Determination of malachite green residues in rainbow trout muscle with liquid chromatography and liquid chromatography coupled with tandem mass spectrometry

Abstract

A method for the determination of malachite green and its major metabolite leucomalachite green in rainbow trout muscle is reported with limits of detection of 0.8 and 0.6 μg kg⁻¹, respectively. Residues were extracted with an acetonitrile–acetate buffer mixture and partitioned into methylene chloride. Clean-up of the extracts was performed on alumina and propylsulfonic acid solid-phase extraction columns using the automated solid-phase extraction system. The chromatographic separation of malachite green and leucomalachite green was achieved on a Chromspher 5B column using an acetonitrile–acetate buffer mobile phase. Leucomalachite green was converted to malachite green by post-column oxidation before spectrophotometric detection at 600 nm. The mean recoveries of malachite green and leucomalachite green from control rainbow trout muscle spiked at 2–50 μg kg⁻¹ were 65% (range 63.4–65.9%, relative standard deviation 3.9–16.1%) and 74% (range 58.3–82.6%, relative standard deviation 3.3–11.4%), respectively. Qualitative confirmation of the determined residues was performed with liquid chromatography coupled with tandem mass spectrometry detection with limits of detection of 2.5 and 1 μg kg⁻¹ for malachite green and leucomalachite green, respectively.

Keywords:

• malachite green
• leucomalachite green
• fish
• automated solid-phase extraction system (ASPEC)
• liquid chromatography (LC)
• LC-VIS
• liquid chromatography coupled with tandem mass spectrometry detection (LC-MS/MS)

Acknowledgements

The authors thank Christina Bäckman and Mervi Rokka, National Veterinary and Food Research Institute (EELA), for support in method development. They also thank Riitta Linjakumpu, EELA, for performing the recovery studies.