title: New approach of validation using internal normalization technique for quantification of related substances in raw material, intermediates and pharmaceutical substances by HPLC.
abstract: Internal normalization (IN) serves as a quantitative tool in gas chromatography. Nevertheless, its utilization in liquid chromatography is not widely employed, as several requirements need to be taken into account. However, IN can be used in case of relative amounts estimation when the absolute concentration is not the crucial factor. This suits very well in pharmaceutical analysis when the relative amount of active pharmaceutical ingredient (API) impurities is to be estimated with a limited knowledge of statistics, such as t-test and linear regression. The determination of three prasugrel impurities in the real sample by means of IN and the comparison of these results with external standard calibration was presented. The IN method was validated by test of population means and variances agreement and the agreement of external calibration and IN was performed by Student t-test. The influence of impurities concentration above and below is also discussed as well as the validation parameters, LOD and LOQ. It was found that the results achieved by external calibration and IN are statistically the same and, therefore, IN is a proper method for relative amount estimation of API impurities.
authors.affiliation: Zentiva, k.s. Praha, a Sanofi Company, U Kabelovny 130, 102 37 Praha 10, Czech Republic. Electronic address: michal.dousa@seznam.cz.
authors.forename: Michal
authors.initials: M
authors.lastname: Douša
authors.affiliation: Zentiva, k.s. Praha, a Sanofi Company, U Kabelovny 130, 102 37 Praha 10, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Gibala
authors.affiliation: Zentiva, k.s. Praha, a Sanofi Company, U Kabelovny 130, 102 37 Praha 10, Czech Republic.
authors.forename: Tomáš
authors.initials: T
authors.lastname: Pekárek
mesh_terms: D002138:Calibration; D002626:Chemistry, Pharmaceutical / Q000592:standards*; D002851:Chromatography, High Pressure Liquid / Q000379:methods / Q000592:standards*; D006863:Hydrogen-Ion Concentration; D057230:Limit of Detection; D004364:Pharmaceutical Preparations / Q000737:chemistry; D000068799:Prasugrel Hydrochloride / Q000032:analysis; D015203:Reproducibility of Results
chemical_list: D004364:Pharmaceutical Preparations; D000068799:Prasugrel Hydrochloride
keywords: Internal normalization; Pharmaceutical substances; Quantification; Validation