基于中西药相互作用的谷红注射液多组分药物代谢与药效相关性研究

The combined application of Chinese and Western medicines has now become widespread in clinic, but its study mainly focuses on efficacy observation and the effect of Chinese medicine on the pharmacokinetics of single western medical component, which is difficult to reflect the correlational feature of multicomponent drug metabolism in vivo and pharmacodynamics when Chinese and Western medicines are used together. This project chooses Guhong injection as the model prescription and combines acetylglutamine and three effective parts of Flos Carthami (flavonoid glycosides, yellow pigments and nucleosides) according to orthogonal test L16(45). On the basis of the fingerprint in vitro, effective ingredients (acetylglutamine, carthamin, hydroxysafflor yellow A and guanosine) or their metabolic compounds of sixteen different dosage ratio prescriptions of Guhong in cerebral ischemia animal after intravenous administration are detected dynamically by HPLC/DAD, LC/MSn or isotope labeling-PET. The potency indicators such as cerebral function, thrombosis and fibrinolysis, free radical oxidation, neurotransmitter release and neuronal apoptosis are assessed simultaneously. Based on the herb-drug interaction (HDI), the conjunctive models of pharmacokinetics and pharmacodynamics which characterise the multiple components of Chinese and Western medicines are built, and then analyzed by various mathematical analysis method. The interaction and action mechanism among multiple compounds of Guhong are interpreted from the aspects of pharmaceutics, pharmacokinetics and pharmacodynamics, which provides new research technology and method for basic research of the combined application of Chinese and Western medicines.

中西药联合用药现已成为临床普遍的用药方式，但其研究目前大多集中在疗效观察、中药对西药单一成分的药代学影响等层面，难以反映中西药联用多组分体内变化与药效变化的相关性特征。 本项目以谷红注射液为示范，正交组合乙酰谷酰胺和红花主要有效部位（黄酮苷类、红花黄色素类和核苷类）组成16个谷红组分配伍方。在构建体外指纹图谱基础上，HPLC/DAD、LC/MSn、同位素标记-PET检测各谷红配伍方静脉给药后在脑缺血动物体内有效成分（乙酰谷酰胺、红花苷、羟基红花黄色素A和鸟苷等）的含量或代谢变化；同步测定各谷红配伍方抗脑缺血损伤的药效学指标（脑功能、血栓与纤溶、自由基氧化、神经递质释放、细胞凋亡等）；基于中西药相互作用原理，引入多种数理分析方法，构建反映中西药多组分特征的药代与药效联合模型，从药剂学、药物代谢、药效学角度阐释谷红多组分相互作用原理和作用机制，为中西药联用的基础研究提供新的技术与方法。

谷红注射液是由乙酰谷酰胺和红花提取物混合制成的灭菌水溶液，为典型的中西药联用制剂。本项目基于中西药相互作用原理，从药剂学、药物代谢、药效学角度阐释了谷红注射液抗脑缺血损伤的药效活性以及组分间相互作用的原理和机制，不仅为谷红注射液的临床合理应用提供了客观的依据，也为中西药联用的基础研究提供了新的技术与方法。.在研究过程中，首先采用UV-VIS、HPLC-DAD 等建立了谷红注射液及其组方药物中有效部位和有效成分的体外含量测定方法，构建了相应的指纹图谱，并结合运用 HPLC-MSn 和血、脑微透析技术等对谷红注射液中化学组分在大鼠体内的代谢变化进行了检测；然后借助 ELISA、Western Blot、RT-PCR、流式细胞术、PET等探讨了谷红注射液抗脑缺血再灌注损伤的作用及作用机制；最后对乙酰谷酰胺和红花主要有效部位进行正交组合，引入数学模型解析谷红多组分体内代谢和抗脑缺血损伤药效学指标之间的相关性。.研究结果表明：谷红注射液中总多酚等有效部位和乙酰谷酰胺、尿苷、腺苷、鸟苷、紫丁香苷、羟基红花黄色素A、脱水红花黄色素B等有效成分的含量相对稳定，指纹图谱的相似度在 0.95 以上，谷红注射液的化学组成较为稳定。谷红注射液中代表性成分乙酰谷酰胺的体内代谢产物主要是乙酰谷氨酸、谷氨酰胺和谷氨酸；羟基红花黄色素A在体内有11种潜在的代谢产物，主要是其氧化、甲基化、或脱水后的衍生物。谷红注射液可有效减轻 MCAO 局灶性脑缺血模型大鼠的神经功能缺损症状，降低脑梗死体积，改善大鼠脑组织的病理异常，具有良好的抗缺血损伤作用，其作用机制与抗氧化、抗血栓、抗炎、抗细胞凋亡、调节氨基酸类神经递质的释放、改善葡萄糖和能量代谢紊乱、保护脑微血管内皮细胞和海马神经元等相关。谷红注射液中多个组分的体内代谢与抗脑缺血损伤药效学指标之间存在着显著的相关性，且其中乙酰谷酰胺、红花组分的生物利用度和抗脑缺血的药效作用均比单用时明显增强，其原因可能是两药的相互作用降低了对方成分与血浆蛋白的结合，使成分更多地被释放出来，从而促进了药效的发挥。