Liquid chromatographic analysis of a formulation containing polymyxin, gramicidin and neomycin

The development of a liquid chromatographic assay system for the stability study of a formulation containing polymyxin, gramicidin and neomycin is described. For the determination of each group of antibiotics, poly(styrene-divinylbenzene) is used as the stationary phase. The mobile phase for the determination of polymyxin consists of an aqueous solution containing 7 g l(-1) of sodium sulfate, 50 ml l(-1) of 1 M phosphoric acid and 160 ml l(-1) of acetonitrile. UV detection is performed at 215 nm. Gramicidin is chromatographed with a mixture of tetrahydrofuran-water (38:62) and detected by UV at 222 nm. An aqueous solution containing 70 g l(-1) of sodium sulfate, 1.4 g l(-1) of sodium octanesulfonate and 50 ml l(-1) of 0.2 M phosphate buffer pH 3.0 is used as the mobile phase for the determination of neomycin. Since neomycin has no UV-absorbing chromophore, pulsed electrochemical detection is chosen to determine neomycin. For each method, the influence of the different chromatographic parameters on the separation, the selectivity towards the other active compounds and the excipients, the repeatability and the linearity were investigated. The stability of the formulation was examined at 0, 6 and 12 months.