Feature-based molecular networking-guided isolation of kaempferol glycosides from Keetia venosa (Oliv.) Bridson leaves and their antibacterial activity

Natural products have made important contributions to the fight against human diseases, including infectious diseases. However, research on natural products still faces many challenges, such as technical barriers to in-depth screening of biomaterials. The genus Keetia is a valuable source of flavonoids, although the specific types and complete profile of most of its species remain undisclosed. The flavonoid profile of the methanol crude extract and derived extracts from Keetia venosa (Oliv.) Bridson leaves was unveiled using a feature-based molecular networking-guided fractionation strategy. This approach enables a comprehensive exploration of the plant chemical diversity, including the detection of minor components often missed out during traditional isolation and characterisation. Four previously undescribed kaempferol glycosides were isolated, together with sixteen known compounds. Their structures were elucidated by examining their physical, mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopic data. Fourteen additional already reported compounds were putatively identified from a comparative analysis of their MS² features with similar analogues available in well-known databases. Furthermore, crude extract, partitioned extracts and isolated compounds were evaluated for their antibacterial activity against both Gram-positive and -negative bacteria, including Salmonella enterica, Pseudomonas aeruginosa and Klebsiella pneumoniae. The n-butanol extract, kotschyanoside A, and kaempferol 7-O-α-l-rhamnopyranoside displayed promising antibacterial activity against Gram-negative bacteria such as S. enterica, P. aeruginosa and K. pneumoniae with MIC values of 3.9-62.5 μg/mL.