Derivatization of N-Acyl Glycines by 3-Nitrophenylhydrazine for Targeted Metabolomics Analysis and Their Application to the Study of Diabetes in Mice

N-Acyl glycines (NAGlys) are an important class of metabolites in the detoxification system of the human body. They have been used in the diagnosis of several metabolic diseases. Liquid chromatography-mass spectrometry (LC-MS) is the most frequently used NAGlys detection platform. Here, we describe a simple and sensitive method of NAGlys detection by LC-MS in plasma and urine samples. This approach is based on the use of a derivatization reagent, 3-nitrophenylhydrazine. The reaction is quick in aqueous solution, and no quenching step is needed. To expand the coverage of NAGlys when standards are not available, NAGlys were first identified based on high-resolution LC-MS. Quantification was subsequently carried out on triple quadrupole LC-MS. This approach allowed a much broader measurement of NAGlys (41 NAGlys in total), especially when authentic standards are unavailable. Comprehensive analysis of NAGlys with this new method was applied in plasma and urine samples of db/db diabetic and non-diabetic db/m+ control mice. The majority of detected NAGlys were altered with high differentiation ability in plasma and urine samples from diabetic and non-diabetic mice. These identified NAGlys hold the potential to be diagnostic biomarkers for type II diabetes and diabetic complications.

Automated summary

In this study, a simple and sensitive method for the detection of N-acyl glycines (NAGlys) in plasma and urine samples using liquid chromatography-mass spectrometry (LC-MS) is described. The use of a derivatization reagent, 3-nitrophenylhydrazine, allows for quick reaction in aqueous solution without the need for a quenching step. NAGlys were first identified based on high-resolution LC-MS and subsequently quantified on triple quadrupole LC-MS. This approach enables a broader measurement of NAGlys and potential identification of diagnostic biomarkers for type II diabetes and diabetic complications.