Method development for quantitative determination of seven statins including four active metabolites by means of high-resolution tandem mass spectrometry applicable for adherence testing and therapeutic drug monitoring

Abstract

BackgroundStatins are used to treat and prevent cardiovascular diseases (CVDs) by reducing the total serum cholesterol concentration. Unfortunately, dose-related side effects and sub-optimal response, attributed to non-adherence amongst others, were described. Therefore, a fast and sensitive liquid chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) method for adherence testing and therapeutic drug monitoring of all currently marketed statins and their active metabolites in human blood plasma should be developed, validated and tested for applicability.MethodsAtorvastatin, fluvastatin, lovastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin, as well as ortho- and para-hydroxy-atorvastatin, lovastatin hydroxy acid and simvastatin hydroxy acid were included and several internal standards (IS) tested. Validation was performed according to the guideline of the European Medicines Agency including selectivity, carry-over, accuracy, precision, matrix effects, dilution integrity and analyte stability. Finally, applicability was tested using 14 patient samples submitted for regular toxicological analysis.ResultsDue to an analytical interference of atorvastatin-d5, diazepam-d5 and pentobarbital-d5 were chosen as IS for positive and negative ionization mode, respectively. All statins and metabolites fulfilled the validation acceptance criteria except for fluvastatin, which could not be quantified reliably and reproducibly, most probably due to instability. Analyses of human plasma samples revealed concentrations of statins and metabolites below the reference plasma concentrations in the case of eight patients. However, nothing was known concerning patients’ adherence and time between intake and sampling.ConclusionsAn LC-HRMS/MS method for identification and quantification of atorvastatin, lovastatin, pitavastatin, pravastatin, rosuvastatin, simvastatin and four active metabolites was successfully developed and applicability demonstrated.

New steps in acute coronary syndrome and antihyperlipidemic treatment: Determination of statins and metabolites by liquid chromatography tandem mass spectrometry

“…Wagmann et al developed a high‐resolution tandem mass spectrometry method based on liquid–liquid extraction with pretreatment steps for the measurement of seven statins and their four metabolites, including atorvastatin and rosuvastatin. The analysis time of the method was 11 min, and intra‐ and inter‐assay CV values ranged between 6 and 15% (Wagmann et al, 2020). In our method, the recovery results of the 250, 31 and 3.9 ng/ml concentrations of atorvastatin were 99.01, 98.87 and 97.17%, respectively; the recovery results of rosuvastatin concentrations of 125, 31 and 3.9 ng/ml were determined as 98.16, 100.3 and 102.05%, respectively.…”

A review on liquid chromatographic methods for the bioanalysis of atorvastatin

“…AT and its metabolites can be detected in both positive and negative ionization modes of MS, yet most of the published work has performed LC-MS/MS analyses of AT in positive ionization mode using electrospray ionization (ESI), but few reported studies used negative ions [54,56,58,67,80,94] of AT for the analysis even though, polarity switching within run is required in ESI − mode [89]. On the other hand, the atomic pressure chemical ionization (APCI) technique has been used by Tahboub for ionization of AT in positive mode [64].…”

“…Various bioanalytical methods used LLE strategy for the sample pre-treatment of AT before the final analysis by using ethyl acetate, methyl-tertiary butyl ether, dichloromethane, or diethyl ether as organic extraction solvents. Instead of the single solvent procedure, various methods have also used a combination of different solvents [17,27,53,54,69,85,86] or various buffering agents such as FA [17,60]. AA [20] and phosphate buffer [33,[70][71][72]85] for the extraction of AT using LLE method.…”

Validation of a novel UPLC‑HRMS method for human whole‑blood cyclosporine and comparison with a CMIA immunoassay

“…Ultra-performance liquid chromatography coupled to high-resolution mass spectrometry (UPLC-HRMS) has already been proven as an important analytical tool for the quantification of drugs (8)(9)(10)(11). Although triple quadrupole mass spectrometers are commonly present in clinical laboratories (12,13), there are major advantages to the application of UPLC-HRMS.…”