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Updated: Aug 9, 2025
Multiplex Therapeutic Drug Monitoring by Isotope-dilution HPLC-MS/MS of Antibiotics in Critical Illnesses
Published on: August 30, 2018
A Validated UHPLC-MS/MS Method to Quantify Eight Antibiotics in Quantitative Dried Blood Spots in Support of
Qian Liu1, Lanyu Liu1, Yu Yuan1
1Division of Biopharmaceutics and Pharmacokinetics, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha 410013, China.
Insights
Dried blood spots (DBS) offer a simple solution for pediatric pharmacokinetic studies. This validated method accurately quantifies eight antibiotics from minimal blood volumes, simplifying drug monitoring in children.
Area of Science:
- Pharmacology
- Analytical Chemistry
- Pediatric Medicine
Background:
- Pediatric pharmacokinetic (PK) studies face challenges due to limited blood sampling volumes.
- Dried blood spots (DBS) present a viable microsampling matrix for pediatric PK analysis.
Purpose of the Study:
- To develop and validate a DBS method for simultaneous quantification of eight antibiotics in pediatric blood samples.
- To utilize the Capitainer® qDBS device for precise 10 µL blood collection.
Main Methods:
- Developed a DBS extraction method using methanol and acetaminophen as internal standard.
- Employed ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) for antibiotic quantification.
- Utilized a 6.5-minute gradient elution method on an Acquity UPLC HSS T3 column.
Main Results:
- The DBS method demonstrated no interfering peaks for the eight antibiotics.
- Achieved lower limits of quantification (LLOQ) between 0.25 and 2.0 μg/mL.
- Exhibited satisfactory accuracy (-16.7 to +13.6%) and precision (1.5-15.6%) for intra/inter-assay analyses.
Conclusions:
- The developed DBS method is simple, robust, and suitable for pediatric blood sampling.
- This microsampling technique facilitates PK studies in children with minimal blood draw requirements.
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.
