Lignocellulosic fruit peel waste represents an abundant, carbon-neutral feedstock for green biohydrogen production via dark fermentation, yet its rigid compact structure and high cellulose crystallinity severely restrict saccharification and fermentative hydrogen yield. In this study, a hydrothermal acidification pretreatment strategy was proposed to boost dark fermentative biohydrogen generation from navel orange peel waste, and systematic investigations were conducted to reveal the regulating mechanisms of key pretreatment parameters (hydrochloric acid concentration, pretreatment temperature, duration) on reducing sugar release and hydrogen-producing performance. Multiscale characterizations including SEM, XRD, FTIR, and TG were integrated to unravel the microstructural and chemical compositional evolution of raw and pretreated substrates. The results demonstrated that hydrothermal acidification effectively disrupted the dense lignocellulosic network of navel orange peel, lowered cellulose crystallinity, and greatly improved substrate accessibility for hydrolytic reactions and microbial adhesion. Under the optimal pretreatment condition (1.0 mol/L HCl, 120 °C, 1 h), the concentration of released reducing sugars reached 10.2 g/L, which was 67.2% higher than that of untreated raw peel. The corresponding maximum cumulative hydrogen yield attained 36.5 mL H₂/g TS, representing a 67.4% improvement relative to the untreated control group. Pearson correlation analysis verified that pretreatment temperature, acid concentration, and duration exhibited strong positive correlations with hemicellulose and cellulose removal efficiencies, while excessive pretreatment (HCl > 1.0 mol/L, temperature > 120 °C, duration > 1 h) generated inhibitory by-products that suppressed microbial hydrogen evolution. This study comprehensively clarifies the structural modification and biohydrogen promotion mechanism of hydrothermal acidification pretreatment on pectin-rich biomass, and delivers a cost-effective, facile technical route for high-value energy valorization and harmless disposal of fruit processing solid wastes.