Abstract: Objective To develop a rapid and accurate gas chromatography-mass spectrometry(GC-MS) method for the determination of nitrite in urine.Methods Through optimizing operation parameters as temperature, time, and pH of derivatization, we developed a 2, 3-diaminonaphthalene derivatization-GC-MS method for the determination of nitrite in urine.Results This method was sensitive, reproducible and accurate.The equation for the regression line was y=0.2071x+0.006(R²=0.999);the linear range was 0.01-20 μg/mL;the limit of detection(LOD) was 0.001 μg/mL;the intra-day relative standard deviation(%RSD) was 3.19-4.06% and the inter-day%RSD was 3.92-5.16%;the recovery was 90.31%-113.5%.Conclusion The method has lower detection limit, better recovery, higher precision, and less time-consuming performance and could be applied in forensic and preventive medicine analysis.