Mechanistic investigation of UV-irradiated neem oil against microbial biofilms and A431 skin cancer cells via Bax/Bcl2/caspase-3 modulation

Abstract

Neem oil (Azadirachta indica) possesses antimicrobial and anticancer properties, but its clinical translation is limited by variable chemical profile and suboptimal bioavailability. Ultraviolet (UV) irradiation is a green, non-thermal physical modification technique that can alter phytochemical composition and bioactivity. The study aimed to characterize chemical changes induced by UV irradiation (254 nm, 120 min) in neem oil and evaluate the enhanced antimicrobial, antibiofilm, and anticancer activities against A431 skin cancer cells, including mechanistic insights into cell cycle arrest and Bax/Bcl-2/caspase-3 signaling. Crude and UV-irradiated neem oils were analyzed by Fourier transform infrared (FTIR) spectroscopy and Gas chromatography–mass spectrometry (GC–MS). Antimicrobial and antibiofilm activities were tested against five pathogens including Methicillin-resistant Staphylococcus aureus (MRSA), Bacillus subtilis, Escherichia coli, Proteus vulgaris and Candida albicans. Antiproliferative activity against A431 cells was assessed by MTT assay, cell cycle analysis (flow cytometry), and qRT-PCR for Bcl-2, Bax, and caspase-3. UV irradiation induced oxidation, cis-to-trans isomerization, and formation of α,β-unsaturated carbonyls (FTIR), along with new bioactive compounds: isochiapin B (3.88%) and a steroid derivative (9.40%) by GC–MS. UV-neem oil significantly enhanced antimicrobial activity (inhibition zones up to 30 mm), reduced MIC/MBC up to eightfold (MRSA, MIC: 125 → 15.6 µg/mL), and showed superior biofilm inhibition (91–95% at 25% MBC). UV-neem oil exhibited a lower IC₅₀ (154.4 vs. 179.3 µg/mL), induced stronger S-phase arrest (S-phase reduced to 9.45%), upregulated Bax (5.26-fold) and caspase-3 (7.65-fold), and downregulated Bcl-2 (0.53-fold). UV irradiation transforms neem oil into a more potent antimicrobial, antibiofilm, and pro-apoptotic agent against A431 cells via oxidative and isomeric modifications. This green photochemical approach offers a promising strategy to enhance the therapeutic efficacy of plant oils.

Graphical abstract

Keywords

Neem oil / UV irradiation / Antimicrobial / Antibiofilm / A431 cells / Apoptosis / Cell cycle / Caspase-3

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Funding

Princess Nourah Bint Abdulrahman University((PNURSP2026R217)

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