Apoptosis-Mediated Cytotoxicity of Costus Comosus–Derived Silver Nanoparticles in Hepatocellular Carcinoma Cells

Abstract

Objective: To synthesize silver nanoparticles using Costus comosus (CC-AgNPs) leaf extract and to evaluate their in vitro antioxidant, in vitro cytotoxic, and apoptosis-inducing effects in HepG2 hepatocellular carcinoma cells. Methods: Silver nanoparticles were synthesized using Costus comosus leaf extract and characterized by UV–spectroscopy, ATR-FTIR, SEM–EDX, HR-TEM, and SAED. Antioxidant activity was assessed using DPPH and FRAP assays. Cytotoxicity was evaluated by the MTT assay. Apoptotic mechanisms were investigated through caspase-3 activity analysis and ELISA-based quantification of apoptotic markers. Results: The aqueous extract of Costus comosus was employed for silver nanoparticle synthesis and confirmed by a characteristic surface plasmon resonance peak at 445 nm. ATR-FTIR spectra revealed prominent bands corresponding to –OH, –NH, and C=O functional groups, indicating the involvement of phenolics and proteins in reduction and stabilization. SEM and TEM analyses demonstrated predominantly spherical, well-dispersed nanoparticles with sizes ranging from 5–12 nm. EDAX analysis showed a strong silver signal around 3 keV, confirming elemental silver along with minor signals from capping biomolecules. SAED patterns exhibited distinct concentric rings, verifying the crystalline nature of the nanoparticles. CC-AgNPs exhibited higher total phenolic (58.62±0.48 mg GAE/g) and flavonoid (19.26±0.74 mg QE/g) contents compared with the crude extract. Enhanced antioxidant activity was observed, with 91.16% DPPH inhibition and FRAP value of 784.21±16.72 µmolFe(II)/g. MTT analysis showed a concentration-dependent reduction in HepG2 cell viability with an IC₅₀ of 18.64±1.84 µg/ml for CC-AgNPs, lower than the crude extract (25.66±2.13 µg/ml). CC-AgNPs treatment significantly increased caspase-3 activity (up to 5.1-fold) and elevated pro-apoptotic markers, including Bax, cytochrome c, and p53, while Bcl-2 expression decreased, indicating activation of the intrinsic mitochondrial apoptotic pathway. The apoptotic response was comparable to the standard drug doxorubicin. Conclusion: Green-synthesized CC-AgNPs exhibit potent antioxidant and apoptosis-mediated anticancer activity against HepG2 cells, highlighting their promise as an eco-friendly nanotherapeutic candidate for hepatocellular carcinoma.