An accurate method for the determination of low concentrations of nitrate in high-saline seawater is now well optimized and validated. The method is based on using reversed-phase ion-pair liquid chromatography with diode-array detection for accurate determination of nitrate in high-saline seawater without interference from highly dominant chloride and sulfate anions. The method showed satisfactory precision (ranging from 5.8% to 9.5%), linearity, and detection limits, and its trueness was confirmed by participation in the international comparisons of CCQM K-161 with seven national metrology institutes. The key comparison and spiking showed recovery rates between 90% and 110%. The validation results confirm the applicability of the optimized method for detecting very low concentrations of nitrate in high-saline seawater.