This paper presents a method of reversed-phase high performance liquid chromatography to determine N,N-methylene-bi-(2-amino-5-mercapto-1,3,4-thiodiazole) (MBAMT) and its decomposition product 2-amino-5-mercapto-1,3,4-thiodiazole (AMT). Recoveries of MBAMT and AMT were 91% and 94% respectively. The detectable limits of them in water samples were 7.19 nmol/L for MBAMT and 2.87 nmol/L for AMT. The resolution between the peaks of MBAMT and AMT was 2.5. The column efficiency and the sensitivity of this method were significantly better in comparison with normal phase chromatography.