biocommaExperiment
Extraction
Weigh 2 g of ground Beef into a 50 mL centrifuge tube. Add100 μL of 0.100 μg/mL internal standard solution. Vortex for 30s, and let stand for 10 min. Then add 15 mL of ethyl acetate.Oscillate for 10 min, and centrifuge at 9500 r/min for 5 min.Pipette the supernatant into another 50 mL centrifuge tube.Repeat the extraction with 15 mL of ethyl acetate, combinethe extract. Evaporate sample to near dryness at 40 ℃ withwater bath. Add 5 mL of 0.1 mol/L hydrochloric acid. Vortex for1 min to dissolve the residue. Add 5 mL of n-hexane. Shakefor 1 min, and centrifuge at 9500 r/min for 5 min. Discard then-hexane layer. Add 5 mL of n-hexane to the lower layer andrepeat degreasing once. The lower aqueous phase is ready forpurification.
Purification (Copure® MCX Cartridge, 60 mg/ 3 mL)
Activation: activate the Copure® MCX Cartridge by 3 mL ofmethanol, then 3 mL of 0.1 mol/L hydrochloric acid.
Loading: add prepared sample to the activated cartridge.
Washing: add 3 mL of 0.1 mol/L hydrochloric acid, then 3 mLmethanol. Drain the cartridge.
Elution: add 5 mL of 5% ammoniated methanol and collect theeluate. Evaporate to near dryness at 40°C. Dilute to 0.5 mL by0.1% formic acid aqueous solution. Vortex for 30 s to dissolvethe residue. After filter by 0.22 μm membrane, the sample isready for analysis.
Preparation of Standard Curve Solution
Accurately measure the standard solution and internal standardsolution. Dilute with 0.1% formic acid aqueous solution toprepare series of standard concentrations of 0.500 μg/L, 1.00μg/L, 2.00 μg/L, 5.00 μg/L, 10.0 μg/L, 25.0 μg/L. The internalstandard is 20.0 μg/L.
Instrumental Conditions
1.Chromatographic conditions
Instrument: Triple Quadrupole LC-MS/MS System (Triple Quad5500)
Chromatographic column: ACQUITY UPLC BEH C18, 1.7 m,2.150 mm column
Mobile phase: A: 0.1% formic acid in water, B: acetonitrile
Mobile phase gradients: initial 95% A, 95% A (0 min~1.00 min),10% A (1.00 min~3.00 min), 10% A (3.00 min~4.20 min), 95%A (4.20 min~4.50 min), 95% A (4.50 min~5.00 min)
Flow rate: 0.300 mL/min
Column temperature: 40 ℃
Injection volume: 4.0 L
2.Mass Spectrometry Conditions
Detection method: multi-reactive ion monitoring (MRM)
Table 2. Targes Characteristic Ions (*Quantifier Ion)
Results
Table 3. Nitroimidazoles in Beef at 2.50 μg/kg
Ordering Information
