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Detection of Melamine in Eggs

Application Introduction

In this paper, the Cleanert® PCX is used for pretreatment of egg samples; Venusil® ASB C8 is used for the detection of chromatographic column; the proportion of buffer salt used as mobile phase and acetonitrile is 93:7;



In this paper, the Cleanert® PCX is used for pretreatment of egg samples; Venusil® ASB C8 is used for the detection of chromatographic column; the proportion of buffer salt used as mobile phase (10mM citric acid +10mM sodium 1-octanesulfonate, pH=3.0) and acetonitrile  is 93:7; the detection wavelength is 240nm and the flow rate 1.0mL/min. The results show that this method has a recovery rate of 97%~121 and good linear relationship (R2=0.999), thus it can be widely used in the detection of melamine in eggs.


Sample preparation

Prepare freshly whipped egg sample by mechanical blending for 3 minutes. Weigh 1.0 g of the sample each time into a centrifuge tube for experiment. Spike samples with 10, 20 and 100 μL of 100 mg/L melamine stock solution, respectively, to obtain three fortified samples at concentrations of 1, 2, and 10 mg/Kg. To each sample, add 10 mL 1% trichloroacetic acid and 2 mL 5% lead acetate solutions, respectively. Mix the sample thoroughly and sonicate it for 20 minutes, and then centrifuge the sample at 8000 rpm for 10 minutes.

Blank sample is prepared with the same procedure as illustrated above without fortification.

SPE Cleanup

Activate the SPE cartridge by adding 3 mL methanol followed by 3 mL water, then discard the eluate.

Transfer supernate of the sample into the cartridge.

Wash the loaded cartridge first with 3 mL water, and then with 3 mL methanol; dry the cartridge and discard the eluate.

Add 5 mL 5% ammonium methanol (v/v) to elute the cartridge and collect the eluate.

Evaporate the collected eluate under nitrogen at 50 °C.

Reconstitute the sample with 1 mL mobile phase and filter through 0.45 μm membrane for HPLC analysis.

Results and Discussion

Figures 1 and 2 show the HPLC profiles of negative blank and spiked samples, respectively. The PCX cleanup procedure removes interference effectively and allows clear detection of melamine. The HPLC method was confirmed by 6 replicate injections of standard melamine solution at 1 and 5 mg/L, respectively. The results are summarized in Table 1, showing the consistence of the method. Calibration curve was established as shown in Table 2 and Figure 3. Recovery data of the spiked samples are shown in Table 3.

Table1 stable data of retention time and peak area ratio

Table 2 Experimental data of standard adjusted curve

Table 3 shows that the recovery ratio of melamine in the eggs is relatively good with the method


It can be seen from the above experiments that this pretreatment method is applicable to the detection of melamine in eggs due to its good purifying effect of matrix, small impurity contamination, high degree of symmetry of chromatographic peak profile, easy operation and high accuracy.

  • Cleanert SLE Products

    Cleanert SLE (Supported Liquid Extraction) plates and cartridges contain a high quality modified diatomaceous earth with an ideal surface with large specific area and low activity. Cleanert SLE plates and cartridges are used to extract analytes from bio-analytical, clinical, forensic, environmental and agrochemical samples, it even can replace most of the Liquid/Liquid extraction (LLE).

  • Venusil ASB Series Columns (C1, C8, C18 and phenyl)

    The Venusil ASB series columns are specially designed for the separation of polar compounds under low (extremely stable at pH=0.8) to medium pH condition. The stationary phase is bonded with unique bulky silanes that sterically protect the siloxane bond. We offer a line of bonding chemistry of C1, C8, C18 or Phenyl groups presenting a broad selection of different polarity for various applications.

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