Targeted HPTLC Profiling and UV Spectrophotometric Estimation of Key Polyphenols Present in Brassica sp.: A Comparative Phytochemical Analysis

Brassica sp., including kale, cabbage, broccoli, and cauliflower, contains bioactive compounds like flavonoids and phenolic acids, which make them nutritionally and therapeutically valuable. This study aimed to detect polyphenolic compounds in selected Brassica sp. (kale, cabbage, broccoli, and cauliflower) and to optimize a mobile phase to obtain better separation of these polyphenolic compounds using High-Performance Thin-Layer Chromatography (HPTLC). Standards including rutin, quercetin, chlorogenic acid, caffeic acid, and caffeine were selected based on their known antioxidant activities and occurrence in leafy vegetables. The dried powdered samples were extracted using methanol-assisted sonication. Multiple mobile phase systems were screened to achieve optimal separation, and the best-performing system, butanol: ethyl acetate:water:formic acid (9:2:1:0.5), was selected for the chromatographic profiling. The choice of mobile phase significantly influenced the resolution, migration behaviour, and detectability of phytochemical constituents. Distinct chromatographic bands were observed in each Brassica sample, indicating the presence of phenolic compounds. However, all standards were not consistently detected, probably due to low concentration, lack of certain compounds, or to extraction difficulties. This study highlights the critical importance of mobile phase selection in HPTLC fingerprinting and emphasizes the need to align extraction and chromatographic conditions with the expected phytochemical profile of the target plant group. Understanding the presence of flavonoids and phenolic acids in Brassica sp. is valuable for both nutritional evaluation and analytical standardization.