高效液相色谱法测定发酵液中的吲哚乙酸含量

Determination of Indole Acetic Acid Content in the Fermentation Broth Using HPLC

建立了发酵液中吲哚乙酸含量测定的高效液相色谱法。样品经乙酸乙酯萃取、分层后,取乙酸乙酯层抽真空浓缩至干,残渣用甲醇溶解定容。色谱条件为:C18柱,以乙腈∶2%醋酸水溶液=1∶4为流动相,采用紫外检测器在280nm处对样品中的吲哚乙酸进行测定高效液相色谱法测定。测定结果表明,吲哚乙酸在0-5mg/L时其峰面积与进样质量的线性相关系数达到0.9998,峰面积测定的相对标准偏差(RSD)3.5%(n=6)。试样加标回收率为97.6-106%。测定结果显示,该方法快速、灵敏、可靠,具有简便、重现性好的特点。

A high performance liquid chromatography was established to determine Indole acetic acid content in fermentation broth. Samples were extracted with ethyl acetate, and the ethyl acetate fraction was collected and vacuum concentrated until dry. The residues were dissolved with methanol and the methanol solution was prepared for detection. The absorbance of the solution was detected at 280 nm by using a UV type detector in the HPLC involving C 18 column with the mobile phase being acetonitrile-2% acetic acid water （1：4）. The quantitative and qualitative analysis was performed with external standard. Determination showed that there was a good linear calibration curve of peak area of the Indole acetic acid versus mass injected in the range from 0 to 5 mg/l （r =0.999 8）, with a relative standard deviation of the samples for the peak area detected being 3.5 % （n=6）. The recovery rate of the spiked sample of Indole acetic acid ranged from 97.6 % to 106 %. The results show that this method is rapid, sensitive and simple with high repeatability and reproducibility.