A Comprehensive Metabolite Profiling Approach Utilizing In vivo Metabolic Reaction-Based Mass Difference Screening with Ultra-High-Performance Liquid Chromatography and Quadrupole-Time-of-Flight Mass Spectrometry for the Huangqi-Danshen Herb Pair

Abstract

The goal of studying traditional Chinese medicine is to determine the biological roles played by the absorbed metabolites
and prototypes in living organisms. However, due to endogenous interference and low metabolite abundance, complete in vivo
component profiling is challenging. Thus, a methodical approach was suggested for the systematic screening and profiling of external
components in biological matrices, using the mass differences between prototypes and their metabolic reaction products as a basis.
Research Tools and Procedures: An extensive database based on metabolic reactions was constructed by collecting 247 metabolic
reactions from the relevant literature on the Huangqi-Danshen (HD) herb pair. The mass differences were automatically computed
using R programming and compared with the database in order to screen candidate components using the preprocessed data from
experimental MS1 that was based on ultra-high-performance liquid chromatography combined with quadrupole-time-of-flight mass
spectrometry. The components were then identified across the MS2 dataset. After oral administration of HD, 164 components were
noted in the plasma samples of the rats. Out of them, 20 were validated using the reference standards. Amino acid dehydration,
methylation, cysteine S-binding, glucuronidation, sulfation, and multistep reactions are prominent metabolic processes. Final
Thoughts: This research paved the way for future studies to clarify HD's therapeutic benefits by revealing its metabolic features and
biotransformation regulation. Beneficial tools for assessing metabolites in complex systems are provided by the suggested method.