A strain with potential hypoglycemic effects was screened from three Cordyceps-associated fungi, namely Penicillium jiangxiense (JX), Ophiocordyceps xuefengensis (XF), and Ophiocordyceps ramosissimum (RS), based on in vitro antioxidant capacity and α-glucosidase inhibitory activity. The in vitro antioxidant and hypoglycemic activities of the three strains were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging assay, total antioxidant capacity measurement, and α-glucosidase inhibition experiment. The results show that JX exhibits the optimal activity in all the above assays. Furthermore, the chemical constituents of JX were analyzed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), and a total of 197 compounds were identified, including L-carnitine, N-feruloyltyramine, and syringic acid. The potential mechanisms underlying the therapeutic effects of the active components of JX on type 2 diabetes mellitus (T2DM) were investigated via network pharmacology and molecular docking techniques. An insulin-resistant HepG2 (IR-HepG2) cell model was established to analyze the hypoglycemic function of JX by determining glucose consumption and glycogen content. The Western Blot analysis was performed to detect the expression levels of PI3K, AKT, p-AKT, and TNF-α proteins. The results of network pharmacology reveals that the active components of JX upregulated the expression or phosphorylation levels of molecules, including EGFR, PIK3CA/PIK3CB/PIK3CD, and AKT1/2/3 proteins, while critical regulatory targets such as TNF, PRKCA, and MTOR are activated, thereby mediating the activation of the PI3K-Akt, Ras, and cyclic adenosine monophosphate (cAMP) signaling pathways. The results of cellular experiments demonstrates that glucose consumption and glycogen content in HepG2 cells are significantly increased by JX treatment compared with model group (P < 0.001, P < 0.05). meantime, the expression of PI3K, AKT, and p-AKT proteins is upregulated, whereas the expression of TNF-α protein is downregulated. In conclusion, it is proposed that the active components of JX may intervene in the pathogenesis of T2DM by synergistically regulating glucose metabolism, lipid metabolism, and oxidative stress processes through multi-target and multi-pathway interactions.