Tranexamic acid (TXA) is an antifibrinolytic drug, with the ability to inhibit lysine binding at plasminogen receptors, used in adult trauma patients with on-going or at risk of significant haemorrhage. To understand the pharmacokinetics and pharmacodynamics of this drug in variable age groups undergoing surgeries with high blood loss, effective methods for determination of TXA in biological samples at sub-μg mL −1 are still required. We describe herein the development and validation of a method based on ultra-high performance liquid chromatography coupled to triple quadrupole-tandem mass spectrometry to quantify TXA in human plasma. An inexpensive, simple and efficient sample clean-up was implemented, not requiring matrix-matching calibration. Sample preparation consisted in protein precipitation using acetonitrile containing 0.5% (v/v) formic acid, followed by hydrophilic interaction based chromatographic separation, with elution in isocratic mode using a combination of acetonitrile and water (75:25, v/v), with quantification of TXA based on selected reaction monitoring. Good linearity was achieved (r 2 > 0.997) for TXA concentrations ranging from 30 to 600 ng mL −1 , with LOD of 18 ng mL −1 in plasma. The developed method proved to be selective, sensitive, accurate (96.4-105.7% of nominal values) and precise (RSD ≤ 4.5%). TXA was found to be stable in plasma extracts standing 24 h at room temperature (20°C) or in the autosampler, and after three freeze-thawing cycles. Mean recovery values of TXA spiked plasma samples were ≥91.9%. No significant matrix effects were observed. The proposed methodology was successfully applied to the clinical study of plasma samples recovered during scoliosis surgery of pediatric patients pretreatment with TXA.
The low bioavailability and nonspecific distribution of dapsone and clofazimine, commonly applied in combination for the treatment of leprosy, can produce toxic effects. Nanotechnological approaches enhance the delivery of these drugs. Therefore, a high-performance liquid chromatography method was developed for the simultaneous determination of dapsone and clofazimine loaded in nanoformulations for quality control purposes. Chromatographic separation was achieved on a reversed-phase Kinetex core-shell C18 column, followed by spectrophotometric detection at 280 nm. Considering the different physicochemical properties of dapsone and clofazimine, elution was performed in gradient mode using an aqueous acetate buffer (50 mmol/L, pH 4.8) and an increasing acetonitrile content from 27 to 63% v/v at a flow rate of 1.0 mL/min with retention times of 6.2 and 14.0 min, respectively. The method was validated according to the European Medicines Agency guideline and it was found to be specific, accurate (99.6-114.0%), and precise for intra- (RSD ≤ 1.8%) and interday assays (RSD ≤ 12.5%). Both drugs showed stability after 24 h at room temperature and over three freeze-thaw cycles with recoveries ≥86.2%. Low temperature (4°C) in the autosampler caused the precipitation of clofazimine and must be avoided. The validated method was successfully applied in the quantification of both drugs in nanoformulations.
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