# Consecutive Recovery of Bioactive Substances from Desmodium canadense at Different Plant Vegetation Phases by Green Extraction with Supercritical CO2 and Increasing Polarity Pressurized Liquids

## TL;DR

This study shows that leaves of Desmodium canadense collected during specific growth stages can be used to extract antioxidant-rich compounds using green extraction methods.

## Contribution

The study introduces a consecutive green extraction method to recover bioactive compounds from D. canadense at different vegetation stages.

## Key findings

- Ethanol PLE extracts showed the strongest antioxidant properties, with high TPC and ABTS•+ values at massive flowering.
- UPLC-Q-TOF-MS/MS identified 37 compounds, including C-glycosyl flavones and flavonol O-glycosides, with citric acid and vitexin being the most abundant.
- Water extracts were rich in low molecular weight organic acids, while acetone and ethanol extracts contained the highest flavonoid levels.

## Abstract

This study used high-pressure extraction to obtain antioxidant-rich fractions from Desmodium canadense leaves harvested at five vegetation phases (intensive growing to end of blooming) and to evaluate their antioxidant activity and phytochemical profile. Supercritical CO2 extraction recovered lipophilic compounds, with the highest yield at massive flowering. The remaining plant material was fractionated by pressurized liquid extraction (PLE) using acetone, ethanol, and water; the highest PLE yield was achieved with water (16.54 g/100 g DW) at the bud formation stage. Antioxidant capacity was measured using total phenolic content (TPC) and ABTS•+, CUPRAC, and ORAC assays. Overall, ethanol PLE extracts showed the strongest antioxidant properties: maximum TPC (282.1 mg GAE/gE) and ABTS•+ (1010 mg TE/gE) at massive flowering, and highest CUPRAC (853.3 mg TE/gE) and ORAC (1882 mg TE/gE) at bud formation. UPLC-Q-TOF-MS/MS profiling identified 37 compounds, mainly C-glycosyl flavones, flavonol O-glycosides, hydroxycinnamic acid derivatives, and low molecular weight organic acids. Water extracts were rich in low molecular weight organic acids, while acetone and ethanol extracts contained the highest flavonoid levels. Citric acid and vitexin were the most abundant compounds. The findings indicate that D. canadense leaves, especially harvested at budding through massive flowering, are a promising source of flavonoid-rich antioxidant extracts for nutraceutical and functional food applications.