Development of a High-Coverage Quantitative Metabolome Analysis Method Using Four-Channel Chemical Isotope Labeling LC–MS for Analyzing High-Salt Fermented Food

Abstract

Metabolome analysis of high-salt fermented food can be an analytical challenge, as the salts can interfere with the sample processing and analysis. In this work, we describe a four-channel chemical isotope labeling (CIL) LC−MS approach for a comprehensive metabolome analysis of high-salt fermented food. The workflow includes metabolite extraction, chemical labeling of metabolites using dansyl chloride, dansylhydrazine, or p-dimethylaminophenacyl bromide reagents to enhance separation and ionization, LC-UV measurement of the total concentration of dansyl-labeled metabolites in each sample for sample normalization, mixing of 13 C-and 12 C-reagent-labeled samples, high-resolution LC−MS analysis, and data processing. Metabolome analysis of fermented foods, including fermented red pepper (FRP) sauce, soy sauce, and sufu (a fermented soybean food), showed unprecedented high metabolic coverage. Metabolome comparison of FRP, soy sauce, and sufu, as well as soy sauce and sufu, indicated great diversity of metabolite types and abundances in these foods. In addition, we analyzed two groups of samples of the same type, FRP with 10% (w/w) and 15% (w/w) salt contents, and detected large variations in multiple categories of metabolites belonging to a number of different metabolic pathways. We envisage that this CIL LC−MS approach can be generally used for metabolomic studies of high-salt fermented food. CIL LC−MS allows high-coverage identification and quantification that could not be done using other methods.

Chemical Composition of Commercial Cannabis

“…However, the performance of our assay was somewhat limited due to the lack of availability of authentic isotopically labeled phenolic standards. The use of chemo-selective labeling methods 65 could potentially increase the analytical coverage by a factor of two or more. Because cannabis is an intensely cultivated crop, it is susceptible to a wide range of bacterial, fungal, parasitic, or insect attacks.…”

“…However, the performance of our assay was somewhat limited due to the lack of availability of authentic isotopically labeled phenolic standards. The use of chemo-selective labeling methods 65 could potentially increase the analytical coverage by a factor of two or more.…”

Challenges and recent advances in quantitative mass spectrometry‐based metabolomics

“…107 It is then necessary to use several reagents, requiring longer and more complex sample preparation. Li et al 105 have described a four-channel chemical isotope labelling approach applied for quantitative metabolome analysis in different food products.…”

“…The use of reagents possessing a characteristic group, such as bromine or quaternary amine group, can facilitate the detection of certain compounds 99–102 . Another advantage is the possibility of using other detection techniques to quantify the total amount of derivatized metabolites for normalization purposes 103–105 . Some derivatization agents, such as dansyl or phenylisothiocyanate, have unique UV absorbance or fluorescence properties amenable to normalization.…”

Enhanced Identification of Ginsenosides Simultaneously from Seven Panax Herbal Extracts by Data-Dependent Acquisition Including a Preferred Precursor Ions List Derived from an In-House Programmed Virtual Library

“…It is crucial to establish precise and fit-for-purpose analytical techniques that can implement the characterization and quantification of metabolites for a number of research areas (e.g., natural product research and food science). , At the same time, it is still confronted with challenges for the detection and analysis of the metabolites owing to the severe endogenous interference and, in most cases, the low content in a complex matrix. In recent years, liquid chromatography/mass spectrometry (LC/MS) has provided a powerful and practical method enabling the systematic characterization of known compounds, as well as the discovery of potentially new components from natural products. − Nowadays, the utilization of multidimensional chromatography (MDC) greatly improves the selectivity and expands the peak capacity, , while various MS strategies are available to maximize the qualitative MS information supporting the identification of metabolites. , The utilization of various scan methods has been demonstrated as potent in the in-depth characterization and multicomponent quantitation of various metabolites from herbal extracts. , …”