Scanning of chicoric acid in different parts of Cichorium intybus by high-performance thin-layer chromatography with quantitation by image analysis

Cichorium intybus, commonly known as chicory, is a widespread plant, valued for its medicinal properties and use as a coffee substitute, chicoric acid being one of its key constituents. This study investigates the variation in chicoric acid content across different parts of the chicory plant-flowers, leaves, root and aerial parts. Three extraction methods, namely maceration in 70% ethanol, ultrasound-assisted extraction (UAE), and enzyme-assisted UAE were compared. A new optimized high-performance thin-layer chromatography (HPTLC) method was used to determine the chicoric acid quantities. The chicoric acid was separated on HPTLC silica gel F₂₅₄ plates developed with a mixture of ethyl acetate‒formic acid‒acetic acid‒water (8:0.2:2:1.8, v/v) as the mobile phase. The detection was performed in UV light at 366 nm after derivatization with natural product (NP) and polyethylene glycol solution (PEG). The quantification was performed by image analysis (IA) using ImageJ software, with the green color channel offering the most sensitive detection (LOD = 2 ng/band and LOQ = 6 ng/band). The analysis revealed that leaves contained the highest concentration of chicoric acid, while roots exhibited the lowest. Enzyme-assisted UAE proved to be the most effective method. Antioxidant activity was tested using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical revealing that the chicoric acid possesses this activity. These findings highlight chicory leaves as a valuable source of chicoric acid and emphasize the importance of choosing effective extraction methods to maximize the yield of bioactive compounds.