Severe Impact of Hemolysis on Stability of Phenolic Compounds

Abstract

Eugénie-Raphaëlle Bérubé has obtained a Bachelor of Science in Biochemistry from Université du Québec à Montréal. She previously worked at the St-Lawrence Center of Environment, Canada, conducting biomarker analysis to measure the impact of contaminants on aquatic species. She has been working in the bioanalysis industry for the past 8 years at Algorithme Pharma, a CRO located in Laval, Canada, becoming a scientist in bioanalytical method development for the quantitation of pharmaceuticals in biological fluids. The presence of hemolyzed plasma samples can negatively impact preclinical and clinical sample analysis. During the method development of morphine, post-extracted instability issues were encountered in human hemolyzed plasma when compared with nonhemolyzed plasma (called normal plasma for simplicity). Investigation revealed that the presence of methemoglobin using a high pH reconstitution solution led to degradation of morphine over time. The degradation probably results from radical oxidation of the ionized phenolic group promoted by the presence of methemoglobin. Pseudomorphine, the product of oxidative dimerization of morphine, was observed as one of the degradation products in hemolyzed plasma. This hypothesis was extended to raloxifene, another phenol-containing compound. On the other hand, no instability was detected for drug products bearing a masked phenol group or carboxylic acid functionality. The issue of morphine instability was resolved by using a reconstitution solution at a pH below the pKa of the phenol moiety.