A novel fully automated method for the measurement of sulphoconjugated catecholamines in urine using the Gilson ASTED-XL sample preparation system and high-performance liquid chromatography
✍ Scribed by G. R. Bayly; W. A. Bartlett; L. V. Harper; A. F. Jones
- Publisher
- John Wiley and Sons
- Year
- 1999
- Tongue
- English
- Weight
- 51 KB
- Volume
- 13
- Category
- Article
- ISSN
- 0269-3879
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✦ Synopsis
Dopamine is produced in the kidney where it causes sodium excretion. Dopamine sulphate is deconjugated in vivo, and may be a physiological reservoir for this active renal dopamine. To investigate the role of dopamine and dopamine sulphate in sodium homeostasis we have developed a fully automated HPLC assay for free, total and sulphoconjugated dopamine. Using the Gilson ASTED-XL sample preparation unit, with temperature controlled racks, urinary free and total dopamine are measured pre-and postincubation with arylsulphatase. Dopamine sulphate is calculated from the difference between free and total measurements. Acidified 24 h urines are processed automatically. Free dopamine assay follows buffering to pH 7.0, addition of internal standard, addition of EDTA to stabilize free catecholamines at neutral pH, and incubation at 37°C for 30 min. This mixture is trace enriched on a HEMA-SB TEC prior to ion-paired HPLC with amperometric detection. To measure total dopamine the entire process is automatically repeated with addition of arylsulphatase (400 mU/mL urine) at the beginning of the 37°C incubation. The working range of the assay is up to 7 mmol/L total dopamine. Within-and between-run imprecision for dopamine sulphate is less than 3 and 7% respectively. Median dopamine sulphate excretion in 12 normotensive subjects was 4.3 mmol/24 h.