Optimization and validation of RP-HPLC method for quantification of piperacillin in pharmaceutical formulation and in human blood plasma

Abstract A broad-spectrum antibiotic known as piperacillin is often used with tazobactam to treat infections caused by anaerobic bacteria as well as different Gram-positive and Gram-negative bacteria. To ensure efficacy, it is important that piperacillin must reach therapeutic levels in the blood. An optimized, high-pressure liquid chromatography (HPLC) serves as an ideal technique for accurate drug concentration analysis. In the current study, a validated reverse-phase HPLC method was developed for the determination of piperacillin in pharmaceutical formulation and human plasma. Acetonitrile was used to deproteinized the plasma samples, vortexed them, and reconstituted the supernatant with the mobile phase prior to HPLC injection. The analysis was performed by using a Waters Alliance HPLC system, which had a UV detector set at 218 nm and a flow rate of 1 mL min −1 . The mobile phase consisted of methanol, water, tetrabutylammonium chloride buffer, and phosphoric acid. The method showed linearity from 0.5 μg mL −1 to 400 μg mL −1 , with 99.9% recovery rate. The detection and quantification limits were 0.25 μg mL −1 and 0.5 μg mL −1 , respectively. The Cmax exhibits variability among patients, even when the same dose is administered to adult individuals. The method is both accurate and reliable, and the findings highlights the importance of individualized dosing based on variations in patients' plasma concentrations.