高效液相色谱法测定凉粉草中天然有机酸——苯甲酸

The analysis of natural acid- benzoic acid from mesona chinensis benth by HPLC

建立了定性、定量测定凉粉草中天然有机酸-苯甲酸的高效液相色谱方法。首先,通过碱性溶液提取、高效液相色谱分离测定,在植物凉粉草中分离鉴定出一种新的天然有机酸成分-苯甲酸。样品经处理后,使用三种不同的色谱柱,在不同的色谱条件下,均呈现出与苯甲酸标准品相同的保留时间,待测物与标准苯甲酸的光谱图也完全相符,证实了植物凉粉草中天然苯甲酸的存在。其次,通过对提取液的种类、提取液的浓度、提取时间和温度进行优化,得到最佳定量提取条件为:凉粉草样品经0.6%的氢氧化钠溶液浸泡30min后,在40℃下超声提取40min。离心后,用1+1的盐酸调节提取液pH至约7.0,用液相色谱进行测定。色谱柱为ZORBAX Eclipse XDB-C18(5μm,4.6mm i.d.×250mm),以v(甲醇)∶v(20mmol/L的乙酸铵溶液)=5∶95为流动相,二极管阵列检测器测定,检测波长230nm。该方法在0.2 10mg/L范围内呈良好线性关系,相关系数为1,定量限(S/N≥10)为0.020g/kg,在0.100、0.500g/kg加标水平下的回收率均为93%,相对标准偏差小于3.9%。本方法可为其他植物中天然有机酸化学成分的定性、定量测定提供参考。

A high performance liquid chromatography （HPLC） was developed for the qualitative and quantitative determination of benzoic acid in mesona chinensis benth.Benzonic acid,a new natural component,was discovered from mesona chinensis benth for the first time and extracted by basic aqueous solution and determined by high performance liquid chromatography.The target compound was proved to be benzonic acid under different conditions of HPLC,such as chromatogram columns,the mobile phase proportion and the spectrogram.Then,based on the extracting solutions,the extracting concentrations,time and temperature were optimized.The optimum extraction condition was：soaking with 0.6％ sodium hydrate for 30min,ultrasonic treatment 40 minutes at 40℃.Mter centrifuge,adjusting pH value to about 7.0 by hydrochloric acid （1 ∶ 1）.The extracts was analyzed by HPLC on ZORBAX Eclipse XDB-C18 （5μm,4.6mm i.d.× 250mm） column.The component of mobile phase was v （methanol） ∶ v （20mmol/L ammonium acetate） =5 ∶ 95,and detection was at 230nm with DAD.The results showed that there is a good linear relationship in the range of 0.2 ～ 10mg/L with correlation coefficient of 1.The limit of quantitation （LOQ） was 0.020g/kg and the recoveries （n =6） at spiked levels of 0.100、0.500g/kg were all 93％ with the relative standard deviation of less than3.9％.This study provides an approach for qualitative and quantitative determination of natural organic acid in other plants.