药效的重要途径之一”，并经预实验初步验证。为阐明圆柏复杂基源的清热药效物质基础，本项目以活性明确的香柏枝叶为切入点，活性导向分离抗补体活性成分，探讨构效关系，以补体介导的急性肺损伤模型验证活性组分的药效。在此基础上，采用UPLC-MSn等技术在线识别多品种、不同部位圆柏中的抗补体活性成分，采用HPLC、HSCCC等快速定向制备各基源的活性指标成分和特征性鉴别成分。最后以活性与成分双重指标评价不同基源的品质，理清藏药圆柏的正品优质药源。研究将大大促进圆柏及其制剂的质量控制与开发利用。

藏药圆柏，又称“徐巴”，来源于青藏高原柏科刺柏属（Juniperus）多种植物的枝叶，能清肺、肝胆、肾热， 治肺炎、关节炎、胆囊炎、肾炎等，有望开发成为防治高原多发重症肺炎、类风湿性关节炎的特色藏药。但药效物质基础不明，基源复杂难辨，有效性和稳定性难以保证。为理清藏药圆柏的正品优质药源，阐明与其清热药效相关的质量标志物，本项目首先采用叶绿体DNA组合序列建立了圆柏8个主要基源种的系统发育树，可用于圆柏的准确鉴定；进而结合前期清热类中药的研究成果及圆柏藏药理论，提出“抑制补体过度激活是圆柏发挥清热药效的重要途径之一”的假说，以抗补体活性为导向分离表征香柏等圆柏主要基源的清热药效物质，阐明构效关系，并以补体介导的流感病毒感染的急性肺损伤模型验证活性组分的药效；在此基础上，采用UPLC-MSn等技术在线识别圆柏多基源的抗补体活性成分和特征性鉴别成分，以活性与成分双重指标评价不同基源的品质。结果表明，不同刺柏属植物枝叶化合物类别相似，但组成与含量不同，导致活性差异，其中黄酮、木脂素、多糖为主要抗补体活性成分，鬼臼毒素类木脂素和多糖有细胞毒活性，挥发油有体外抗炎活性，二萜类成分无明显活性；圆柏的抗补体活性多糖对病毒感染急性肺损伤有明显疗效，但不同基源的体内抗炎药效差异明显，且与体外抗补体活性差异一致，证明抗补体活性可作为评价圆柏品质的可靠指标之一；对于抗补体活性，不同基源种间差异明显大于种内居群间差异，香柏、高山柏、小果垂枝柏、方枝柏和滇藏方枝柏是圆柏的优质基源；LCMS分析识别出不同种的特征性成分以及藏药圆柏的活性指标成分，基于HILIC色谱和FACE电泳的糖谱分析找到与圆柏多糖抗补体活性密切相关的四糖片段。本研究为圆柏等传统清热类藏药的药效物质基础与质量控制研究搭建了良好的技术平台，也为西藏刺柏属植物资源在新型抗病毒性肺炎新药的开发利用方面提供了科学依据。