Quantitative fluorometric in-situ determination of netilmicin and 4 gentamicins on TLC-RP-layers

Summary

This investigation describes a thin-layer chromatographic method for the quantitative determination of netilmicin and gentamicins C₁, C_1a, C₂ and C_2a in pharmaceutical preparations. The individual components are separated on C₈ or C₁₈ reversed phase layers with a mobile phase consisting of methanol and ammonia with added lithium chloride. After optimization of the post-chromatographic derivatization with 2,2-diphenyl-1-oxa-3-oxonia-2-boratanaphthalene (DOOB) quantitative determination takes place directly on the layer. The calibration curves are linear for netilmicin in the range 50–250 ng/zone and for the gentamicin components in the range 30–140 ng/zone (this corresponds to 100 to 400 ng/zone total complex). The determination limits per zone are 50 ng total gentamicin complex and 10 ng netilmicin per zone. No clean-up is required for analysis because of the great specificity of the derivatization reaction. The reproducibility of the method for independently carried out measurement series can be described by coefficients of variation between CV = ±1.7%–4.2%.