Evaluation of three methods for recovery of sulfamethazine metabolites from swine tissue

Recent investigations have clearly established the presence of desaminosulfamethazine (I), N4-acetylsulfamethazine (II), and N4-D-glucosyl sulfamethazine (III) in the tissues of swine which have been dosed with sulfamethazine (V). N4(1-Deoxy-D-glucuronyl) sulfamethazine (IV) has frequently been found in the urine and feces of animals. These metabolites have generally not been investigated in current sulfamethazine methods; therefore, it is not clear to what extent they might be measured by these procedures. Metabolites I, II, III, and IV were synthesized and characterized by several chemical and physical measurements. These metabolites and their parent, V, were added to swine tissue and recoveries were measured by a gas-liquid chromatographic (GLC), a colorimetric, and a gas chromatographic/mass spectrometric method with electron ionization-selected ion monitoring (GC/MS EI-SIM). With modifications of the gas chromatographic parameters, the GLC method can quantitate compounds, I, II, and V simultaneously. If a hydrolysis step is added, all 3 methods can quantitate compound II as V and, in addition, the GC/MS EI-SIM and colorimetric methods can quantitate III as V. In none of these methods do metabolites I, II, III, or IV interfere in the determination of V.