Quantification of ascorbic acid and acetylsalicylic acid in effervescent tablets by CZE-UV and identification of related degradation products by heart-cut CZE-CZE-MS.

Capillary electrophoresis is commonly applied for the analysis of pharmaceutical products due to its high separation efficiency and selectivity. For this purpose, electrospray-ionization-(ESI)-interfering additives or electrolytes are often required, which complicates the identification of impurities and degradation products by mass spectrometry (MS). Here, a capillary zone electrophoresis (CZE) method with ultraviolet (UV) absorption detection for the simultaneous determination and quantification of ascorbic acid and acetylsalicylic acid in effervescent tablets was developed. Related degradation products were identified via CZE-CZE-MS. Systematic optimization yielded 100 mM tricine (pH = 8.8) as appropriate background electrolyte, resulting in baseline separation of ascorbic acid, acetylsalicylic acid, and related anionic UV-active degradation products. The CZE-UV method was successfully validated regarding the guidelines of the Food and Drug Administration. The validated method was applied to trace the degradation rate of the active pharmaceutical ingredients at defined ambient conditions. A heart-cut CZE-CZE-MS approach, including a 4-port-nL-valve, was performed for the identification of the observed degradation products. This 2D setup enables a precise cutting of accurate sample volumes (20 nL) and the independent operation of two physically separated CZE dimensions, which is especially beneficial regarding MS detection. Hence, the ESI-interfering tricine electrolyte components were separated from the analytes in a second electrophoretic dimension prior to ESI-MS detection. The degradation products were identified as salicylic acid and mono- and diacetylated ascorbic acid. This setup is expected to be generally applicable for the mass spectrometric characterization of CZE separated analytes in highly ESI-interfering electrolyte systems. Graphical Abstract A CZE-UV method for the quantification of effervescent tablet ingredients and degradation products was developed and validated. In order to identify unknown degradation products separated in the CZE-UV, a 2D heart-cut approach was performed applying a mechanical 4-port-valve. The unknown substances were transferred from the 1st to the 2nd dimension followed by the separation of ESI-interfering tricine from the analytes prior to mass spectrometric detection.