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. 2020 Feb 22;25(4):987.
doi: 10.3390/molecules25040987.

Determination of Eight Coccidiostats in Eggs by Liquid-Liquid Extraction-Solid-Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry

Bo Wang  1   2 , Jianyu Liu  2   3 , Xia Zhao  2   3 , Kaizhou Xie  2   3 , Zhixiang Diao  2   3 , Genxi Zhang  2   3 , Tao Zhang  2   3 , Guojun Dai  2   3
Affiliations

Determination of Eight Coccidiostats in Eggs by Liquid-Liquid Extraction-Solid-Phase Extraction and Liquid Chromatography-Tandem Mass Spectrometry

Bo Wang et al. Molecules. .

Abstract

A method for the simultaneous determination of robenidine, halofuginone, lasalocid, monensin, nigericin, salinomycin, narasin, and maduramicin residues in eggs by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. The sample preparation method used a combination of liquid-liquid extraction (LLE) and solid-phase extraction (SPE) technology to extract and purify these target compounds from eggs. The target compounds were separated by gradient elution using high-performance liquid chromatography (HPLC) and ultra-performance liquid chromatography (UPLC). Tandem mass spectrometry was used to quantitatively and qualitatively analyze the target compounds via electrospray ionization (ESI+) and multiple reaction monitoring mode. The HPLC-MS/MS and UPLC-MS/MS methods were validated according to the requirements defined by the European Union and the Food and Drug Administration. The limits of detection and limits of quantification of the eight coccidiostats in eggs were 0.23-0.52 µg/kg and 0.82-1.73 µg/kg for HPLC-MS/MS, and 0.16-0.42 µg/kg and 0.81-1.25 µg/kg for UPLC-MS/MS, respectively. The eggs were spiked with four concentrations of the eight coccidiostats, and the HPLC-MS/MS and UPLC-MS/MS average recoveries were all higher than 71.69% and 72.26%, respectively. Compared with the HPLC-MS/MS method, utilizing UPLC-MS/MS had the advantages of low reagent consumption, a short detection time, and high recovery and precision. Finally, the HPLC-MS/MS and UPLC-MS/MS methods were successfully applied to detect eight coccidiostats in 40 eggs.

Keywords: HPLC–MS/MS; LLE; SPE; UPLC–MS/MS; coccidiostats; eggs.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Comparison of the recovery of eight coccidiostats in eggs via different solid-phase extraction cartridges. Note: 1. CNW C18 cartridge; 2. SIMON C18 cartridge; 3. BOSHI C18 cartridge.
Figure 2
Figure 2
HPLC–MS/MS total ion chromatograms (TICs) and extracted ion chromatograms (XICs) (a–h: robenidine (ROB), halofuginone (HAF), lasalocid (LAS), monensin (MON), nigericin (NIG), salinomycin (SAL), narasin (NAR), and maduramicin (MAD)) of eight coccidiostats in a blank egg sample. Note: “Int ...” is represented as "Intensity, cps" in the figure, the following is the same.
Figure 3
Figure 3
UPLC–MS/MS TICs and XICs (a–h: ROB, HAF, LAS, MON, NIG, SAL, NAR, and MAD) of eight coccidiostats in a blank egg sample.
Figure 4
Figure 4
HPLC–MS/MS TICs and XICs (a–h: ROB, HAF, LAS, MON, NIG, SAL, NAR, and MAD) of a blank egg sample spiked with ROB and HAF at 25.0 μg/kg, MON at 37.5 μg/kg, MAD at 60.0 μg/kg, and LAS, NIG, SAL, and NAR at 15.0 μg/kg.
Figure 5
Figure 5
UPLC–MS/MS TICs and XICs (a–h: ROB, HAF, LAS, MON, NIG, SAL, NAR, and MAD) of a blank egg sample spiked with ROB and HAF at 25.0 μg/kg, MON at 37.5 μg/kg, MAD at 60.0 μg/kg, and LAS, NIG, SAL, and NAR at 15.0 μg/kg.
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