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Urinary 5_HIAA measurement using automated on-line solid-phase extraction-high-performance liquid chromatography- tandem mass spectrometry

Summary 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-d2) 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-d2) 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 (R2 = 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.
Category 3. Symbiosis User Application
These are applications that have been made by Symbiosis customers. They are validated and have been published.
Matrix Urine
Market Clinical
Cartridge HySphere Resin GP
Detection MS (Waters Quattro Premier XE)
Compounds 5-HIAA, 5-Hydroxyindole-3-acetic acid, 5-HT, 5-Hydroxytryptamine
Compound Group Serotonin, Serotonin metabolite
Sample_volume 50 ÁL (2.5ÁL of urine)
Contact Wilhelmina (Helma) H.A. de Jong, Ido P. Kema, Kendon S. Graham (Waters)
Account Department of laboratory medicine, UMCG, Groningen Waters Corporation, Manchester, UK
Published J. of Cromatography B, 2008
Date 8-8-2008
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Remarks The total cycle time 5 min. 5-HIAA and I.S. were retained with on the cartridge a high yield of 81.5 - 98%.
Quantification of 5-HIAA in urine is useful for the diagnosis and follow-up of patient with carcinoid tumors and for monitoring the Serotonin metabolism in various disorders
Users UMCG
Status Finished