ammeline and cyromazine

Milk-derived ingredients are widely used around the world in the manufacturing of nutritional products. They are prone to economically motivated adulteration with nitrogenous compounds such as melamine and its analogs in order to increase the nitrogen content of these ingredients. The need to rapidly screen milk-derived ingredients to detect adulteration is of paramount public health concern. A liquid chromatography/mass spectrometry (LC/MS)-based method using a single quadrupole mass spectrometer has been developed for the rapid frontline analysis of six nitrogenous protein adulterants, i.e. melamine, ammeline, ammelide, amidinourea, cyromazine and cyanuric acid, in three key milk-derived ingredients, i.e. whole milk powder, nonfat milk powder and whey protein concentrate.. The sample preparation scheme involves both 'dilute and shoot' as well as solid-phase extraction (SPE)-based methods. The 'dilute and shoot' scheme uses a tenfold dilution of sample with water followed by protein precipitation using 2% formic acid in acetonitrile. The SPE scheme involves tenfold dilution of sample with water, followed by protein precipitation using acetonitrile, and further cleanup through Strata Melamine SPE cartridges. Sample extracts were analyzed by hydrophilic interaction chromatography/single quadrupole mass spectrometry (HILIC/MS) in both positive and negative electrospray ionization mode. Accurate quantitation was achieved using stable isotope labeled internal standards.. A multi-day method validation study was conducted using three different milk-derived ingredients. Average accuracies, relative standard deviations (RSD) and method detection limits (MDL) for all analytes in whole milk powder were 65-118%, 7-11% and 0.9-30 mg/kg, using the 'dilute and shoot' extraction procedure. The SPE procedure results were 102-111%, 5-13%, and 0.4-2.5 mg/kg, respectively, for melamine, ammeline, ammelide and cyromazine only.. A rugged and simple to use analytical method to screen for the presence of nitrogenous economic adulterants in milk-derived ingredients has been developed for routine frontline use. Copyright © 2016 John Wiley & Sons, Ltd.

Topics: Animals; Chromatography, High Pressure Liquid; Food Analysis; Food Contamination; Limit of Detection; Milk; Milk Proteins; Nitrogen Compounds; Solid Phase Extraction; Spectrometry, Mass, Electrospray Ionization; Triazines; Urea

A novel molecularly imprinted solid-phase extraction-ultra-performance liquid chromatography (MISPE-UPLC) method for effective separation and simultaneous determination of cyromazine, melamine, and their metabolites (ammeline and ammelide) in milk samples was developed. Molecularly imprinted polymers (MIP) were synthesized in an ethanol-water system, with melamine as the template and methacrylic acid as the organic functional monomer. The MIP were applied as a specific sorbent for the selective solid phase extraction of cyromazine, ammelide, melamine and ammeline. The molecular recognition mechanism was investigated by molecular simulation and the experiment was validate by using Fourier transform infrared spectroscopy and (1)H nuclear magnetic resonance spectroscopy. A new mechanism based on the formation of both an amido group and hydrogen bonds was developed. A binding study demonstrated that the MIP showed excellent affinity to and high selectivity for melamine and related compounds. Under optimized conditions, we achieved good linearity of the calibration curves with correlation coefficients >0.999. Low limits of quantification (LOQ) for the method were determined to be 1.25, 1.25, 2.59, and 6.42 µg/kg for cyromazine, ammelide, melamine, and ammeline, respectively, which were 3 orders of magnitude smaller than the maximum residue limit (MRL). The high sensitivity of this method allows detection at the microgram per kilogram level. The proposed MISPE-UPLC method is a highly selective and sensitive method for determination of cyromazine, melamine, and their metabolites (ammeline and ammelide) for use in the control and quality assurance of milk.

Topics: Animals; Chromatography, High Pressure Liquid; Limit of Detection; Magnetic Resonance Spectroscopy; Methacrylates; Milk; Molecular Imprinting; Polymers; Solid Phase Extraction; Triazines