Determination of Anthelmintic Drug Residues in Milk Using Ultra High Performance Liquid Chromatography–Tandem Mass Spectrometry
A modified QuEChERS-type extraction method is used with an additional concentration step to detect 38 anthelmintic (nematicides, flukicides, endectocides) drug residues, consisting of benzimidazoles, avermectins, and flukicides in milk. UHPLC–MS-MS instrumentation detects most analytes at <1 µg/kg.
Don Shelly and Craig A. Perman, UCT, LLC
A modified QuEChERS-type extraction method is used with an additional concentration step to detect 38 anthelmintic (nematicides, flukicides, endectocides) drug residues, consisting of benzimidazoles, avermectins, and flukicides in milk. UHPLC–MS-MS instrumentation detects most analytes at <1 µg/kg.
Materials and Instruments
1) Sample Preparation
a) Weigh 10.0 g milk into a 50 mL centrifuge tube
b) Add IS and allow to sit for 15 min
c) Add 12 mL acetonitrile (MeCN) and the contents of ECMSSC50CT-MP (4000 mg anhydrous MgSO4, 1000 mg NaCl) pouch
d) Shake vigorously then centrifuge for 12 min @ 3500 rpm
2) Dispersive Sample Cleanup
a) Add the supernatant to ECMSC1850CT (1500 mg anhydrous MgSO4 and 500 mg C18) in a 50 mL centrifuge tube
b) Vortex sample for 30 s
c) Centrifuge for 10 min @ 2500 rpm (1449g)
d) Transfer 6 mL of supernatant to an evaporation tube
e) Add 0.25 mL DMSO and vortex for 1 min
f) Evaporate the MeCN @ 50 °C using a Turbovap® or similar apparatus to 0.25 mL
g) Filter extract using 0.2 µm PTFE syringe filter
h) Sample is ready for UHPLC analysis
3) Analysis by UPLC–MS-MS
- Waters Acquity UPLC system (Milford, Massachusetts) or equivalent
- Analytical column SS HSS T3 (or equivalent) (100 × 2.1 mm, particle size 1.8 µm) with in-line filter unit and 0.2 µm stainless steel filter
- Maintain column at 60 °C
- Pump flow rate of 0.6 mL/min
- Binary gradient:
1) mobile phase A 0.01% HOAc in water: MeCN (90:10, v/v)
2) mobile phase B 5 mM ammonium formate in MeOH: MeCN (75:25 v/v)
Injection volume 5 µL
- Waters Quattro Premier XE triple quadrupole mass spectrometer
- 200 electrospray ionization (ESI) interface
- System control 201 by MassLynx™ software and data was processed using TargetLynx™ Software (Waters)
- Parent ion masses are calculated from the elementary compositions
Table 1
Information on standards available in full article summary on the UCT website.
*Used with permission from Whelen, M., Kinsella, B., "Determination Of Anthelmintic Drug Residues In Milk Using Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry With Rapid Polarity Switching", doi:10.1016/j.chroma.2010.05.007, CHROMA 351049, J. of Chromatography A.
Listing of instrument manufacturers and standards suppliers does not constitute endorsement by UCT. Equivalent systems may be used.
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