定量核磁共振波谱法测定川芎中藁本内酯含量

A validated ~1H-qNMR method for direct quantification of ligustilde in Ligusticum chuanxiong Hort

建立氢核磁定量方法(~1H-qNMR)测定川芎中藁本内酯含量。以川芎中藁本内酯含量为指标,采用单因素实验优选提取工艺;以1,4-二硝基苯为内标、氘代DMSO-d6为溶剂,观测频率600 MHz,脉冲延迟时间1 s,采样次数32次,采集藁本内酯的1H-NMR谱图,定量峰为藁本内酯9位质子信号δ6.18~6.19;采用高效液相色谱法(HPLC)对氢核磁定量方法进行验证。结果表明优选出的藁本内酯提取工艺条件为80%乙醇、料液比20 mg/mL、静置30 min、超声波辅助提取30 min;~1H-qNMR结果显示藁本内酯在115.3~3 546.7μg/mL浓度范围内呈现良好的线性关系(r=0.999 9),精密度相对标准偏差为0.017%,回收率为96.30%~102.87%,相对标准偏差为2.43%,说明该方法准确性良好。~1H-qNMR测定川芎中藁本内酯平均含量为1.805%,HPLC测定的藁本内酯的平均含量为1.830%,两种方法所得数据基本一致。提示所建立的氢核磁定量方法操作简便、结果准确、可靠,可用于川芎中藁本内酯含量测定。

To establish a quantitative1 H-NMR method(1 H-qNMR)for determining the ligustilide in Ligusticum chuanxiong Hort. Single factor experiment was used to optimize the extraction process with the content of ligustilide as index,in this experiment,deuterated DMSO-d6 was used as the test solvent,1,4-dinitrobenzene was used as the internal standard,the observation frequency was 600 MHz,the pulse delay time was 1 s,and the sampling times were 32 times. The1 H-NMR spectra of ligustilide were collected,and the quantitative peak was of ligustilide was δ6.18 6.19(H-9). High performance liquid chromatography(HPLC) was used to validate the quantitative method of hydrogen nuclear magnetic resonance. The optimum extraction conditions was as follows:80% ethanol,20 mg·mL-1 of solid-liquid ratio,30 min of standing,30 min of ultrasound-assisted extraction;1 H-qNMR results showed that ligustilide showed a good linear relationship in the concentration range of 115.33 546.7 g·mL-1(r=0.999 9),the relative standard deviation of precision was 0.017%,and the recovery was 96.3%102.87% and the relative standard deviation was 2.43%,which showed that the method was accurate. The average content of ligustilide of Ligusticum chuanxiong Hort was 1.805% by1 H-qNMR and 1.830% by HPLC,which were basically consistent with each other. The results showed the method was simple,accurate and reliable,and could be used for the determination of ligustilide in Ligusticum chuanxiong Hort.