Method for isolation and determination of pyridostigmine and metabolites in urine and blood

A procedure is presented for the unique isolation and determination by liquid chromatography of pyridostigmine and its metabolites in urine and blood. Isolation is accomplished by an extension of paired-ion theory using a reversed-phase chromatographic column. The assay of pyridostigmine is linear in concentrations from 40 to 5000 ng/ml. Separation and quantitation of pyridostigmine and metabolites in urine and blood are rapid. An analysis can be performed in 15 min. The method has been applied to the determination of urinary excretion and plasma levels of pyridostigmine administered intramuscularly in rats and to the isolation of acetylcholine, neostigmine, and edrophonium from aqueous solutions.