Fluorimetric determination of phytic acid in urine based on replacement reaction

Abstract

A sensitive fluorimetric method for determination of phytic acid in human urine samples was described. The method was based on a fluorimetric replacement reaction, in which the added phytic acid replaced the Cu 2+ ion from Cu 2+–gelatin complex, liberating the fluorescent gelatin molecule. The fluorescence of the solution was accordingly recovered proportionally to the amount of the foreign phytic acid. The excitation wavelength was 273.5 nm and the characteristic emission wavelength was 305.0 nm, respectively. The calibration graph was obtained by plotting the recovered fluorescent intensity at maximum 305.0 nm against the added standard phytic acid, and was divided into two sections. One section was linear over the range of 0.40–2.40 mg L −1 with a linear regression equation of I f = −0.895 + 15.146 c ( R 2 > 0.9993), and the other over the range of 2.40–9.20 mg L −1 with a linear regression equation of I f = −29.526 + 26.113 c ( R 2 > 0.9996), respectively. The relative standard deviation (R.S.D.) at 95% confidence degree for a 2.0 mg L −1 of standard phytic acid within 1 month was less than 1.26% ( n = 5), indicating the procedure is reproducible. The detection and the quantification limits of phytic acid were estimated to be 0.23 and 0.40 mg L −1, respectively. The proposed method was applied to the determination of phytic acid in urine samples and the found concentrations of phytic acid in urine were in the range of 0.49–0.75 mg L −1 with recoveries of 96.2–108.8%. Comparison of the obtained results with the reported HPLC was performed, indicating the proposed method was reliable.