Fractionation-Guided Evaluation of Justicia secunda and Kalanchoe pinnata as Inhibitors of DNA Gyrase and Penicillin-Binding Proteins in Multidrug-Resistant Acinetobacter baumannii and Klebsiella pneumoniae

Abstract

Background: The increasing prevalence of multidrug-resistant (MDR) bacterial pathogens necessitates alternative antimicrobial agents; thus, this study combined fractionation-guided in vitro evaluation with in silico target-based analysis to assess the antibacterial potential and possible mechanisms of action of Justicia secunda and Kalanchoe pinnata against MDR Acinetobacter baumannii and Klebsiella pneumoniae. Methods: Crude extracts of J. secunda and K. pinnata were prepared by maceration and subsequently fractionated. Phytochemical profiling was performed using high-performance liquid chromatography (HPLC). Antibacterial activity against MDR A. baumannii and K. pneumoniae was evaluated using agar well diffusion, while minimum inhibitory concentrations (MICs) were determined by broth dilution. Molecular docking with MM/GBSA refinement assessed interactions of identified compounds with DNA gyrase (A. baumannii) and penicillin-binding protein (K. pneumoniae). ADMET and drug-likeness properties were predicted in silico. Results: HPLC analysis revealed diverse phenolic acids and flavonoids in both plants, with abundant compounds including naringin, myricetin, baicalin, apigenin, catechin, and mangiferin. Fractionation enhanced antibacterial activity relative to crude extracts. In J. secunda, the ethyl acetate fraction showed the strongest activity (zones up to 27.33 mm; MIC 3.125 mg/mL). In K. pinnata, n-hexane and ethyl acetate fractions produced inhibition zones up to 27.00 mm, while the aqueous fraction exhibited the lowest MIC (3.125 mg/mL). Docking studies identified several phytochemicals with higher predicted binding affinities than standard antibiotics, alongside favorable ADMET profiles and Lipinski compliance. Conclusion: Justicia secunda and Kalanchoe pinnata contain bioactive phytochemicals with significant antibacterial activity against MDR pathogens, supporting their potential as sources of novel antimicrobial leads.