Short communicationUPLC MS/MS assay for routine quantification of dabigatran – A direct thrombin inhibitor – In human plasma
Introduction
Dabigatran etexilate is a novel oral direct thrombin inhibitor. In recent studies, dabigatran demonstrated its efficacy for prophylaxis and treatment of thromboembolic event during orthopedic surgery and curative treatment of hypercoagulability in atrial fibrillation [1], [2].
Contrary to vitamin K antagonist (VKA), no laboratory monitoring was advised for this drug because its pharmacokinetic profile is supposed to be more predictable [3], [4]. However EMEA underlined the low bioavailability and very large interindividual variability of dabigatran exposition in case of renal impairment, elderly patients or extreme weights [5]. From that point, some arguments are in favor of a monitoring assay [6]. First, dabigatran is eliminated at 80% by kidney leading to accumulation in renal impaired patients. Second, it is also a substrate of P-glycoprotein (P-gp) and may be subject to drug–drug interactions [7], [8]. P-gp inductors or inhibitors can respectively decrease or increase its bioavailability. In these situations, dabigatran accumulation could increase bleeding risk and may require therapeutic drug monitoring.
Several methods were previously published for quantification of dabigatran in phase II/III studies [1], [3], [7], [8], [9], [10], [11]. None of them presented the complete analytical process used and validation criteria according the FDA guidelines [12]. The purpose of this work was to develop a fully validated and routinely available ultra performance liquid chromatography coupled with tandem mass spectrometry (UPLC MS/MS) method by for quantification of dabigatran in human plasma.
Section snippets
Chemicals and reagents
Dabigatran and [13C6]-dabigatran were purchased from Alsachim (Strasbourg, France). LC MS grade methanol and acetonitrile were obtained from Fisher Scientific GmbH (Schwerte, Germany) and distilled water from Aguettant (Lyon, France). Hydrochloric acid 0.1 N was obtained from Sigma–Aldrich (St. Quentin Fallavier, France). A 0.2 μm polyvinylidene fluoride filter was used for mobile phases and was provided by Interchim (Montluçon, France).
Stock solutions, calibration standards and quality control samples
Stock solutions of dabigatran and internal standard [13C6
Liquid chromatography
Representative chromatograms of drug free, LLOQ and supposed over exposed human plasma samples are shown in Fig. 1. Dabigatran and IS retention times and peak shape did not varied over the whole validation period (data not shown). Retention time was near 1.6 min for dabigatran and IS, total run time of 4.5 min allowed return to initials conditions.
Mass spectrometry
In positive electrospray ionization, quantification was performed by addition of 472.19→324.17 m/z and 472.19→306.13 m/z MRM transitions for [dabigatran
Conclusion
In conclusion, this paper described the development and full validation of a single step preparation, rapid (about 10 min), sensible and accurate ultra performance liquid chromatography method using tandem mass spectrometry detection for dabigatran quantification in human plasma. This method is totally compatible with other UPLC MS/MS routine activities and could be applied to 24/7 clinical toxicology to monitor dabigatran accumulation and therapeutic drug monitoring [13].
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A chemometric comparison of different models in fluorescence analysis of dabigatran etexilate and dabigatran
2021, Spectrochimica Acta - Part A: Molecular and Biomolecular SpectroscopyCitation Excerpt :But these methods are usually inferior in accuracy, sensitivity, selectivity and universality [14]. Some direct determination methods such as liquid chromatography or ultra performance liquid chromatography with tandem mass spectrometry (LC-MS/MS or UPLC-MS/MS) [15–25], high performance liquid chromatography (HPLC) combined with ultraviolet detection (UV) [26–29], diode array detection (DAD) [30,31] or fluorescence detection (FLD) [32] have been also reported to measure DABE or DAB levels in different matrices. However, the instruments for mass spectrometric methods are expensive and require individuals with specialized experience, which makes it difficult for routine clinical applications.
Gradient RP-HPLC method for the determination of potential impurities in dabigatran etexilate in bulk drug and capsule formulations
2019, Arabian Journal of ChemistryCitation Excerpt :Dabigatran etexilate converted to a major active metabolite known as dabigatran by non-specific plasma and liver esterase (Blech et al., 2008). Several research papers have been reported in the literature for the determination of DAB (Bernardi et al., 2013a,b; Damle and Bagwe, 2014; Geetharam et al., 2014; Pani Kumar et al., 2015; Khan et al., 2014; Reddy and Rao, 2014) and these papers were limited to the assay of alone DAB from its degradants and concurrently DAB in human plasma and other biological stuff, based on UPLC MS/MS (Delavenne et al., 2012), HPLC and LC/MS/MS (Bernardi et al., 2015; Nouman et al., 2015; Laizure et al., 2014; Li et al., 2014; Härtter et al., 2012). Monitoring and control of by-products or impurities at each stage of synthesis is required to get a final pure form of DAB as per predefined specifications.
An improved extraction protocol for therapeutic dabigatran monitoring using HPLC-MS/MS
2019, Journal of Chromatography B: Analytical Technologies in the Biomedical and Life SciencesCitation Excerpt :However, the INR does not provide an accurate assessment of anticoagulant activity of new oral anticoagulants such as dabigatran, so other tests are required to assess or to adjust dosing in emergency situations. Gas-liquid chromatography [3], chromogenic tests [4–6], high-performance liquid chromatography combined with mass-selective spectrometry detection [7–9], have been used for the quantification of dabigatran in plasma. The HPLC-MS/MS method provides a highly selective and direct determination of the dabigatran in human plasma over a wide analytical concentration range.
Measurement of apixaban, dabigatran, edoxaban and rivaroxaban in human plasma using automated online solid-phase extraction combined with ultra-performance liquid chromatography-tandem mass spectrometry and its comparison with coagulation assays
2018, Clinica Chimica ActaCitation Excerpt :Accuracy, intra- and inter-assay precision of our online SPE-UPLC-MS/MS assay for all four DOACs were high, as presented in Table 3. These assay parameters of our method are generally comparable with those of previously described LC-MS/MS assays of DOACs, albeit with most of these assays being limited to measurement of only one, two, or three DOACs [11,13–21,39,40], and only three of them measuring all four DOACs [14,20,21], however, none of which used automated online solid-phase extraction for sample preparation to obtain high-purity samples which prevent mass spectrometer contaminations. The recovery of our assay was good for dabigatran (>90%), edoxaban (>85%), rivaroxaban (>90%), and moderate for apixaban (>60%) (Table 3).
Microdosing Cocktail Assay Development for Drug–Drug Interaction Studies
2018, Journal of Pharmaceutical SciencesCitation Excerpt :The interconversion between the 2 forms depends on pH, enzyme activity, sample processing temperature, and time.29 Methods for the determination of dabigatran in human plasma have also been reported,30-34 but these methods lack the sensitivity required to support microdosing studies. Several methods exist for midazolam,35-37 one assay was sensitive and was utilized for a microdose study; however, sample preparation was expensive and complex.36