Numerous studies have demonstrated that Chlorella pyrenoidosa exerts potent anti-diabetic effects, yet its underlying mechanism remains elusive. Notably, no research has explored the anti-diabetic efficacy of its fermented product to date. In this study, we developed a novel compound probiotic-fermented C. pyrenoidosa product (CH-F), and systematically evaluated its anti-diabetic efficacy and mechanism. Furthermore, untargeted metabolomics analysis revealed that fermentation significantly altered the metabolite profile of C. pyrenoidosa, leading to the enrichment of potential anti-diabetic compounds such as acetylcholine and citicoline. In streptozotocin-induced T2DM mice, CH-F effectively ameliorated hyperglycemia, dyslipidemia, and hepatic/pancreatic histological injuries, showing markedly superior efficacy compared to unfermented C. pyrenoidosa. Additionally, CH-F modulated the gut microbiota and improved the levels of short-chain fatty acids (SCFAs) in the intestines of T2DM mice. We demonstrate that CH-F exerts anti-diabetic effects via gut microbiota modulation and SCFA regulation, highlighting its great potential as a functional food ingredient and dietary supplement for the adjuvant management of T2DM.