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Determination and Confirmation of Leucomalachite Green in Salmon using No-Discharge Atmospheric Pressure Chemical Ionization LC-MSn Sherri B. Turnipseed, Wendy C. Andersen, José E. Roybal Animal Drugs Research Center, Food and Drug Administration, Denver, CO
ABSTRACT An LC-MS method utilizing no-discharge atmospheric pressure chemical ionization (APCI), in conjunction with an ion trap instrument to generate product ion spectra, was developed to quantitate and confirm residues of leucomalachite green (LMG) in salmon tissue after conversion to the chromic malachite green (MG) in the extraction process. For the most efficient use of laboratory resources, the same extracts prepared for an LC-VIS method can be used for LC-MS analysis. The sample preparation procedure involves extraction of tissue into acetonitrile/buffer, partition of LMG residue into methylene chloride, conversion of LMG to MG using an organic oxidizing agent, followed by the isolation of MG on alumina/propyl sulfonic solid phase extraction cartridges. The validation of this method was performed by fortifying salmon with different levels of LMG, and then detecting the residue as MG. LC-MS conditions, including a comparison of electrospray and no-discharge atmospheric pressure chemical ionization (APCI), were evaluated and optimized. Confirmation criteria for retention time and product ion relative abundances that need to be met for positive identification of MG are provided. MG was not confirmed in any of the control tissue extracts, and all fortified samples analyzed in the validation of this method met the confirmation criteria as described. In addition to providing confirmatory data, this method can provide an alternative quantitation method for MG in salmon. The recoveries of LMG, measured as MG by this LC-MS method, at all fortification levels (1-10 ppb) were very high (87-109 %), with low relative standard deviations (
