Melamine contamination in food became an issue in recent years after the discovery of it, and related compounds, in pet food and baby formula. It was discovered that melamine was intentionally added to inflate nitrogen content, often the sole measure of the amount of protein in these products. The tainted food led to numerous illnesses, several fatalities, and massive product recalls. Currently, imported raw materials, namely wheat gluten and rice protein used to make these foods, as well as the actual consumer-ready foods, may undergo testing to ensure the absence of these compounds. We have detailed preparation and analytical procedures for these adulterants, using HPLC-MSMS instrumentation in previous publications (1,2). In this article, we focus on the analysis of melamine and related compounds with the use of more economical gas chromatography-mass spectrometry (GC-MS) instrumentation.
The United States Food and Drug Administration (US FDA) adopted a screening method in October 2008 for the GC-MS analysis of melamine and related compounds in a variety of matrices (3). Per the method, 0.5 g of the sample is mixed thoroughly with 20 mL of an extraction solvent mixture (10:40:50 diethylamine:water:acetonitrile). Following sonication (30 minutes) and centrifugation (10 minutes), an aliquot is filtered and evaporated to dryness. Sylon™ BFT and pyridine are then added along with an internal standard. The extract is then incubated (70 °C for 45 minutes) so that trimethylsilyl (TMS) derivatives of each analyte are formed. The resulting derivatized extract is then analyzed by GC-MS. The method allows the operation of the MS in the scan mode (m/z from 50-450 amu) or the selected ion monitoring (SIM) mode. Table 1 shows the structures of the four analytes (melamine, ammeline, ammelide, and cyanuric acid) plus 2,6-diamino-4-chloropyrimidine, the internal standard (I.S.) specified by the method.
Table 1.Melamine and Related Compounds
For this work, we chose a common dry dog food obtained from a local grocery store. The following samples were prepared:
All standards and extracts were analyzed with the MS operating in the scan mode, and again later with the MS operating in the SIM mode.
The following results were observed:
Figure 1.100 ng/mL Calibration Standard (SIM Mode) (referenced above is 28471-U)
Figure 2.Laboratory Blank (SIM Mode)
Figure 3.Dog Food Sample (SIM Mode)
Figure 4.Spiked Dog Food Sample (SIM Mode)
Our observation is that the method is very easy to perform and provides good sensitivity. In particular, the use of the specified extraction solvent mixture was found to be very effective in solubilizing and extracting all target analytes. Additionally, the formation of TMS derivatives allows these analytes to be analyzed by GC, with symmetrical peak shapes, high signal-to-noise ratios, and low detection levels.