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.
|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|
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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