Free 3-methoxy-4-hydroxyphenylglycol determined in plasma by liquid chromatography with coulometric detection

We describe a simple "high-performance" liquid-chromatographic method for determining free 3-methoxy-4-hydroxyphenylglycol (MHPG) in plasma, with coulometric detection and with 4-methoxy-3-hydroxyphenylglycol (iso-MHPG) as the internal standard. MHPG and iso-MHPG are extracted from plasma into ethyl acetate and the extract is washed with a sodium bicarbonate solution, evaporated, reconstituted, and injected into a 25 x 0.4 cm column packed with 3-micron particles of C18 material. We used a mobile phase of acetate buffer (100 mmol/L, pH 5.0) and methanol (92:8 by vol) and an oxidation voltage of 0.5 V. The detection limit of the assay is 0.1 micrograms/L. The interassay CV for a sample with a mean MHPG concentration of 3.84 micrograms/L was 5.2%. The average absolute recovery for the method was 37%.