Simultaneous Determination of 5 Nitrogen-Rich Adulterants in Milk and Powdered Milk by LC-MS/MS

A method of liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed for the determination of five nitrogen-rich adulterants (dicyclanil, dicyandiamide, biuret, cyromazine, melamine) in milk and powdered milk samples. These compounds were potential to be used in economic adulteration to enhance the nitrogen content in milk products. The protein of sample was precipitated, and the extracts were dissolved and distilled with acetonitrile. The supernatant solution was extracted with n-hexane to remove the fat, and then cleaned up with SPE HLB cartridges. The quantitative detection was performed on LC-MS/MS by multiple reaction monitoring (MRM) mode under positive electrospray ionization (ESI⁺). Isotopes dilution internal standard method (melamine) or external standard method (dicyclanil, dicyandiamide, biuret and cyromazine) was used to determine the residue contents in samples. The limits of quantification (LOQs, S/N=10) are dicyclanil (0.5 μg/kg), dicyandiamide (2.5 μg/kg), biuret (20 μg/kg), cyromazine (5 μg/kg), melamine (15 μg/kg) for milk samples and dicyclanil (8 μg/kg), dicyandiamide (35 μg/kg), biuret (50 μg/kg), cyromazine (40 μg/kg), melamine (50 μg/kg) for powered milk samples. Single-laboratory method validation data are determined, the calibration standard curves concentration are 0, 50, 100, 200, 400 μg/kg and the correlation coefficients are more than 0.991. The average recovery is 72.4%-102.8%, and the relative standard deviation (RSD) is 1.3%-12.3%. The residues of 5 compounds in 55 batches of milk and milk powder samples are determined by LC-MS/MS, and the results are consistent with the national standard method (GB/T 22388—2008). This method is suitable for determination and confirmation of these compounds and provides a convenient, rapid and effective approach to proactively combat economically motivated adulteration in milk and powdered milk sample.