Precision Meets Affordability: A Highly Sensitive HPLC-FLD Technique for Accurate Pitavastatin Quantification in Human Plasma †

Abstract

High-Performance Liquid Chromatography (HPLC) combined with ultraviolet/visible (UV/Vis) or diode array detection (DAD) is routinely used for drug quantification in R&D around the world. However, it may lack the sensitivity required for bioanalytical studies. On the other hand, HPLC with fluorescence detection (FLD) is a cost-effective alternative that significantly increases drug signals, enabling the detection of compounds at very low concentrations. Pitavastatin is a lipid-lowering drug that contains the structure of quinoline, a highly fluorescent molecule. Recently, it has gained interest due to its pleiotropic effects on different conditions. Bearing this in mind, an HPLC-FLD method was developed and validated for the quantification of pitavastatin in human plasma. Overall, a signal gain of 54–70 times compared to that of UV detection was achieved when using fluorescence. Sample preparation included one-step protein precipitation with acetonitrile, followed by centrifugation and filtration prior to injection. Pitavastatin was separated from endogenous matrix interferents using a C18 column and by applying gradient elution. Atorvastatin was used as an internal standard. Accordingly, the method was shown to be selective, specific, and sensitive, with a lower limit of quantification of 3 ng/mL and complete absolute and relative recoveries higher than 94%. The method was linear over the wide concentration range of 3–900 ng/mL (R2 = 0.998), accurate (bias < 7.15%), and precise (RSD < 9.63%). This method allows for the therapeutic monitoring of patients treated with pitavastatin but can also support novel clinical studies of this drug in human plasma.