Effect-directed profiling of Ficus religiosa leaf extracts for multipotent compounds via 12 effect-directed assays

The interest in the therapeutic values of natural compounds from plants is growing worldwide because the development of modern synthetic drugs has not lived up to expectations. The tree Ficus religiosa native to India, China and Southeast Asia is traditionally used for curing almost 50 ailments, although the majority of the individual active compounds are not known. Hence, a hyphenated high-performance thin-layer chromatography (HPTLC) method was newly developed. It allowed a physicochemical, but especially effect-directed profiling of individual compounds present in Ficus religiosa leaves obtained from four locations (in India and Germany). Extracts of different polarities were screened for bioactivity responses and most bioactivities were found in the ethyl acetate extracts. A multi-imaging via 26 different detection modes was performed, i. e. UV/Vis/FLD, 11 microchemical derivatizations and 12 effect-directed assays (EDA). By HPTLC-UV/Vis/FLD-EDA, antibiotics against Gram-positive and Gram-negative bacteria as well as acetylcholinesterase, butyrylcholinesterase, tyrosinase, α-amylase, α-glucosidase and β-glucosidase inhibitors and radical scavenging compounds were detected. Estrogen-like or gentotoxic compounds were not detected at higher extract amounts of even 5 mg/band applied. For further characterization of three most important, multipotent, bioactive compound zones, HPTLC was hyphenated with heated electrospray ionization high-resolution mass spectrometry including fragmentation (HPTLC-HESI-HRMS/MS). Multipotent bioactive compounds discovered in the extracts were equivalently calculated in reference to well-known reference inhibitors.