The medicinal fungus Cordyceps militaris is recognized for producing cordycepin, a bioactive nucleoside with anticancer, immunomodulatory, and antioxidant properties. However, conventional culture media often entail high production costs and limited sustainability, prompting the search for alternative nutrient sources. This study evaluated onion, green onion, and garlic peel extracts—agricultural by-products rich in flavonoids, phenolics, and sulfur-containing antioxidants—as sustainable substrates for enhancing mycelial biomass and cordycepin biosynthesis in C. militaris. Liquid cultures supplemented with peel extracts (1–5%) were assessed for growth, cordycepin production (HPLC), and antioxidant activity (DPPH assay). Onion peel extract (OPE) showed the strongest growth-promoting effect, yielding 8.2 g/L of biomass at 5% and achieving a 19% increase in cordycepin concentration at 3% compared with the control. Antioxidant activity strongly correlated with cordycepin accumulation (R = 0.96, p < 0.001), indicating that secondary metabolite production contributed significantly to radicalscavenging capacity. Response surface methodology using a Box–Behnken design revealed that extract concentration, pH, and incubation period significantly influenced cordycepin production (p < 0.05), with the quadratic model showing excellent fit (R² = 0.9924). Optimal conditions were identified as 3% extract concentration, pH 6.0, and 12 days of incubation, under which cordycepin reached 0.995 mg/L, substantially higher than the control (0.693 mg/L). These findings demonstrate that agricultural by-product extracts, particularly onion peel, can serve as effective and economical substrates for enhancing cordycepin biosynthesis while supporting sustainable bioprocessing strategies in C. militaris cultivation.

ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
    Mycelial culture preparation
    Preparation of liquid culture media
    Cultivation of C. militaris
    Biomass yield determination
    Determination of cordycepin content
    Antioxidant activity measurement
    Optimization using response surface methodology(RSM)
    Statistical analysis
RESULTS AND DISCUSSION
    Mycelial biomass yield of C. militaris in differentsubstrates
    Quantitative analysis of cordycepin in C. militaris
    Assessment of antioxidant properties in C. militarisculture broth
    Optimization of culture conditions using responsesurface methodology
CONCLUSION
ACKNOWLEDGEMENTS
REFERENCES

• Si Young Ha(Department of Environmental Materials Science, Institute of Agriculture & Life Science, Gyeongsang National University, Jinju 52828, Republic of Korea)
• Woo Seok Lim(Department of Environmental Materials Science, Institute of Agriculture & Life Science, Gyeongsang National University, Jinju 52828, Republic of Korea)
• Hyeon Cheol Kim(Department of Environmental Materials Science, Institute of Agriculture & Life Science, Gyeongsang National University, Jinju 52828, Republic of Korea)
• Jae-Kyung Yang(Department of Environmental Materials Science, Institute of Agriculture & Life Science, Gyeongsang National University, Jinju 52828, Republic of Korea) Corresponding author