Groundwater nitrate contamination is challenging and requires efficient solutions for nitrate removal. This study aims to investigate nitrate removal using a novel adsorbent, biochar-supported aluminum-substituted goethite (BAG). The results showed that an increase in the initial Al/(Al + Fe) atomic ratio for BAGs from 0 to 20% decreased the specific surface area from 115.2 to 75.7 m²/g, but enhanced the surface charge density from 0.0180 to 0.0843 C/m². By comparison, 10% of Al/(Al + Fe) led to the optimal adsorbent for nitrate removal. The adsorbent’s adsorption capacity was effective with a wide pH range (4–8), and decreased with increasing ionic strength. The descending order of nitrate adsorption inhibition by co-existing anions was SO₄²⁻, HCO₃⁻, PO₄³⁻, and Cl⁻. The adsorption kinetics and isotherms agreed well with the pseudo-first-order equation and Langmuir model, respectively. The theoretical maximum adsorption capacity was 96.1469 mg/g. Thermodynamic analysis showed that the nitrate adsorption was spontaneous and endothermic. After 10-cycle regeneration, the BAG still kept 92.6% of its original adsorption capacity for synthetic nitrate-contaminated groundwater. Moreover, the main adsorption mechanism was attributed to electrostatic attraction due to the enhancement of surface charge density by Al substitution. Accordingly, the BAG adsorbent is a potential solution to remove nitrate from groundwater.