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Bioanalysis. 2011 May;3(9):951-65. doi: 10.4155/bio.11.59.

Impact of methylation of acyl glucuronide metabolites on incurred sample reanalysis evaluation: ramiprilat case study.

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  • 1Algorithme Pharma Inc., 575 Armand-Frappier, Laval, Québec, Canada.



Reanalysis of incurred samples showed that the bioanalytical method for the quantification of ramipril and ramiprilat was generating irreproducible results for ramiprilat.


An additional peak interfering with ramiprilat was observed in the incurred samples but not in the calibrant and quality control samples. A similar interference was detected for ramipril, but it was chromatographically separated. Interferences were produced during sample preparation, which involves strong cation exchanger cartridges. The interfering products corresponded to the methylation of ramipril and ramiprilat glucuronide.


Following this discovery, a reproducible method was developed and successfully validated for ramipril and ramiprilat. Additional stability tests were performed in the presence of glucuronide and diketopiperazine metabolites of ramipril and ramiprilat to demonstrate the method specificity.

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