Microbial-derived carbon plays a crucial role in mitigating climate change by forming stable carbon components through the soil microbial carbon pump. However, related studies have ignored the contribution of extracellular polymeric substances (EPS) as microbial extracellular metabolites to soil organic carbon (SOC), particularly in deeper soils. This study explored the distribution of EPS in six typical soil profiles (0-120 cm) from two parent materials (limestone and shale) and three land use types (dryland, woodland, and paddy land). The contribution of microbial biomass carbon (MBC) to SOC was significantly higher than that of EPS-carbon (EPS-C) in surface soils (0-40 cm), while EPS-C constituted a larger proportion in deeper soils (80-120 cm). The EPS accumulation efficiency (EPS-protein/MBC and EPS-polysaccharide/MBC) gradually increased with soil depth. This accumulation was strongly correlated with the abundance of g_Zixibacteria, g_Zavarzinella, g_Xylohypha, g_Xanthothecium, and g_Xanthagaricus. Data analysis revealed that beta-glucosidase (BG) activity and total nitrogen (TN) content had significant negative effects on the EPS/SOC ratio. Additionally, extracellular enzyme analyses confirmed that low nitrogen availability in deeper soils enhanced the EPS accumulation efficiency, thereby increasing the EPS-C/ SOC ratio along the soil profile. Overall, this study provides new insights into the composition of deep soil carbon pools and highlights the important role of EPS in deep soil carbon storage.