一测多评同时测定款冬花中10个成分的含量

目的: 建立款冬花中10个活性成分(新绿原酸、绿原酸、隐绿原酸、咖啡酸、芦丁、金丝桃苷、异绿原酸B、异绿原酸A、异绿原酸C及款冬酮)的一测多评含量测定方法。 方法: 采用Kromasil C₁₈色谱柱(250 mm×4.6 mm,5 μm);以乙腈-0.1%磷酸水溶液为流动相;梯度洗脱(0→10→25→30→35→40 min,10%→16%→30%→90%→95%→95%乙腈);分段变波长测定(0~15 min,326 nm,检测新绿原酸、绿原酸、隐绿原酸及咖啡酸;15~20 min,254 nm,检测芦丁、金丝桃苷;20~30 min,326 nm,检测异绿原酸B、异绿原酸A及异绿原酸C;30~40 min,220 nm,检测款冬酮);流速1.0 mL·min^-1;柱温30 ℃。以绿原酸为参照物建立与其余成分的相对校正因子,并计算其含量,实现一测多评。同时采用外标法测定该10个成分的含量,并比较二者的差异,以验证一测多评法的准确性和可行性。 结果: 在一定的线性范围内,绿原酸与新绿原酸、隐绿原酸、咖啡酸、芦丁、金丝桃苷、异绿原酸B、异绿原酸A、异绿原酸C及款冬酮的相对校正因子分别为0.997、0.998、1.762、0.601、0.865、1.229、1.232、1.233、0.743;且在不同实验条件下重现性良好[相对校正因子的RSD(n=6)<2.4%];6批款冬花中10个成分的计算值与实测值间无显著差异。 结论: 本文建立的一测多评法控制款冬花的质量可行、准确。

Objective: To establish a method for simultaneous assay of 10 main active constituents (neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, rutin, hyperin, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C and tussilagone) in Farfarae Flos by quantitative analysis of multi-components by single marker (QAMS). Methods: The chromatographic separation was achieved on a Kromasil C₁₈ column (250 mm×4.6 mm,5 μm), the mobile phase was acetonitrile-0.1% phosphoric acid aqueous solution with gradient elution(0→10→25→30→35→40 min, 10%→16%→30%→90%→95%→95% acetonitrile). The detection wavelengths were changeable (0-15 min, 326 nm for detection of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid and caffeic acid; 15-20 min, 254 nm for detection of rutin and hyperin; 20-30 min, 326 nm for detection of isochlorogenic acid B, isochlorogenic acid A and isochlorogenic acid C; 30-40 min, 220 nm for detection of tussilagone). The flow rate was 1.0 mL·min^-1 and the column temperature was 30 ℃. Chlorogenic acid was used as the reference, the calibration factor of other 9 constituents to that of chlorogenic acid were calculated respectively to get each relative calibration factor.And the contents of active constituents were calculated by the relative calibration factor to realize QAMS.At the same time, external standard method was used to detect the contents of the 10 constituents,the two methods were compared to validate the accuracy and feasibility of the new method. Results: Relative correction factors of neochlorogenic acid, cryptochlorogenic acid, caffeic acid, rutin, hyperin, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C and tussilagone with reference to chlorogenic acid were 0.997, 0.998, 1.762, 0.601, 0.865, 1.229, 1.232, 1.233, and 0.743 respectively, and repeatability was good in different experimental conditions[RSD of ele correction factors less than 2.4% (n=6)]. No significant differences were found in the quantitative results of the 10 active constituents in 6 batches of Farfarae Flos between the measured value and the calculated value. Conclusion: The QAMS is feasible and accurate to evaluate the contents of the 10 active constituents in Farfarae Flos.