Ion pair high-performance liquid chromatographic assay for ceftriaxone

Area of Science:

• Analytical Chemistry
• Pharmacokinetics

Background:

• Ceftriaxone, a third-generation cephalosporin, requires accurate quantification in biological fluids for therapeutic drug monitoring.
• Standard reverse-phase HPLC methods often struggle with polar compounds like ceftriaxone, leading to poor retention and peak tailing.

Purpose of the Study:

• To develop and validate a high-performance liquid chromatographic (HPLC) assay for the precise measurement of ceftriaxone in serum, urine, and cerebrospinal fluid.
• To address the challenges of analyzing polar ceftriaxone using ion-pairing chromatography.

Main Methods:

• Development of an ion-pairing HPLC assay using a mobile phase of acetonitrile and water (pH 9.0) with hexadecyltrimethylammonium bromide.
• Utilized moxalactam as an internal standard and a silica-packed precolumn to protect the analytical column.
• Validated linearity, correlation with bioassay, reproducibility, and recovery.

Main Results:

• The assay demonstrated excellent linearity (r = 0.999) over a concentration range of 1 to 250 µg/ml.
• High correlation (r = 0.990) was observed with an agar diffusion bioassay.
• Good reproducibility (intrarun CVs 2.3–6.4%, interrun CVs 3.2–21.4%) and high absolute recoveries (91–97%) were achieved.
• No interference was detected from over 40 common drugs, including other cephalosporins, or from patient sera with renal or hepatic disease.

Conclusions:

• The developed ion-pairing HPLC assay is a robust, accurate, and reproducible method for quantifying ceftriaxone in various biological matrices.
• This validated assay is suitable for clinical applications, including therapeutic drug monitoring and pharmacokinetic studies.
• The use of a precolumn effectively prolonged the analytical column's lifespan under alkaline conditions.