Yibar, Artun, et al. Kafkas Üniversitesi Veteriner Fakültesi Dergisi, 2013, 19(6), 1023-1028.
To see if boiling reduced the nitrofuran 3-amino-2-oxazolidinone (AOZ) residues in eggs, nitrofuran AOZ residues in raw and boiled eggs were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The nitrofuran AOZ, a protein-bound side-chain metabolite, levels in eggs actually increased after they were boiled (perhaps due to higher extraction efficiency in boiled eggs).
Sample preparation
· Scrambled raw eggs (liquid) and 2 ± 0.03 g volumes were added to 50 mL centrifuge tubes. The other egg parts were boiled for 10 minutes in 100°C water and placed in an ice bath. It was also done for another batch of boiled eggs (solid). 100 µL of 2-NP-AOZ-D4 internal standard, 5 mL 0.1 M HCl were added to each sample, then vortexed and homogenised. Added 300 µL of 50 mM 2-nitrobenzaldehyde in DMSO and stirred for 1 minute. Then the samples were shaker-incubated at 37°C for 16 hours at 50 rpm.
· Samples were brought to room temperature after incubation and neutralised using 700 µL of 1 M NaOH/0.1 M K2HPO4 + 3H2O (2:1, v/v). Add 5 mL of ethyl acetate and shake for 10 minutes at room temperature. In order not to make an emulsion, 3 mL of n-hexane was added. It was centrifuged and 6 mL of the organic upper phase was added to a 15 mL glass tube and dried in nitrogen under 5.0 psi pressure at 42°C. Then wetted with 1 mL of n-hexane, vortexed and added 750 µL of MeOH/deionised water (1:4, v/v). The result was again vortexed and centrifuged. The lower aqueous phase was collected, the 0.45 µm filter was used, and 50 µL of the filtrate was transferred to a microvial for LC-MS/MS analysis.
