Biogenic nanoparticle-mediated enhancement of antioxidant responses and secondary metabolites in Capsicum annuum callus cultures

Capsaicin, a pharmacologically and nutraceutically important alkaloid, faces production challenges due to limitations in
conventional extraction and chemical synthesis. In this study, callus cultures of Capsicum annuum L. were established
from cotyledonary explants on Murashige and Skoog (MS) medium supplemented with 2 mg/L 2,4-D and 0.5 mg/L kinetin.
The friable calli exhibited sigmoidal growth, with biomass peaking at 3 g FW/L on day 28. Interestingly, maximum
capsaicinoid accumulation (65.63 mg/100 g FW) occurred earlier, on day 21, highlighting a temporal separation between
growth and secondary metabolism. Nanoparticles (Ag, AgN, and RAg) were evaluated as abiotic elicitors to enhance
metabolite production. AgNPs at 5 ppm significantly increased biomass, while RAg at 5 ppm produced the highest capsaicinoid
yield. Chlorophyll and carotenoid contents were also positively influenced, with Ag 10 ppm promoting the highest
chlorophyll levels and RAg supporting greater carotenoid accumulation. In addition, nanoparticle treatments enhanced
total phenolic content (TPC), total flavonoid content (TFC), and antioxidant capacity. RAg elicited a 124% increase in
TPC, while AgN 20 ppm induced the highest flavonoid accumulation. Antioxidant assays (TAA, FRAP, and DPPH) further
confirmed that nanoparticle elicitation stimulated redox-related defense pathways, though responses varied with nanoparticle
type and concentration. Overall, the results demonstrate that nanoparticle-mediated elicitation effectively enhances
biomass, capsaicinoids, pigments, and antioxidant metabolites in Capsicum callus cultures. This approach represents a
scalable and sustainable strategy for producing high-value secondary metabolites, with potential applications in the food,
pharmaceutical, and nutraceutical industries.