A Validated UHPLC-MS/MS Method to Quantify Eight Antibiotics in Quantitative Dried Blood Spots in Support of Pharmacokinetic Studies in Neonates.

Objectives: Conduction of pharmacokinetic (PK) study in pediatric patients is challenging due to blood sampling limits. The dried blood spots (DBS) method represents a potential matrix for microsampling in support of PK studies in children. Herein, we used the Capitainer qDBS device to develop a DBS method that can collect an exact 10 µL volume of blood on a paper card. This DBS method was developed to simultaneously quantify the concentrations of eight antibiotics, including sulbactam, tazobactam, ampicillin, meropenem, cefotaxime, cefoperazone, piperacillin, and metronidazole using ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS).

Methods: The prepared DBS samples were extracted in methanol containing acetaminophen as the internal standard at 20 °C on a block bath shaker at 500 rpm for 30 min. The extracted antibiotics were eluted on an Acquity UPLC HSS T3 column (2.1 × 50 mm, 1.8 µm) using gradient elution with a total chromatographic run time of 6.5 min. The precursor and product ions of the analytes were detected by use of the multiple reaction monitoring (MRM) mode.

Results: No interfering peaks at the respective retention times of the analytes were observed in DBS samples. The lower limits of quantification (LLOQ) for the antibiotics were between 0.25 and 2.0 μg/mL, and satisfactory accuracies (intra/inter-assay bias -16.7 to +13.6%) and precisions (intra/inter-assay coefficient of variations 1.5-15.6%) were obtained for the analytes. As a proof of concept, the method was applied to DBS samples obtained from neonatal patients treated with ampicillin and piperacillin/sulbactam.

Conclusions: The DBS method is simple and robust, and it can be used in children with limited blood sampling.