A simplified routine assay for urinary 3-methoxy-4-hydroxyphenylglycol

Abstract

Urinary 3-methoxy-4-hydroxyphenylglycol (MHPG) level may provide us with a biochemical index for central noradrenergic activity. Previous methods for assaying MHPG in urine often lacked specificity, sensitivity, cost-effectiveness or convenience. We now describe a simplified routine assay for urinary MHPG by high-pressure liquid chromatography using electrochemical detection. For convenience and cost-effectiveness within a typical batch assay of total MHPG, 0.5 mL of urine sample, 50 microL of glusulase and 4 micrograms iso-vanillyl alcohol (internal standard) were used to hydrolyze conjugated MHPG at 37 degrees C overnight. Alternatively, for a same-day operation, it is efficient to hydrolyze the sample at 50 degrees C for 3 hours. Each sample was separated isocratically on a reversed-phase column (Ultracarb 5 ODS) at 25 degrees C with the flow rate at 1 mL/min. Intra- and inter-assay coefficients of variation were found to be 4.0% (n = 10) and 5.0% (n = 27), respectively, for MHPG at a mean concentration of 1.9 mg/L. Sulfatase or glucuronidase can be substituted for glusulase to obtain either sulfate- or glucuronide-conjugated MHPG. This procedure requires smaller sample amounts and less preparation time without compromising sensitivity and reproducibility.