Nontargeted Modification-Specific Metabolomics Study Based on Liquid Chromatography–High-Resolution Mass Spectrometry

Abstract

Modifications of genes and proteins have been extensively studied in systems biology using comprehensive analytical strategies. Although metabolites are frequently modified, these modifications have not been studied using -omics approaches. Here a general strategy for the nontargeted profiling of modified metabolites, which we call "nontargeted modification-specific metabolomics", is reported. A key aspect of this strategy was the combination of in-source collision-induced dissociation liquid chromatography-mass spectrometry (LC-MS) and global nontargeted LC-MS-based metabolomics. Characteristic neutral loss fragments that are specific for acetylation, sulfation, glucuronidation, glucosidation, or ribose conjugation were reproducibly detected using human urine as a model specimen for method development. The practical application of this method was demonstrated by profiling urine samples from liver cirrhosis patients. Approximately 900 features were identified as modified endogenous metabolites and xenobiotics. Moreover, this strategy supports the identification of compounds not included in traditional metabolomics databases (HMDB, Metlin, and KEGG), which are currently referred to as "unknowns" in metabolomics projects. Nontargeted modification-specific metabolomics opens a new perspective in systems biology.

Global Metabolic Profiling of Plasma Shows that Three-Year Mild-Caloric Restriction Lessens an Age-Related Increase in Sphingomyelin and Reduces L-leucine and L-phenylalanine in Overweight and Obese Subjects

“…Third, although a large number of metabolites were detected by UPLC-LTQ-Orbitrap MS in this study, most of them are currently unidentified. Large databases of endogenous biomolecules have not been constructed yet for use with LC-MS-based techniques for metabolomics research [47]. Fourth, the relatively small sample size used in this study may not be sufficiently large to detect all long-term MCD-associated metabolic changes.…”

Systems-based approaches to unravel multi-species microbial community functioning

“…As discussed above, targeted metabolomics can be powerful in the context of diagnostics [80]. Also, methodologies are being developed to gain some insights into the large fraction of unknown metabolites typically identified in untargeted experimental strategies [77], [78]. Despite the wealth of information that can be derived from omics datasets, pathway connectivity and microbial interrelationships are not easily accessed.…”

“…In this study, metabolomics was combined with isotope labelling, which led to significant improvements in chemical formula prediction particularly for large metabolites [76]. In order to gain some insights into unknown metabolites typically detected in untargeted investigations, modification-specific metabolomics was developed [77]. This novel approach involves the detection of metabolite modification encompassing acetylation, sulfation, glucuronidation, glucosidation and ribose conjugation.…”

“…This novel approach involves the detection of metabolite modification encompassing acetylation, sulfation, glucuronidation, glucosidation and ribose conjugation. The inclusion of modification information to the mass feature during database searches drastically reduces the number of matches for metabolite identification and therefore significantly decreases the time required for this process [77]. Similarly, in order to improve metabolite identification rates in untargeted metabolite profiling, Mitchell et al (2014) developed an algorithm for the detection of functional groups within metabolite databases [78].…”

Coupled Enzymatic Treatment and Mass Spectrometric Analysis for Identification of Glucuronidated Metabolites in Human Samples

“…Mass spectrometry (MS) is the most frequently reported technique for investigating glucuronides in human samples . The most common MS methods are multiple reaction monitoring (MRM), neutral loss analysis, and precursor ion scans . Targeted analyses of glucuronides have mainly been reported in the fields of drug metabolism, doping control, and diet metabolism so far .…”