Rapid Analysis of Monosaccharides in Sub-milligram Plant Samples Using Liquid Chromatography-Mass Spectrometry Assisted by Post-column Derivatization

摘要

Monosaccharides play important roles in plant growth and development, and their biofunctions are closely related to their endogenous contents. Therefore, the determination of monosaccharides is beneficial for the further study of monosaccharide biofunction, In this work, we developed a liquid chromatography-mass spectrometry analytical method assisted by a post-column derivatization technique (LC-PCD-MS) for the fast and automatic determination of 16 monosaccharides in samples. Post-column chemical derivatization of monosaccharides was performed by a reaction of monosaccharides with 4-benzylaminobenzeneboronic acid (4-PAMBA) through boronate ester formation in a three-way connector. 4-PAMBA worked as a derivatization reagent to improve the selectivity and sensitivity of monosaccharide detection by MS. The developed LC-PCD-MS method integrates LC separation, chemical derivatization, and MS detection in one run, thus greatly reducing the analysis time for each sample. The limits of detection and limits of quantification for 16 monosaccharides were in the range of 0.002-0.1 and 0.007-0.5 ng/mL, respectively. Good linearity was obtained from the linear regression, with a determination coefficient (R-2) ranging from 0.9928 to 1.0000. The relative recoveries were in the range of 80.7-117.8%, with the intra- and interday relative standard deviations less than 19.7 and 16.5%, respectively, indicating good accuracy and acceptable reproducibility of the method. Finally, the method was successfully applied to investigate the spatial and temporal distribution of 16 monosaccharides in the developing flower and germinating seed of Arabidopsis thaliana.