Abstract: Background This study focused on exploring how biochar application with varying doses and frequencies regulates soil microbial necromass carbon (MNC) in the saline-alkali farmlands of the Yellow River Delta, a region critical for China’s agricultural carbon sequestration and food security. Given the lack of systematic research on the “dose-frequency” coupling effect of biochar on MNC in coastal saline-alkali soils, a four-year field experiment was conducted at the Dongying Base of Shandong Academy of Agricultural Sciences. Methods Ten treatments were designed, including a control (C0, no biochar), annual biochar application at 4 (C1), 8 (C2), and 12 t/ha (C3), biennial application at 4 (C4), 8 (C5), and 12 t/ha (C6), and one-time application at 4 (C7), 8 (C8), and 12 t/ha (C9). The biochar used was pyrolyzed from rice straw at 800°C, with key properties: 73% carbon content, pH 8.35, and 0.32 mm particle size. The experiment analyzed the effects of these treatments on soil organic carbon (SOC), MNC (including fungal necromass carbon, FNC, and bacterial necromass carbon, BNC), and key physicochemical properties (dissolved organic carbon, DOC; total nitrogen, TN; total phosphorus, TP; etc.). Results Results showed that all biochar treatments except C3 (annual 12 t/ha) and C7 (one-time 4 t/ha) significantly increased total soil amino sugar carbon—an important marker of microbial residue accumulation—by 15.73%-44.30%. Notably, only the annual low-to-medium dose treatments (C1: 4 t/ha, C2: 8 t/ha) achieved a significant MNC increase of 26.26%-30.82%, with FNC identified as the primary driver. This was attributed to biochar’s porous structure (2-10 μm pore size, matching fungal hyphal diameter) and aromatic carbon supply, which reduced hyphal disturbance and provided decomposable substrates for fungi, while high-dose or one-time treatments disrupted microbial metabolism via rapid adsorption saturation or long-term carbon source depletion. Correlation analysis revealed distinct interactions: SOC showed a significant negative correlation with BNC (r = -0.59), likely due to the alkaline soil environment (pH 8.47-8.55) inhibiting bacterial activity; in contrast, FNC was positively correlated with DOC, TN, and TP (r = 0.44-0.50), indicating biochar-facilitated nutrient microdomains promoted fungal proliferation. All biochar treatments improved soil nutrients (e.g., TN increased by 12.50%-33.33%, TP by 9.91%-77.48%), but only C1, C4, and C5 enhanced DOC (33.71%-80.74%), as high-temperature biochar in other treatments reached DOC adsorption-desorption equilibrium. Conclusions In conclusion, annual application of low-dose biochar (4-8 t/ha) is the optimal strategy for sustained MNC accumulation in the Yellow River Delta’s saline-alkali farmlands. Biochar exerts a stronger promoting effect on fungus-derived carbon than bacteria-derived carbon, reshaping the microbial residue carbon pool and strengthening soil carbon sequestration. This study provides theoretical support for formulating targeted farmland carbon sink enhancement strategies in the region. In order to further clarify the long-term regulatory mechanism of biochar on the soil carbon pool, future research should design long-term field-based controlled experiments, combined with high-throughput sequencing and 13C isotope tracing techniques, to ultimately clarify the "application mode-microorganisms-carbon pool" coupled regulatory mechanism, and provide more systematic support for enhancing soil carbon sequestration in the Yellow River Delta.