Fast determination of free metronidazole in rat plasma and peritoneal fluid using HPLC-UV method
Michele Vaz Dos Anjos1, Larissa Bergoza1, Gisele Fonseca1
1Universidade de Caxias do Sul.
Abstract:
Peritonitis refers to the inflammatory reaction of the peritoneum to aggression. In addition, it contributes significantly to sepsis. The presence of free concentrations of antimicrobials above the minimum inhibitory concentration at the site of infection is critical to therapeutic response. Metronidazole (MTZ) is an antimicrobial used to treat peritonitis because of its effectiveness against anaerobic microorganisms. This study investigates free MTZ concentrations in peritoneal microdialysate in Wistar rats. A C18 column (150 × 4.0 mm, 5 μm) was used for the analysis conducted at 40°C under isocratic conditions. The mobile phase consisted of acetonitrile and an aqueous solution of 50-mM monobasic phosphate buffer and 0.1% triethylamine, with pH 3.0 (10:90, v/v). MTZ calibration was linear in the range of 0.5-30.0 μg/ml. The intra- and inter-day precision was satisfactory with relative standard deviation ≤5.67%. The accuracy ranged from 90.64 to 103.77%, and the lower limit of quantification was 0.5 μg/ml. The developed method was successfully applied in a pilot pharmacokinetic study after MTZ administration (30 mg/kg, intravenously) in rats. The main advantage of the employed method is that it does not require sample processing and protein removal steps. This is the first study to be conducted using MTZ in rats.
Insights
This study developed a novel method to measure free metronidazole (MTZ) concentrations in peritoneal fluid. The findings are crucial for optimizing MTZ treatment in peritonitis and sepsis cases.
Area of Science:
- Pharmacology
- Analytical Chemistry
- Infectious Diseases
Background:
- Peritonitis, a peritoneal inflammation, significantly contributes to sepsis.
- Achieving adequate antimicrobial concentrations at the infection site is vital for treatment success.
- Metronidazole (MTZ) is effective against anaerobic bacteria implicated in peritonitis.
Purpose of the Study:
- To develop and validate a method for quantifying free metronidazole (MTZ) in rat peritoneal microdialysate.
- To investigate the pharmacokinetic profile of MTZ in the peritoneal cavity of Wistar rats.
- To establish a reliable analytical technique for assessing antimicrobial efficacy in peritonitis.
Main Methods:
- High-performance liquid chromatography (HPLC) with a C18 column was employed for MTZ analysis.
- The mobile phase consisted of acetonitrile and a phosphate buffer with triethylamine at pH 3.0.
- The method demonstrated linearity (0.5-30.0 μg/ml), good precision (RSD ≤ 5.67%), and accuracy (90.64-103.77%).
Main Results:
- A sensitive method with a lower limit of quantification of 0.5 μg/ml was successfully developed.
- The validated method was applied to a pilot pharmacokinetic study in Wistar rats.
- The analysis requires no sample processing or protein removal, simplifying the procedure.
Conclusions:
- A novel, efficient HPLC method for measuring free MTZ in peritoneal microdialysate was established.
- This method facilitates pharmacokinetic studies of MTZ in peritonitis models.
- The findings support optimizing MTZ dosing strategies for effective peritonitis and sepsis management.
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