Optimization and Validation of a Capillary MEKC Method for Determination of Proteins in Urine

Abstract

Proteinuria, i.e. increased excretion of proteins in urine, is a common sign indicating renal or urinary tract diseases. In this study, a fast and simple procedure for urine sample preparation and capillary micellar electrokinetic chromatographic analysis is presented, without any sample pretreatment prior to the analysis. The developed MEKC method was employed for simultaneous determination of albumin (ALB), haemoglobin (HGB), and myoglobin (MYO) in human urine samples obtained from patients with diagnosed proteinuria. Optimum conditions for detection and separation of ALB, HGB, and MYO are 50 mmol L−1 borate buffer containing 20 mmol L−1 SDS (pH 9.3), injection 40 mbar × 20 s, voltage 25 kV, temperature 30 °C, and detection wavelength 200 nm. The method was shown to be specific, accurate, linear (correlation coefficients r2 > 0.99), and precise (RSD below 3.75 and 7.23% for migration time and peak area, respectively). Multi-variable-at-a-time (MVAT) approach for robustness testing shows no significant variations in accuracy, specificity, and precision as RSD values were lower than 5 and 10% for migration time and peak area, respectively. The presented method is applicable for routine analyses of urine samples as a screening method for patients with excess ALB, HGB, and MYO.