Abstract
Measurements of 3-hydroxyanthranilic acid and anthranilic acid in urine are described. The urine adjusted to pH 3.0 was saturated with NaCl, and 3-hydroxyanthranilic acid and anthranilic acid were extracted with diethyl ether. The method for 3-hydroxyanthranilic acid measurement employed a Shim-pack CLC-ODS (150 mm × 6.0 mm i.d.) column eluted with 100 mM potassium dihydrogenphosphate (pH 4.5)-acetonitrile (9:1, v/v) at a flow rate of 1.5 ml/min. The applied volage was set at +300mV vs. Ag/AgCl, the detection limit being 0.2pmol (30.62 pg) at a signal-to-noise ratio of 5:1. The method for anthranilic acid measurement employed a Chemcosorb 5-ODS-H (150 mm × 4.6 mm i.d.) column eluted with 80 mM potassium dihydrogenphosphate (pH adjusted to 3.0 by phosphoric acid)-acetonitrile (65:35, v/v) at a flow rate of 1.0 ml/min, with estimation at an excitation wavelength of 340 nm and an emission wavelength of 410 nm. The detection limit was 0.3pmol (41.13pg) at a signal-to-noise ratio of 5:1. These techniques were applied to the analyses of riboflavin-deficient rat urine. The total analysis times for 3-hydroxyanthranilic acid and anthranilic acid were ca. 12 min and 7 min, respectively.
