Simultaneous liquid-chromatographic analysis for 4-hydroxy-3-methoxymandelic acid and 4-hydroxy-3-methoxyphenylacetic...

We describe a rapid, reliable "high-performance" liquid-chromatographic method of simultaneously analyzing for 4-hydroxy-3-methoxymandelic acid (I) and 4-hydroxy-3-methoxyphenylacetic acid (II) in urine. Paired-ion chromatography and amperometric detection are used in the method. A 5-mL aliquot of urine is adjusted to pH 6.1 and subjected to anion-exchange chromatography (Dowex AG 1-X4, 1.8 X 0.7 cm column). The two analytes are eluted from the column with 10 mL of 3 mol/L sodium chloride, and 2 microL of the eluate, injected directly into the liquid chromatograph, is chromatographed at 40 degrees C. This column contains mu Bondapak C18, and the mobile phase is 100 mmol/L potassium phosphate buffer, pH 6.7, containing 0.5 mmol of tetrabutylammonium phosphate per liter. Detector sensitivity is 50 nA full scale. Analytical recoveries of I and II are 80--84% and 48--49%, respectively; the coefficients of variation for various concentrations in urine are about 10%.