Demand for natural red colorants has increased due to regulatory and consumer pressure to replace synthetic additives; however, their industrial application remains limited by challenges related to production, stability, and safety. In this study, the production, physicochemical stability, antimicrobial activity, and preliminary cytotoxicity of an azaphilone-type red colorant produced by Talaromyces sp. C1I3 under submerged fermentation were evaluated. Two sequential Box–Behnken designs identified optimal production conditions at approximately pH 5.8, 60 rpm, and 15.15 g·L⁻¹ monosodium glutamate. Under these conditions, colorant production increased from 1.46 AU_500nm on day 5 to 9.9 AU_500nm on day 10, with estimated kinetic parameters of µmax 0.0109 h⁻¹ and Yx/s 0.63 g·g⁻¹. The colorant showed high thermal stability, retaining approximately 95% of its color after heating at 80 °C for 10 min, and remained stable over a broad pH range, with only 18% degradation after 168 h of light exposure. No significant antimicrobial activity was detected, indicating that the colorant primarily functions as a color additive without additional bioactive effects. Furthermore, no acute cytotoxicity was observed in human neonatal dermal fibroblasts at concentrations ranging from 0.5 to 15 µg·mL⁻¹. These findings demonstrate the potential of this fungal colorant as a stable and safe natural colorant for future industrial applications.