Abstract A method based on dispersive solid phase extraction-high performance liquid chromatography-tandem mass spectrometry for determination of 19 kinds of nonprotein nitrogen compounds including melamine, cyromazine, amidinourea, aminotriazine, 3-aminotriazole, 4-aminotriazole, allantoin, cyanuric acid, dicyandiamide, thiourea, semicarbazide, L-leucine, L-isoleucine, L-arginine, L-hydroxyproline, L-theanine, ammeline, ammelide and guanidine in powdered formulas was presented. The nonprotein nitrogen compounds were degreased by chloroform and extracted by acetonitrile, with MgSO4 to remove water and C18 to clean up. The samples were separated on Merck ZIC HILIC column (150 mm í2. 1 mm, 5 μm, 20 nm) with gradient elution. The electrospray ionization was operated in the positive mode and negative mode, and monitored by the multiple reaction monitoring ( MRM) mode. Allation was quantified by external standard method and the other 18 kinds of nonprotein nitrogen compounds were quantified by internal standard method. All of the correlation coefficients (r) were higher than 0. 99. The limits of quantitation (LOQ) were 0. 05-5. 0 mg/kg, the average recoveries were between 82 . 2% and 115 . 0%, and the relative standard deviations were less than 20%.