Rapid and sensitive high-resolution procedure for digitalis glycoside analysis by derivatization liquid chromatography.

The separation and quantitative determination of digitalis glycosides by high performance liquid chromatography following derivatization with 4-nitrobenzoylchloride (4-NBC1) is described. The compounds of primary interest were the digitalis glycosides and aglycones of the pharmaceutically important A, B and C series, The derivatization step results in higher extinction values at a more favourable wavelength (260 nm), which permits the use of low-cost ultraviolet detectors. Detection limits are below 20 ng/ml for all of the glycosides tested. The chromatographic properties are also improved by reducing the polarity without a decrease in selectivity. The use of low-polarity and low-viscosity solvent systems on silica gel adsorbents permits rapid isocratic separations of complex mixtures as they usually occur in pharmaceutical products and extracts. The quantitative potential of this method was demonstrated by analyzing ampoule solutions containing desacetyl lanatoside C as the active compound. The active substance, by-products and degradation products were determined down to 0.1% of the total glycoside concentration in one ampoule.