Instability of calcium channel antagonists during sample preparation for LC–MS–MS analysis of serum samples

Abstract 

1,4-Dihydropyridines calcium channel antagonists (1,4-DHP CCAs) are photolabile and the products of their photodecomposition have no pharmaceutical activity. In our previous work we have presented a screening procedure for eleven 1,4-DHPs in plasma by LC–MS–MS using multiple reaction motoring. The laboratory process includes preparation and storage of stock solutions, plasma storage, solid-phase extraction, reconstitution of extracts and storage time in an autosampler for LC–MS–MS analysis. Prior to validation of the analytical procedure, we have tested the stability of these compounds by exposure to light. Methanolic solutions have been exposed to laboratory and UV light and the stability of the compounds in plasma was tested by exposure of spiked plasma samples to laboratory light at room temperature. Stability during freeze–thaw cycles and stability during 2 month storage at −20°C have been tested as well. Products of photodecomposition have been identified after forced degradation and the degree of degradation has been quantified using LC–UV-DAD and LC–MS–MS, respectively. A 96% degradation after only 2h has been observed when solutions of nifedipine or nisoldipine were exposed to laboratory light in clear glass vials. In plasma samples degradation was 25% in only 2h for both compounds. The main degradation product was produced by oxidation of the dihydropyridinic ring resulting in the pyridine analogue that has been described as the first metabolite in the metabolic pathway. Only minor degradation was found for the other tested compounds after 2h light exposure in methanolic solutions. Furthermore, lercanidipine and nicardipine were also degradated by esterhydrolysis. Several additional minor degradation products were found for the other tested 1,4-DHPs, however, some of them could not be identified. Preconditions for storage and handling of plasma samples prior to and during analysis for 1,4-DHP CCAs are suggested in order to avoid photodecomposition of the analytes.

Keywords: Instability, 1,4-Dihydropyridine calcium channel antagonists, LC–MS–MS, Plasma