# Comparison of the phenolic and antioxidant potential of five European herbal remedies by effect-directed analysis using offline two-dimensional liquid chromatography-high resolution mass spectrometry

## TL;DR

This study compares the antioxidant properties and phenolic compounds in five European herbal plants to identify which species and plant parts have the strongest antioxidant activity.

## Contribution

The study introduces an effect-directed analysis workflow combining offline two-dimensional liquid chromatography and high-resolution mass spectrometry to link antioxidant activity with specific compounds in herbal remedies.

## Key findings

- Flavan-3-ol oligomers like procyanidin C1 were identified as major antioxidants in Agrimonia eupatoria.
- Sambucus nigra showed high antioxidant activity linked to rutin and other phenolics.
- Angelica archangelica had a coumarin-rich profile with lower antioxidant activity compared to flavonoid-rich species.

## Abstract

Herbal remedies contain various phenolic compounds. However, it remains difficult to identify the most important bioactive components and compare their effectiveness in different plant species. In this study, effect-directed analysis has been applied to five European medicinal plants. Angelica archangelica, Angelica sylvestris, Agrimonia eupatoria, Sambucus ebulus, and Sambucus nigra have been analyzed to unravel and compare their phenolic profiles and antioxidant potential. Plant extracts obtained by a sustainable microwave-assisted extraction method were fractionated using semi-preparative liquid chromatography to yield continuous fractions, and a miniaturized ABTS radical scavenging assay of the fractions was used to screen for antioxidant activity. Highly active fractions were selected for a second HPLC fractionation and analyzed with a quadrupole time-of-flight mass spectrometer using a non-targeted workflow that successfully linked antioxidant effects to specific compounds or compound classes. In the richest antioxidant fractions, flavan-3-ol oligomers such as procyanidin C1 were found in A. eupatoria, the flavonol glycoside rutin and other co-eluting phenolics in S. nigra. In contrast, A. archangelica showed a distinct metabolite profile rich in coumarins (e.g., bergapten, umbelliferone), but they contributed less to antioxidant activity compared to the flavonoid-dominated profiles of the other species. Overall, leaves and flowers contained the highest diversity and quantity of phenolic antioxidants among the plants studied. The effect-directed analysis of multiple European medicinal plants demonstrated its utility in exploring the major antioxidant compounds and highlighted significant differences in phenolic composition and antioxidant activity between species and plant parts.

The online version contains supplementary material available at 10.1007/s00216-026-06319-2.