Simple Method for Monitoring Melamine in Food by Ion Pair Chromatography

A simple, rapid, precise and accurate, ion-pair reversed-phase high-performance liquid chromatography (HPLC) method is developed to determine melamine in a wide range of food matrices, including several milk-based products. The sample is prepared by protein precipitation with 6M hydrochloric and methanol (1:9). As melamine is present in supernatant in the cationic form, the chromatographic separation in reversed-phase LC requires the use of anionic ion pair reagent, such as 1-Hexanesulfonic acid sodium. This allows a satisfactory chromatographic retention and peak shape in all type of food samples investigated. The separation is performed on a Vetical VertiSepᵀᴹ pH endure C18 column (4.6 id x 250 mm, 5 µm particle size) at room temperature. The mobile phase (0.01 M 1-Hexanesulfonic acid sodium in 50 mM NaH2PO4) is pump at a flow rate of 1.0 ml/min with detection at 215 nm. Melamine elutes at 5.4 min. A linear response (r = 0.999) is observed for sample ranging from 5-20 µg/mL. The method provides recoveries of 97.8-100.7% (mean=99.4) with the RSDs (n=5) of 3.34-3.82% in the concentration 5 - 20 µg/mL. The limit of detection (LOD) and limit of quantitation values is 0.16 and 0.53 µg/mL(s/n=5) respectively.