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Published on: November 8, 2015
Development and Validation of a High-Performance Liquid Chromatography With Ultraviolet Detection Method to
Ola Ramadan1, Patrick Opitz, Georg Hempel
1Department of Pharmaceutical and Medical Chemistry, Clinical Pharmacy, University of Münster, Münster, Germany.
Background:
In neonatal and pediatric intensive care units, Gram -positive infections are a significant cause of morbidity and mortality. The increase in infections caused by methicillin-resistant Staphylococcus aureus and methicillin-resistant coagulase-negative Staphylococci have led to the increased use of glycopeptides, which treat invasive infections caused by Gram -positive organisms, particularly those resistant to beta-lactam antibiotics. Teicoplanin has bacteriostatic activity against Gram -positive bacteria, but its pharmacokinetics in children is highly variable, with most children failing to reach target levels at the recommended dose. This study aimed to develop a cost-effective method for determining concentrations using dried blood spot (DBS).
Methods:
A method to determine the concentrations of teicoplanin in 20 µL blood or plasma using the Whatman 903 Protein Saver filter was evaluated. High-performance liquid chromatography with ultraviolet detection high-performance liquid chromatography with ultraviolet/vis was used, with internal standard ketoconazole. In addition, a method to quantify teicoplanin using 50 µL of liquid plasma was established to compare the results with the values obtained by DBS and dried plasma methods.
Results:
The method was successfully developed and validated for 20 µL DBS. Furthermore, 50 µL of plasma was used to quantify teicoplanin with a lower limit of quantification of 10 mg/L. Precision and accuracy ranged from 2.3% to 10.7% and 95%-114.2%, respectively. A consistent factor (1.15) was used to calculate teicoplanin plasma concentrations from whole blood, indicating the reliability of the DBS method for therapeutic drug monitoring of teicoplanin.
Conclusions:
A simple, reliable, and cost-effective method using high-performance liquid chromatography with ultraviolet/vis was established to determine pediatric teicoplanin concentrations in both small plasma sample volumes and whole blood using DBS, and an accurate correlation factor for estimating teicoplanin plasma concentrations from DBS was identified. This method is suitable for the use in pediatrics.
Insights
A new dried blood spot (DBS) method accurately measures teicoplanin levels in children, improving therapeutic drug monitoring for Gram-positive infections. This cost-effective approach ensures optimal dosing for pediatric patients.
Area of Science:
- Pharmacology
- Clinical Chemistry
- Pediatric Infectious Diseases
Background:
- Gram-positive infections are a major cause of morbidity and mortality in pediatric intensive care units.
- Increased resistance to beta-lactam antibiotics necessitates the use of glycopeptides like teicoplanin.
- Teicoplanin pharmacokinetics in children are highly variable, often leading to sub-therapeutic levels with standard dosing.
Purpose of the Study:
- To develop a cost-effective method for determining teicoplanin concentrations in pediatric patients using dried blood spots (DBS).
- To validate a high-performance liquid chromatography (HPLC) method for teicoplanin quantification in small blood volumes.
- To establish a reliable correlation for estimating plasma teicoplanin concentrations from DBS samples.
Main Methods:
- Evaluation of a method for teicoplanin determination in 20 µL of whole blood or plasma using Whatman 903 Protein Saver filter paper.
- Utilized high-performance liquid chromatography with ultraviolet/vis detection (HPLC-UV/Vis) and ketoconazole as an internal standard.
- Established a parallel method for quantifying teicoplanin in 50 µL of liquid plasma for comparative analysis.
Main Results:
- Successfully developed and validated a method for quantifying teicoplanin in 20 µL DBS samples.
- Achieved a lower limit of quantification of 10 mg/L for teicoplanin in plasma.
- Demonstrated high precision (2.3%–10.7%) and accuracy (95%–114.2%), with a consistent correlation factor (1.15) for estimating plasma concentrations from DBS.
Conclusions:
- A simple, reliable, and cost-effective HPLC-UV/Vis method was established for pediatric teicoplanin concentration determination using DBS.
- The validated method allows accurate quantification from small plasma volumes and whole blood, suitable for therapeutic drug monitoring.
- An accurate correlation factor was identified, making DBS a practical tool for optimizing teicoplanin therapy in children.
