Determination of 3-methoxy-4-hydroxyphenylethylene glycol in urine using reversed-phase liquid chromatography with...

A column-switching method was developed for the determination of total 3-methoxy-4-hydroxy-phenylethyleneglycol (MHPG) in urine. This was performed by first treating samples with beta-glucuronidase, followed by extraction with ethyl acetate. The reconstituted extracts were injected onto an HPLC system containing an amperometric detector and tandem Nucleosil C18 and C8 reversed-phase columns connected by a switching valve. The total analysis time for MHPG was 12 min. The limit of detection was 0.18 ng, or 9 micrograms/l for 20-microliters injections of a 1.0-ml reconstituted extract prepared from 1.0 ml of urine. The linear range extended up to 80 mg/l. The within-day precision for a urine sample containing 170 micrograms/l total MHPG was +/- 6% and the day-to-day precision was +/- 15%. The average levels determined by this method for total MHPG in normal subjects showed good agreement with previous literature values. This approach could be modified for the determination of free MHPG by using only ethyl acetate extraction for sample pretreatment.