Journal of Analytical & Bioanalytical Techniques
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Cardiovascular disease is the biggest killer in the UK and yet some 60% of patients are reported not to take their
medication correctly. The use of blood spot collection cards was investigated as a means of obtaining small volume samples
for the analysis of therapeutic drugs for assessing medication adherence in cardiovascular therapy. An accurate mass liquid
chromatography-high resolution mass spectrometry (LC-HRMS) method was developed and validated for the determination of
the top three prescribed cardiovascular drugs in the UK, bisoprolol, ramipril and simvastatin, in dried blood spots (DBS).
For the preparation of DBS samples whole blood spiked with the three target analytes was used to produce 30 μl blood
spots on specimen collection cards. An 8 mm disc was cut from the dried blood spot and extracted using methanol: water (70:30,
v/v) containing the internal standard, atenolol. Extracts were vortexed, sonicated and then centrifuged. Analysis was performed
using gradient chromatographic elution and a mobile phase flow rate of 0.6 ml/min and the column oven temperature at 40?C
with a run time of 3 min. MS detection was carried out in electrospray positive ion mode at accurate mass m/z 326.2326 for
bisoprolol, m/z 417.2384 for ramipril, m/z 441.2611 for simvastatin and m/z 276.1703 for the IS. The developed bioanalytical
method was applied to blood spots samples taken from adult volunteers previously administered one or more of the target drugs.
Drug recoveries from spiked blood spots were ≥92% for bisoprolol and ramipril and ~43% for simvastatin and the
drugs were stable in DBS for at least 12 weeks. Validation of the LC-HRMS method showed good linearity and the accuracy
and precision values were within the pre-defined limits of ≤15% at all concentrations. Factors with potential to affect drug
quantification measurements such as the matrix effects and volume of blood applied onto the collection card were investigated.
The accurate mass LC-HRMS DBS based method successfully identified control volunteers who were known to
be either adherent or non-adherent. There were no false positives from volunteers taking other cardiovascular drugs or from
volunteers receiving no medication. Initial research has already identified one case of medication taken incorrectly. This
demonstrates that the DBS based assay has the potential to assess patient adherence for a range of therapeutic drugs.
Sangeeta Tanna is a reader in Pharmaceutical Bioanalysis in the Leicester School of Pharmacy at De Montfort University. Her expertise and research
interest is in the bioanalysis and drug delivery fields. This has led to the development of micro-analytical methodologies for the determination of
therapeutic drugs from dried blood spots (DBS) based on LC-MS and LC-MS/MS studies for a range of clinical applications. This research was
awarded the Royal Society of Chemistry Analytical Methods Prize in 2010. Applications of this work to patient care include improved medication for
babies and to people with cardiovascular diseases.
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