稚儿灵颗粒HPLC指纹图谱及多指标成分的含量测定

Study on HPLC Fingerprint and Multi-index Determination of Zhierling Granules

目的:建立稚儿灵颗粒HPLC指纹图谱,测定其中4种成分的含量,并进行化学模式识别分析。方法:采用高效液相色谱法,Agilent ZORBAX Eclipse Plus(4.6 mm×250 mm,5μm)色谱柱;流动相为乙腈-0.05%磷酸(梯度洗脱);检测波长:230 nm;柱温:30℃;流速:1.0 mL·min^(-1);进样量:10μL。以芍药苷为参照,绘制19批稚儿灵颗粒的HPLC指纹图谱,采用《中药色谱指纹图谱相似度评价系统(2012版)》进行相似度评价,确定共有峰;采用相同的HPLC法测定稚儿灵颗粒中4种成分的含量;采用SPSS19.0软件进行聚类分析。结果:19批样品HPLC指纹图谱确定10个共有峰,各批样品相似度均在0.94以上。芍药内酯苷、芍药苷、甘草苷、橙皮苷检测质量浓度的线性范围分别为1.9384~38.7688μg·mL^(-1)(r=0.9999)、7.7362~116.0437μg·mL^(-1)(r=1)、1.9494~38.9880μg·mL^(-1)(r=0.9999)、3.4384~68.7685μg·mL^(-1)(r=1);精密度、稳定性、重复性试验的RSD均<2%;平均加样回收率分别为97.9%(RSD=1.1%,n=6)、97.9%(RSD=0.8%,n=6)、98.8%(RSD=1.5%,n=6)、103.2%(RSD=0.6%,n=6)。聚类分析显示,19批样品可聚为4类:S1﹑S2﹑S7为一类;S3~S6为一类;S8~S10为一类;S11~S19为一类。结论:所建指纹图谱特征性强且操作简便,可用于该品的质量控制;含量测定方法准确、可靠,可用于同时测定其中4种有效成分的含量。

Objective:To establish HPLC fingerprint of Zhierling granules,determine their contents of four components and carry out chemical pattern recognition analysis.Method:HPLC was done with an Agilent Zorbax eclipse plus(4.6 mm×250 mm,5μm)column.The mobile phase was acetonitrile-0.05%phosphoric acid with a gradient.The flow rate was 1.0 mL·min^(-1).The injection volume was 10μL.The detection wavelength was 230 nm.The HPLC fingerprints of 19 batches of Zhierling granules were drawn with reference to paeoniflorin,and the Similarity Evaluation System of Chromatographic Fingerprints of Traditional Chinese Medicine(2012 Edition)was used to determine the common peaks.Meanwhile,the same HPLC method was used to determine the contents of four components in Zhierling granules;the cluster analysis was carried out by the SPSS 19.0 software.Results:The HPLC fingerprints of 19 batches of Zhierling granules determined 10 common peaks,and the similarity of each batch of the samples was above 0.94.The linear ranges for determination of paeoniflorin,paeoniflorin,glycyrrhizin and hesperidin were 1.9384-38.7688μg·mL^(-1)(r=0.9999),7.7362^(-1)16.0437μg·mL^(-1)(r=1),1.9494-38.9880μg·mL^(-1)(r=0.9999)and 3.4384-68.7685μg·mL^(-1)(r=1),respectively.The RSD values of precision,stability and repeatability tests were less than 2%;the average recoveries of sample addition were 97.9%(RSD=1.1%,n=6),97.9%(RSD=0.8%,n=6),98.8%(RSD=1.5%,n=6)and 103.2%(RSD=0.6%,n=6),respectively.The results of cluster analysis showed that the 19 batches of samples could be clustered into 4 groups:S1,S2 and S7;S3-S6;S8-S10;S11-S19.Conclusion:The established fingerprint is characteristic and easy to operate,which can be used for the quality control of Zhierling granules,and the content determination method is accurate and reliable,which can be used for the simultaneous determination of four effective components.