Supporting non-target identification by adding hydrogen deuterium exchange MS/MS capabilities to MetFrag

Area of Science:

• Analytical Chemistry
• Environmental Science
• Biochemistry

Background:

• Liquid chromatography-high resolution mass spectrometry (LC-HRMS) with tandem MS/MS is vital for non-targeted analysis of complex samples.
• Mass spectra alone may lack sufficient information for definitive structural elucidation of unknown compounds.

Purpose of the Study:

• To evaluate the added value of hydrogen deuterium exchange (HDX) experiments for structural characterization in LC-HRMS.
• To enhance the in silico fragmenter MetFrag with new scoring terms for improved compound identification using HDX data.

Main Methods:

• Incorporation of new scoring terms into the MetFrag in silico fragmenter to process normal and deuterated mass spectra.
• Testing the enhanced MetFrag on 762 environmental compounds and 80 metabolites.
• Analysis of hydrogen/deuterium exchange on heteroatom-bound hydrogens, observed as [M+D]+ and [M-D]- ions.

Main Results:

• HDX experiments significantly increased correct identifications (78 vs 49) and top 10 rankings in positive and negative modes for environmental compounds.
• Compounds with dual functionality showed substantial retention time shifts (up to minutes) with HDX.
• Top 10 rankings for metabolites also improved substantially when HDX information was included.

Conclusions:

• Hydrogen deuterium exchange (HDX) provides crucial additional information for the structural identification of unknown compounds using LC-HRMS.
• The enhanced MetFrag tool with HDX scoring improves the accuracy and reliability of compound identification in complex mixtures.
• The findings were validated using standard measurements and analysis of environmental samples, demonstrating practical applicability.