Validated HPLC-MS-MS method for determination of azithromycin in human plasma

Area of Science:

• Analytical Chemistry
• Pharmacokinetics
• Biomedical Science

Background:

• Accurate quantification of azithromycin (AZI) in biological matrices is essential for pharmacokinetic studies.
• Existing methods may lack the required sensitivity, selectivity, or efficiency for routine analysis.

Purpose of the Study:

• To develop and validate a highly sensitive and selective High-Performance Liquid Chromatography tandem Mass Spectrometry (HPLC-MS-MS) method for determining azithromycin concentrations in human plasma.
• To apply the validated method for assessing the pharmacokinetic profile of azithromycin.

Main Methods:

• Quantitative determination of azithromycin in human Na2EDTA plasma using High-Performance Liquid Chromatography coupled with tandem Mass Spectrometry (HPLC-MS-MS).
• Roxithromycin (ROX) served as the internal standard.
• Ultrafiltration using Centrifree Micropartition devices followed by isocratic HPLC-MS-MS analysis in Multiple Reaction Monitoring (MRM) mode.

Main Results:

• The developed HPLC-MS-MS method demonstrated excellent linearity over a concentration range of 2.55-551.43 ng/mL.
• Both inter-day and intra-day precision and accuracy were found to be less than 8% (R.S.D. and percentage deviation).
• The limit of quantification (LOQ) was established at 2.55 ng/mL, indicating high sensitivity.

Conclusions:

• A robust, sensitive, and selective HPLC-MS-MS method for azithromycin quantification in human plasma has been successfully developed and validated.
• The method is suitable for pharmacokinetic studies, enabling accurate assessment of drug disposition.
• This validated assay provides a reliable tool for therapeutic drug monitoring and clinical research involving azithromycin.