HPLC Determination of Spectinomycin in Feed Premixes and Dosage Forms Using 1-Naphthyl Isocyanate Precolumn Derivatization with UV Detection

A new, simple, and sensitive HPLC method was developed for the determination of spectinomycin hydrochloride in dosage forms and feed premixes through derivatization with 1-naphthyl isocyanate. The separation was achieved on a C18 column using a mobile phase consisting of acetonitrile/water (50:50 v/v, pH 3.2) in a flow rate of 1 mL/min with UV detection at 230 nm. The factors influencing the derivatization reaction yields were carefully studied and optimized. The method was linear over the concentration range of 10–100 μg/mL with a limit of detection 0.25 μg/mL and limit of quantitation 1.75 μg/mL. The developed method was successfully applied to the analysis the drug in the commercial dosage forms and spiked feed premix samples; the average recoveries were 99.79 and 99.56, respectively. The analytical performance of the method was fully validated and the results were satisfactory. A proposal of the reaction pathway was presented.