激光拉曼光谱分析中药复方制剂中青蒿素含量的研究

从中药材黄花蒿中提取的青蒿素，是一种含有过氧基团的倍半萜内酯药物，目前已成为世界卫生组织推荐的抗疟疾药物。含有青蒿素的复方制剂具有抗疟、抑菌和调节免疫功能等作用，然而含青蒿素的复方制剂没有可量化的统一质量标准，因此难以控制和评价这些复方制剂的质量。目前对于青蒿素复方制剂的研究主要集中在定性分析其有效成分上，但具体如何定量分析其中青蒿素含量的研究工作却并不多，迫切需要一种简便、快速、无损的方法监控青蒿素的生产和使用。激光拉曼光谱是一种分子联合的光散射现象，能够提供样品分子的成分及结构，对研究青蒿素具有重要的意义。首先对青蒿素标准样品进行激光拉曼光谱定性分析测试，获得青蒿素标准品的拉曼特征图谱，这些标准图谱为以后分析中药复方制剂中的青蒿素奠定了基础。通过拉曼测试分析得到位于724 cm-1处的拉曼峰是与青蒿素中过氧基团直接相关的特性声子振动模式，可用于检测过氧桥键的存在，也是决定其抗疟活性的关键；而位于1 736 cm-1处的拉曼峰强度大、周围无其他振动峰干扰，是与青蒿素中内酯基团相关的特性声子振动模式，也可用来检测分析青蒿素。因此，拟采用青蒿素分子中位于724 cm-1处和1 736 cm-1两处的拉曼特征峰进行定性定量分析。然后，在实验室制备了一系列具有不同青蒿素质量百分数的青蒿素/面粉混合样品，确定出混合样品中青蒿素724 cm-1处和1 736 cm-1处拉曼光谱参数。并以峰面积比值A₇₂₄/A_{1 736}的平均值作为横坐标，青蒿素质量百分数作为纵坐标，拟合青蒿素质量百分数与峰面积比的函数关系，得到二次函数关系式为y=0.907 22x2+0.465 93x(0<x<0.9)，相关系数r为0.992 65。最后，将该关系式用于中药复方制剂青蒿素哌喹片中青蒿素的含量检测分析，按照此方法计算得到的青蒿素含量值与真实值（14.29%）比较，相对误差较小（<10%），可以初步建立起青蒿素含量的拉曼光谱测试方法。本文提出应用激光拉曼光谱技术，以期实现原位、无损和快速检测。在青蒿素标准样品实验研究基础上，确定青蒿素拉曼光谱参数和实验条件，建立起复方制剂中青蒿素含量的实验分析方法，为青蒿素的研究和应用开创新的局面。

Quantitative Measurement of Artemisinin Content in Chinese Traditional Compound Medicine by Raman Spectroscopy

Artemisinin is a kind of sesquiterpene lactone drug containing peroxide groups, which is extracted from Artemisia annua L. Now it has become an antimalaria drug recommended by the world health organization. The compound preparation of Chinese material medica containing artemisinin has strong anti-malarial, antibacterial and immunomodulatory effects. However, there is no measurable and generalized standard for controlling and evaluating the quality of the compound preparations. Currently, the main researches of artemisinin compound preparations focus on the qualitative analysis of the active ingredients, and only less work is aimed at how to quantitatively analyse the artemisinin content. Therefore, it is urgent to develop a convenient, rapid and nondestructive method to monitor the production and use of artemisinin. Laser Raman spectroscopy is a kind of molecule scattering spectroscopy, which is characterized by frequency excursion that is caused by interactions of molecules and photons, to obtain information on molecules. Laser Raman spectrometry is a potential method for quantitatively analyzing artemisinin content. In this study, we demonstrate that Laser Raman spectroscopy is a suitable and non-destructive technique to qualitatively analyze artemisinin. The spectral characteristics of artemisinin will provide standard Raman spectra for identification and analysis of artemisinin in Chinese traditional compound medicine. It is found that the vibrational mode at 724 cm-1 is related to the vibration of peroxide groups, and also it is the key to verifying its anti-malarial function, in Artemisinin. In addition, the phonon mode at 1 736 cm-1 is not affected by the surrounding vibrational modes and its intensity is strong. The vibrational mode at 1 736 cm-1 corresponds to the vibrational mode of lactone bond, so it could be used for detecting artemisinin. Hence, this paper attempts to adopt the characteristic vibrational modes at 724 and 1 736 cm-1 simultaneously to analyze Artemisinin qualitatively and quantitatively. A series of artemisinin/flour binary mixture with different artemisinin mass percentage were synthesized with the aim of obtaining Raman spectral parameters at 724 and 1 736 cm-1. The average of Raman peak area ratios in accordance with the abscissa, the ordinate artemisinin content for result analysis. Afterwards the results are analyzed through nonlinear regression in order to get the function between the artemisinin content and Raman peak area ratios. The quadratic function is y=0.907 22x2+0.465 93x(0<x<0.9), and its correlation coefficients is 0.992 65. In addition, measurements of artemisinin content in Chinese traditional compound medicine by Laser Raman analysis were carried out on artemisinin-piperaquine tablets. The values of artemisinin content obtained by Laser Raman spectroscopy technology and true values (14.29%) are in good agreement with each other (relative errors <10%). This research provides a fundamental analysis tool for determining artemisinin content quantitatively by using Laser Raman spectroscopy. Experiment results demonstrate that this method has the potential for obtaining artemisinin content in Chinese traditional compound medicine.