title: Urinary 5-HIAA measurement using automated on-line solid-phase extraction-high-performance liquid chromatography-tandem mass spectrometry.
abstract: Quantification of 5-hydroxyindole-3-acetic acid (5-HIAA) in urine is useful for diagnosis and follow-up of patients with carcinoid tumors and for monitoring serotonin (5-hydroxytryptamine) metabolism in various disorders. We describe an automated method (XLC-MS/MS) that incorporates on-line solid-phase extraction (SPE), high-performance liquid chromatography (HPLC) and tandem mass spectrometric (MS/MS) detection to measure urinary 5-HIAA. Automated pre-purification of urine was carried out with HySphere-Resin GP SPE cartridges containing strong hydrophobic polystyrene resin. The analyte (5-HIAA) and internal standard (isotope-labelled 5-HIAA-d(2)) were, after elution from the cartridge, separated by reversed-phase HPLC and detected with tandem MS. Total cycle time was 5 min. 5-HIAA and its deuterated internal standard (5-HIAA-d(2)) were retained on and eluted from the SPE cartridges in high yields (81.5-98.0%). Absolute recovery was 96.5-99.6%. Intra-assay (n=20) and inter-assay (n=20) CVs for the measurement of 5-HIAA in urine in three concentration levels ranged from 0.8 to 1.4% and 1.7 to 4.2%, respectively. For urine samples from patients (n=78) with known or suspected metastatic carcinoid tumors, results obtained by XLC-MS/MS were highly correlated (R(2)=0.99) with the routinely used fluorometric method. This XLC-MS/MS method demonstrated lower imprecision and time per analysis (high-throughput) than manual solvent extraction methods and higher sensitivity and specificity than non-mass spectrometric detection techniques.
authors.affiliation: Department of Laboratory Medicine, University Medical Center, Groningen, University of Groningen, Groningen, The Netherlands.
authors.forename: Wilhelmina H A
authors.initials: WH
authors.lastname: de Jong
authors.forename: Kendon S
authors.initials: KS
authors.lastname: Graham
authors.forename: Elisabeth G E
authors.initials: EG
authors.lastname: de Vries
authors.forename: Ido P
authors.initials: IP
authors.lastname: Kema
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D006897:Hydroxyindoleacetic Acid / Q000652:urine*; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D006897:Hydroxyindoleacetic Acid