Derivatisation efficiency in the tissue matrix was determined by adding the underivatised analytes (metabolites) to blank tissue homogenate before the derivatisation step and adding the corresponding amounts of 2-NBA derivatives to identical samples after the derivatisation procedure. Recovery of solid-phase extraction was determined by adding the 2-NBA derivatives before and after the SPE step.
2.7. Contaminated tissue
In the course of an animal feeding study at the Bundesanstalt für Tierseuchenbekämpfung (Mödling, Austria), furazolidone and furaltadone were administered in therapeutic doses to one pig per investigated drug. For these studies, 3.5 g of each drug were dissolved in 10 ml ethanol and over a period of 3 days, 1-ml aliquots of the solutions were added to pig feed rations (approximate dosage: 5 mg per kg body weight per portion). Three to four rations were offered to the pigs for three consecutive days. After the end of the third day, the pigs were killed and muscle samples collected. Samples were stored at −20°C prior to analysis.
3. Results and discussion
3.1. Method development
Several crucial aspects regarding sample preparation, chromatography and MS detection had to be considered during the development of this assay.