基于中红外光谱技术的短瓣金莲花抗氧化谱－物、谱－效关系及抗氧化水平快速分析方法的研究

The oxidation resistance of medicinal materials is one of the hot spots in the field of traditional Chinese medicine (TCM). Trollius ledebouri is selected as the study object in this research project. A direct and fast sampling technique, attenuated total reflection (ATR), is used to examine the mid-infrared (MIR) fingerprint spectra of different extracts of the flowers of Trollius ledebouri, and ultraviolet-visible spectrophotometry, high performance liquid chromatography, high performance liquid chromatography-mass spectrometry tandem technology are applied as well. Multiple linear regression and bivariate correlation are used to set up the relation between HPLC chromatographic fingerprint spectra and antioxidant activity, and the antioxidant constituents of the flowers of Trollius ledebouri will be confirmed by combining with HPLC-MS tandem technology and HPLC fast screening methods. On this basis, backward method is used to construct the equation which expressing the relationship between total flavonoids, antioxidant constituents, DPPH free radical scavenging ability, hydroxyl free radical scavenging ability, superoxide free radical scavenging ability and MIR fingerprint spectra, several pattern recognition methods are used to construct the MIR model of the several antioxidant indexes, as to discuss the interrelation in infrared fingerprint spectra-antioxidant constituents and infrared fingerprint spectra-antioxidant effectiveness and establish a fast analyzing method for the antioxidant level by infrared spectroscopy as well. Thus, a fast and effective method for the evaluation of the bioactivity and the analysis of bioactive substance of traditional Chinese medicine is provided, and new development direction in the field of TCM for the application of infrared spectroscopy is explored.

药材的抗氧化性是近年来中医药领域研究的热点之一。本课题以抗氧化性中蒙药材―短瓣金莲花（Trollius ledebouri）作为研究对象，利用直接、快速的衰减全反射采样技术获得MIR指纹图谱，结合紫外-可见分光光度法和化学计量学方法，在利用多元线性回归与双变量相关分析方法以及液-质联用技术和HPLC快速筛选方法确定短瓣金莲花抗氧化活性成分的基础上，利用向后消元法建立各抗氧化指标（总黄酮含量、各抗氧化活性成分含量、总抗氧化能力、DPPH自由基、羟基自由基和超氧阴离子自由基清除能力）与MIR谱图的相关方程，对短瓣金莲花的抗氧化谱-物、谱-效关系进行分析；并采用多种模式识别方法构建各抗氧化指标的MIR预测模型，以期建立利用中红外光谱技术对该药材抗氧化水平进行快速分析的方法。该课题为中药材质量评价的探讨及药效物质基础的分析提供了快速有效的方法，亦为红外光谱技术拓展了在中药现代化研究领域中的应用。

本课题利用直接、快速的衰减全反射MIR指纹图谱技术，结合紫外-可见分光光度法和化学计量学方法，对抗氧化性中蒙药材―短瓣金莲花（Trollius ledebouri）进行了系统的研究，具体包括：（1）利用 HPLC离线筛选技术筛选短瓣金莲花提取物的抗氧化活性色谱峰，并应用 UPLC-MS 方法对短瓣金莲花提取物的部分成分进行定性和定量分析，发现荭草苷、荭草素-2''-O-β-L-半乳糖苷、藜芦酸、牡荆苷、田蓟苷等成分均表现出不同程度的抗氧化活性；（2）以高效液相色谱法获得短瓣金莲花指标成分（荭草苷、牡荆苷）的含量作为参考数据，以傅里叶变换红外光谱技术获取其红外光谱图，利用化学计量学方法将指标成分与红外光谱图数据关联，构建了短瓣金莲花药材指标成分（荭草苷、牡荆苷）的快速预测模型。(3)对短瓣金莲花进行黄酮、酚酸、糖和木质纤维素含量的测定，并利用三级红外光谱技术对化学成分相似的短瓣金莲花不同部位进行鉴别和区分，实现了对短瓣金莲花质量的快速评价；（4）化学成分与抗氧化活性相关性分析表明，短瓣金莲花不同部位甲醇和超纯水提取物的黄酮含量和酚酸含量与 DPPH 自由基、羟基自由基、超氧阴离子自由基清除能力之间均成正相关关系，说明短瓣金莲花中的黄酮和酚酸对其抗氧化活性有贡献作用；（5）将不同溶剂体系的总黄酮含量与红外光谱建立合理模型，实现了利用数学模型对短瓣金莲花总黄酮含量进行预测。本课题对短瓣金莲花的抗氧化成分、抗氧化水平及抗氧化谱效关系进行了系统的研究，为短瓣金莲花抗氧化活性的物质基础分析及相关深入研究提供了有价值的数据支撑。.