A Copper Determination Method Based on the Reaction between 2-(5-Nitro-2-Pyridylazo)-5-(N-Propyl-N- Sulfopropylamino) Phenol (Nitro-PAPS) and Copper for Use in Urine Copper Measurement and Application to Automation

Authors: OSMAN METİN İPÇİOĞLU, ÖMER ÖZCAN, MUSTAFA GÜLTEPE

Abstract: Aim: To develop a sensitive automated colorimetric urine copper determination method using a water soluble compound, 2-(5-nitro-2-pyridylazo)-5-(N-propyl-N-sulfopropylamino) phenol (nitro-PAPS), as a ligand. Materials and Methods: The new photometric method using sodium dodecyl sulfate (SDS)-ascorbic acid to dissociate copper from proteins in acetate buffer (pH 3.2) and nitro-PAPS as ligand was also adapted to an automated analyzer. Copper concentrations were determined by atomic absorption spectrophotometric method and by photometric method in 24 h urine samples of patients (n = 100) with various types of diseases including Wilson's disease (n = 12), diabetes mellitus (n = 34), osteoporosis (n = 27), nephrotic syndrome (n = 17), and rheumatoid arthritis (n = 10). Results: There was a statistically significant correlation (P < 0.01) between our new method and the atomic absorption spectrophotometric method (y = 0.994x + 0.207, Sy/x = 1.776), and the photometric method was linear up to 200 µg/l concentration. Other complexing metals (Zn^{+2} and Fe^{+2}) had no interfering effect on the Cu-nitro-PAPS reaction. The analytical recovery of copper was between 90% and 107% (mean 98%). Within-run and between-run CVs were <5% in normal and high copper containing urine pools. Conclusions: We have developed a new sensitive and reliable automated colorimetric urine copper determination method.

Keywords: Urine copper, photometric method, nitro-PAPS

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