LC-MS/DIA-based strategy for comprehensive flavonoid profiling: an Ocotea spp. applicability case

Area of Science:

• Plant biochemistry
• Analytical chemistry
• Metabolomics

Background:

• Flavonoids are crucial plant secondary metabolites with diverse biological activities.
• Comprehensive profiling of glycosylated flavonoids, especially in the Ocotea genus, remains challenging.
• Existing methods often lack the depth and reliability for simultaneous precursor and fragment analysis.

Purpose of the Study:

• To develop and validate a liquid chromatography-mass spectrometry with data-independent acquisition (LC-MS/DIA) strategy for comprehensive glycosylated flavonoid profiling.
• To apply this strategy to analyze flavonoids in six Ocotea plant species.
• To identify novel and known flavonoid structures within the Ocotea genus.

Main Methods:

• Utilized a liquid chromatography-mass spectrometry with data-independent acquisition (LC-MS/DIA) approach.
• Employed the open-source MZmine 3 software for data processing, enabling simultaneous analysis of precursors and fragments.
• Applied gas-phase fragmentation reactions for structural elucidation and confirmation.

Main Results:

• Successfully profiled glycosylated flavonoids across six Ocotea species, annotating over 50 flavonoid backbones per species.
• Identified 49 potentially novel and 45 known flavonoids within the genus, with kaempferol and quercetin O-glycosides being prevalent.
• Achieved the first-time annotation of the apigenin flavone backbone in most examined Ocotea species, primarily as C-glycosides.

Conclusions:

• The developed LC-MS/DIA strategy offers a robust and comprehensive approach for glycosylated flavonoid profiling.
• The study significantly expands the known flavonoid diversity in Ocotea, revealing new insights into their chemical composition.
• Highlights the potential of LC-MS/DIA data for broad and reliable flavonoid analysis in plant metabolomics.