Lin Guang-Huey, Chang Chung-Yu, Lin Huei-Ru
Department of Microbiology, Tzu Chi University, Hualien, Taiwan.
Institute of Medical Biotechnology, Tzu Chi University, Hualien, Taiwan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2015 Apr 15;988:53-8. doi: 10.1016/j.jchromb.2015.02.025. Epub 2015 Feb 24.
Indole-3-acetic acid (IAA) is produced from tryptophan through five synthesis pathways. A comprehensive method for the quantification of IAA and biosynthesis-related intermediates in a culture medium was developed. Sample preparation was simple with protein precipitation. The analytes were separated on a superficially porous C18 silica column and detected by electrospray ionization-tandem mass spectrometry in the positive ion multiple reaction monitoring mode. The limit of detection was 0.05 μM, and the lower limits of quantification ranged from 0.05 to 2 μM. The intra-day and inter-day precision and accuracy were less than 13.96%. Ion suppression was observed, and the deuterated internal standards were used to compensate for the matrix effect. The method was applied to analyze changes in tryptophan catabolism in a culture medium of Pseudomonas putida. The proposed method is robust and suitable for the systematic profiling of IAA biosynthesis in culture supernatant.
吲哚 - 3 - 乙酸（IAA）通过五条合成途径由色氨酸产生。开发了一种用于定量培养基中IAA和生物合成相关中间体的综合方法。通过蛋白质沉淀进行样品制备很简单。分析物在表面多孔C18硅胶柱上分离，并通过电喷雾电离串联质谱在正离子多反应监测模式下进行检测。检测限为0.05μM，定量下限范围为0.05至2μM。日内和日间精密度和准确度均小于13.96％。观察到离子抑制现象，并使用氘代内标来补偿基质效应。该方法用于分析恶臭假单胞菌培养基中色氨酸分解代谢的变化。所提出的方法稳健，适用于培养上清液中IAA生物合成的系统分析。
