Exogenous 2-Keto-L-Gulonic Acid Supplementation Promotes L-Ascorbic Acid Biosynthesis in Drosophila melanogaster

L-ascorbic acid (ASA) is an essential micronutrient critical for antioxidant defense and metabolic regulation in animals. Unlike many vertebrates, <i>Drosophila melanogaster</i> possesses the ability to synthesize ASA endogenously, yet the regulatory mechanisms governing this biosynthesis remain unclear. 2-keto-L-gulonic acid (2KGA), a key precursor in industrial vitamin C production, has been shown to enhance ASA accumulation in plants, but its role in invertebrates is unknown. This study systematically investigated the effect of exogenous 2KGA supplementation on ASA biosynthesis in <i>Drosophila</i>. Fruit flies were reared on media with or without 2KGA (1.6 g/L; <i>n</i> = 30 per group) for 12 days, followed by ASA quantification via high-performance liquid chromatography (HPLC). Results showed that 2KGA treatment increased mean ASA content from 0.00853 ± 0.0012 to 0.01064 ± 0.0015 μg/fly (24.74% increase; <i>p</i> = 0.0194, <i>r</i>² = 0.558) compared to the control group, indicating that 2KGA acts as a metabolic regulator to promote ASA biosynthesis. Sex-separated analyses further revealed that this effect was primarily driven by male flies (<i>p</i> = 0.0057, <i>r</i>² = 0.879), whereas females showed no significant response (<i>p</i> = 0.1783), pointing to a sex-dependent regulation of 2KGA-mediated ASA biosynthesis. These findings provide the first evidence that 2KGA modulates ASA levels in an invertebrate disease model and suggest that fruit flies can serve as a useful platform to explore conserved redox-regulatory mechanisms relevant to human health and disease.