Identification and semiquantitation of monensin sodium in animal feeds by thin layer bioautography

A bioautographic technique for the determination of monensin sodium contamination in animal feeds is described. The feeds are extracted in aqueous methanol and the initial monensin extracts are isolated by filtration through an alumina column. These eluates are partitioned between 5% NaCl and methylene chloride, and are further purified through a Sephadex LH-20 column. A 10 mL eluate containing the monensin is collected from the Sephadex column and evaporated, and the residue is dissolved in methylene chloride. Aliquots are spotted on a thin layer plate and monensin is detected by a thin layer bioautographic technique, using Bacillus subtilis as the test organism. The reliable limit of sensitivity is 100 ppb, but 10 ppb can be detected. This technique can be used to semiquantitate monensin by comparing the zones of inhibition of unknown test samples against monensin standards.