ESSENTIAL PEAK AREA NORMALIZATION FOR QUANTITATIVE IMPURITY CONTENT DETERMINATION BY CAPILLARY ELECTROPHORESIS

A number of papers have been published [1-5] which mention that normalisation of CE peak areas (ie division of peak areas by migration time) is necessary to ensure correct quantitation. However, there is a general unawareness of the impact of not performing this simple data manipulation upon impurity results when expressed as %area/area. An exercise has been conducted to exemplify the need for this normalisation using the separation of selected pharmaceuticals as illustrative examples. The impact of normalisation on %area/area results was demonstrated using the pharmaceutical ranitidine, and a synthetic precursor, as test solutes. The UV absorbance of each compound was determined and found to be equivalent. A solution of ranitidine hydrochloride was then spiked with a weighed amount of precursor. The %area/area CE results, when normalised, matched the known weighed radio. Use of uncorrected peak area data in this instance would have resulted in a severe underestimation of impurity levels. A quantitative analysis of drug related impurities was conducted at three levels of operating voltage whilst keeping all other operating parameters constant. This produced electropherograms having identical peak profiles but each peak having a different retention time and peak area. However, when normalised, the results were identical at each level of operating voltage confirming the validity and necessity of normalisation. A chiral separation of a racemic pharmaceutical was conducted. The uncorrected peak area data indicated the test sample was not racemic whilst the corrected data correctly confirmed that the compound was racemic. The significance and impact upon reported purity data of not normalising peak area data has been clearly demonstrated in this paper.