Quantitative liquid chromatography/tandem mass spectrometry determination of chloramphenicol residues in food using sub-2 microm particulate high-performance liquid chromatography columns for sensitivity and speed

The use of chloramphenicol (CAP)--a highly effective broad-spectrum antibiotic used in animal husbandry--is banned in many countries. Therefore, a very low minimum required performance limit (MRPL) of 0.3 microg/kg CAP in meat for human consumption has been defined. Analytical methods capable of quantifying and confirming such low residue levels require sophisticated instrumentation. Preferably sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) or gas chromatography/mass spectrometry (GC/MS) methods have been used. This paper suggests the use of sub-2 microm particulate high-performance liquid chromatography (HPLC) columns to gain additional sensitivity and improve resolution as well as speed. Depending on the operating conditions, higher chromatographic resolution and speed can be obtained at the price of a significantly increased operating pressure, requiring dedicated LC equipment. A 3-4-fold overall improvement of the signal-to-noise ratio for CAP was obtained compared to more classical 5 microm particulate HPLC columns. The proposed analytical methodology includes an enzymatic digestion, which liberates glucuronide-bound CAP from kidney tissue. The extracts obtained after an Extrelut clean-up are sufficiently pure to permit routine injection of biological samples into the sub-2 microm particulate HPLC column, without observing rapid deterioration of peak shape or column clogging problems. The time for one chromatographic run was 4.2 min. The described method was validated for two particularly difficult matrices (kidney and honey). Decision limits (CC alpha) were 0.007 microg/kg (honey) und 0.011 microg/kg (kidney), which are significantly below the current MRPL.