UHPLC-MS/MS And NMR Analysis Of Multiple Aromatic Metabolites

The shikimate pathway is a vital biochemical process for biosynthesis of aromatic metabolites. These aromatic metabolites are also the precondition for the interaction between mammalian and its intestinal flora through co-metabolism, thus, quantitative analysis of these substances become essential.Here,26 metabolites of shikimate pathway were assigned by nuclear magnetic resonance (NMR) offering basic data for the NMR-based metabonomics studies.A sensitive and accurate method was developed for simultaneous quantification of these metabolites using ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). The linear correlation coefficients of the quantitative standard curves for these compounds are above 0.99 (r2>0.99), the limit of detection (LOD) is 0.003～0.52 pmol whilst limit of quantification (LOQ) is 0.01～2.00 pmol, which is 3～980 fold more sensitive than reported methods.Furthermore, we applied the quantitative method to study the difference content of metabolites between the rice sheath with diverse gene background, and compared the response of metabolism between them. Thirteen metabolites mediated by shikimate pathway had been detected. As a result, the variation trend of metabolites between the susceptible TN1 and the resistant B5 (including Bph14 and Bph15 genes) rice sheath is different, and the change tendency becomes closer when the gene background becomes more similar, for example, the metabolic response of TN1 and Bph15 (the resistant isogenic of TN1) rice after BPH attacked almost the same. In addition, shikimic acid, quinic acid and benzoic acid perhaps have some important roles in resisting pests attack.Moreover, we developed a method for simultaneous quantifying 50 metabolites related to gut microbiota with UHPLC-MS/MS. This study supports for the clinical research on the diagnosis of metabolic diseases and provide technical assistance for deep study in the three amino acid (tyrosine, phenylalanine and tryptophan) metabolic pathways. In order to improve separation efficiency and separation flux, variable flow rate and positive and negative mode switching have been used.