Unveiling the Chemical Composition Differences in Ginseng Extract Fermentation Using LC/Q-TOF Technology and Multilevel Qualitative Analysis Strategies

Significance of topic

Ginseng has been used for centuries in traditional medicine due to its diverse therapeutic activities attributed primarily to ginsenosides. Fermentation can transform these bioactive compounds, potentially unlocking new molecules and enhancing efficacy. Understanding how fermentation alters ginsenoside profiles is critical for developing more potent ginseng-based products and advancing functional food research.

Study objectives and overview

This work compares pre- and post-fermentation ginseng extracts to:

• Identify differential ginsenosides and other metabolites
• Map potential biotransformation pathways induced by microbial fermentation
• Assess how these chemical changes may enhance bioactivity

Used instrumentation

The analysis employed:

• Agilent 1290 Infinity II liquid chromatography system
• Agilent 6546 quadrupole time-of-flight mass spectrometer (LC/Q-TOF)
• Agilent InfinityLab Poroshell EC-C18 column (150 mm)
• MassHunter Explorer software with TCM PCDL library
• SIRIUS CSI:FingerID and feature-based molecular networking tools

Methodology

Pre- and post-fermentation ginseng samples were extracted in water/methanol, ultrasonicated, centrifuged, and analyzed by a 35-minute LC gradient from 10 % to 100 % organic. MS data were acquired in negative mode for m/z 60-1500. A pooled quality control sample enabled iterative MS/MS acquisition for in-depth structural annotation. Differential features were filtered by fold change ≥ 2 and p-value < 0.01.

Main results and discussion

Principal component analysis clearly separated fermented and unfermented samples. Volcano plots revealed dozens of significantly up- or downregulated compounds. Key findings include:

• Increase in mono- and di-glycosyl ginsenosides as a result of enhanced glycosidic hydrolysis
• Decrease in acylated and high-polysaccharide forms due to faster acyl-group removal
• Production of oxidized or rearranged compounds, notably OCO-type ginsenosides

Rapid annotation against the TCM PCDL library identified 33 known ginsenosides, including a marked rise in Rg3 after fermentation. Unmatched features were characterized by SIRIUS CSI:FingerID, unveiling novel malonylated and hydroxylated ginsenosides. Feature-based molecular networking further highlighted clusters of related analogs and suggested biotransformation paths for acyl hydrolysis, glycosyl hydrolysis, and rearrangement reactions.

Benefits and practical applications

Enhanced conversion to more bioavailable mono-glycosides and generation of unique OCO-type compounds suggest that fermented ginseng extracts may offer superior pharmacological effects. This refined profiling supports targeted development of functional foods, nutraceuticals, and standardized quality control methods in industrial or research laboratories.

Future trends and applications

Emerging opportunities include:

• Microbial strain optimization to tailor ginsenoside transformation
• High-throughput screening combining LC/Q-TOF with machine learning for rapid metabolome profiling
• Integration of fermentation insights into large-scale production of value-added ginseng derivatives

Continued advances in software-driven structural annotation and molecular networking will deepen understanding of complex biotransformations in plant extracts.

Conclusion

Multilevel qualitative analysis using LC/Q-TOF combined with library matching, SIRIUS CSI:FingerID, and molecular networking has elucidated how fermentation reshapes ginseng’s chemical makeup. The study demonstrates targeted conversion of ginsenosides into more bioactive forms and discovers novel derivatives, laying the groundwork for improved ginseng-based products.

References

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