Development of liquid chromatographic method for the analysis of kanamycinresidues in varicella vaccine using phenylisocyanate as a derivatization reagent

A liquid chromatographic method for the determination of the aminoglycoside kanamycin in varicella vaccine is described. Kanamycin sulfate was derived with phenylisocyanate (PIC) and triethylamine for 10 min at 70 degreesC an d chromatographed on a alkylamide-bonded column, Suplex pKb-100. A derivati ve of kanamycin sulfate was attached to four phenylisocynato groups and tha t molecular mass was confirmed with liquid chromatography-electrospray ioni zation mass spectrometry (LC-ESI-MS). The kanamycin-PIC derivative was foun d to have a retention time of 11.7 min using an eluent composed of 40% acet onitrile in water at 1.2 ml/min column flow-rate. Detection was at a wavele ngth of 240 nm. Recoveries ranging from 97.5 to 99.8% were found. The corre lation coefficient was greater than 0.9998 over the range between 10 and 10 0 mug/ml. The method precision of within-day assay showed a 0.5 to 4.0% coe fficient of variation (n = 5) ranging from 10 to 70 mug/ml of kanamycin con centration levels. Kanamycin-PIC derivative in reaction solution was stable for 24 h at room temperature. A simple and efficient method for the analys is of the kanamycin in varicella vaccine was developed and validated. (C) 2 001 Elsevier Science B.V. All rights reserved.