Development and Validation of a Dried Blood Spot Assay Using UHPLC-MS/MS to Identify and Quantify 12 Antihypertensive Drugs and 4 Active Metabolites: Clinical Needs and Analytical Limitations.

Purpose: As nonadherence to antihypertensive drugs (AHDs) can increase the risk of cardiovascular events, hospitalization, and higher costs, there is a need for a reliable, objective, and easy method to assess nonadherence in patients. The dried blood spot (DBS) sampling method used to measure drug concentrations meets these requirements. For detecting nonadherence, identification is more important than quantification. Owing to their use in clinical practice, it is important to measure multiple AHDs with a single method. Therefore, we developed and validated a single DBS method for 17 commonly used AHDs and 4 active metabolites using ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS).

Methods: Analytical validation of the DBS assay was performed in accordance with the guidelines on bioanalytical method validation of the European Medicines Agency and US Food and Drug Administration as well as the International Association of Therapeutic Drug Monitoring and Clinical Toxicology guidelines.

Results: We validated 12 of the 17 AHDs according to the European Medicines Agency and Food and Drug Administration requirements for bioanalytical method validation. Eleven AHDs were validated for both identification and quantification of drug concentrations, whereas nifedipine was only validated for identification. However, 5 of the 17 AHDs were excluded due to suboptimal validation results. Lercanidipine was excluded due to nonlinearity, and all 4 AHDs measured in the negative mode of UHPLC-MS/MS were not in accordance with one or more of the acceptance criteria and were therefore excluded.

Conclusions: The described method accurately measured AHDs in DBS and can be used to determine nonadherence in patients. However, method validation revealed a challenging balance between analytical limitations and clinical needs when analyzing multiple drugs using the same method.