Rapid Determination of Nitroimidazole Residues in Honey by Liquid Chromatography/Tandem Mass Spectrometry

Abstract

Abstract We developed a rapid and efficient means of determining residues of four nitroimidazoles—i.e., dimetridazole, ipronidazole, metronidazole, and ronidazole—and three hydrophilic metabolites—i.e., 2-hydroxymethyl-1-methyl-5-nitroimidazole, 1-methyl-2-(2′-hydroxyisopropyl)-5-nitroimidazole, and 1-(2-hydroxyethyl)-2-hydroxymethyl-nitroimidazole—in honey. We applied a QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) procedure improved to suit a nitroimidazole analysis, which is fast (approximately 30 min) and uses less organic solvent. The procedure involves initial single-phase extraction of 5 g of honey with acetonitrile containing 1% acetic acid, followed by liquid–liquid partitioning involving the addition of 5 g sodium chloride, 1.5 g trisodium citrate dihydrate, and 4 g magnesium sulfate. Moreover, matrix from honey was reduced by an SPE method with an alumina-N cartridge. The samples were analyzed using LC/MS/MS. Chromatographic separation of these nitroimidazoles and metabolites was performed in the gradient mode on a pentafluorophenylpropyl-bonded silica column (150 × 2.0 mm, 3 μm particle size) at 40°C. The mobile phase consisted of a 0.01% acetic acid solution and acetonitrile, and the flow rate was 0.2 mL/min. The method was validated using honey spiked with these nitroimidazoles from 0.1 to 0.5 μg/kg. The overall recovery of the seven nitroimidazoles ranged from 76.1 to 98.5%; intra- and interassay CV values were <9.5 and <14.2%, respectively. The LOQ ranged from 0.1 to 0.5 μg/kg. LC/MS/MS coupled with the QuEChERS method showed good potential as a method for determining nitroimidazole residues in honey.