Ultrafiltration-based sample preparation and HPLC-UV determination of diclofenac in human plasma samples

Authors: MERVE NENNİ, AYŞEGÜL DOĞAN, MUSTAFA ÇELEBİER, MURAT SOYSEVEN, MUSTAFA SİNAN KAYNAK, HASSAN Y. ABOUL-ENEIN, GÖKSEL ARLİ

Abstract: The sample preparation step is the initial step in pharmaceutical analysis. While ultrafiltration is a well-known technique used in the food and pharmaceutical industries, it has rarely been used to measure the plasma concentration of active pharmaceutical ingredients. This study aimed to analyze diclofenac sodium (DS) in human plasma samples using ultrafiltration-based sample preparation before high-performance liquid chromatography (HPLC) analysis. The advantages and limitations of ultrafiltration-based sample preparation in bioanalysis were evaluated by comparing the results with conventional methods. The precipitating agent was used before ultrafiltration. The analysis was carried on an HPLC-UV system with a C18 column (250 ×4.6 mm, 5 μm) and acetonitrile : phosphate buffer (pH 3.0, 10 mM) (70 : 30 v/v) was used as the mobile phase. The bioanalytical method was validated according to FDA guidelines and applied to spiked samples of DS in commercial human plasma samples. The LOD and LOQ values were 0.006 μg mL^-1 and 0.020 μg mL^-1, respectively. The method was linear in the range of 0.025-0.50 μgmL^-1 with excellent determination coefficients (R² > 0.9991). The findings of this analysis with low LOD and LOQ values and high recovery values with high trueness and precision proved the matrix minimizing the effect of the presented sample preparation technique.

Keywords: Centrifugal filters, diclofenac, high-performance liquid chromatography, HPLC, plasma, ultrafiltration

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