UPLC-Triple TOF-MS/MS based metabolomics approach to reveal the influence of nitrogen levels on Isatis indigotica seedling leaf

Isatis indigotica Fortune represents a significant economic resource in China, which the leaves (Daqingye) remain until now important and popular herbal drug in traditional Chinese medicine (TCM). Nitrogen nutrition is in great demand during plant growth stage, and nitrogen levels are usually related with plants biosynthesis of primary and secondary metabolites. To investigate the influence of nitrogen nutrition levels on various metabolites of Isatis leaf, untargeted metabolomics analysis based on LC-MS (UPLC-Triple-TOF system) was used to obtain the response of Isatis leaf under different nitrogen levels: 0 mmol.L-1(deficient), 7.5 mmol.L-1 (low) and 15 mmol.L-1 (high). Based on the results, dihydroxyindole, anthranilic acid, N-acetylanthranilic acid significantly discriminated the three treatments. Low nitrogen generally led to the decrease of most amino acids, while the increase of phenylalanine, indole-3-pyruvate acid, and some specialized metabolites including quercetin, indigo and indirubin were observed. The change in content of indole-3-acetic acid (IAA) was speculated to be the potential low nitrogen tolerance mechanism of I. indigotica. Our findings suggest that nitrogen levels significant affect metabolic capacity of Isatis leaf; low nitrogen treatments induce the accumulation of active ingredients. This result will provide leads into understanding the physiological adaption to nitrogen levels of Isatis leaf and quality improvement research of Daqingye.