A rapid liquid chromatography-tandem mass spectrometry method for the analysis of urinary 5-hydroxyindoleacetic acid (5-HIAA) that incorporates a 13C-labelled internal standard. (November 2021)

• Record Type: Journal Article
• Authors: Grant, Kirsten M; Livie, Craig; Smith, Karen; Leung, Chui Ha; Johnston, Susan
• Is Part Of: Annals of clinical biochemistry. Volume 58:Number 6(2021)
• Page Start: 622
• Page End: 631
• Publication Date: 2021-11
• Subjects: Mass spectrometry -- laboratory methods -- chromatography -- laboratory methods -- tumour markers -- analytes; Clinical chemistry -- Periodicals; Clinical biochemistry -- Periodicals; 616.075
• DOI: 10.1177/00045632211038021
• Languages: English
• ISSNs: 0004-5632

Abstract

Background: Urinary 5-hydroxyindoleacetic acid (5-HIAA) is a first-line investigation for gastrointestinal neuroendocrine tumours that secrete serotonin. It also has clinical utility for monitoring disease progression and therapeutic response. Aim: To develop and validate a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for urinary 5-hydroxyindoleacetic acid that incorporates a supported liquid extraction and 13 C-labelled internal standard. Methods: Samples were diluted in ammonium acetate containing a 13 C-labelled internal standard (5-hydroxyindole-3a, 4, 5, 6, 7, 7a- 13 C6 -3-acetic acid). Supported liquid extraction was performed followed by chromatographic separation using the 2.1 × 30 mm CORTECS® UPLC® T3 column. Mass spectrometry detection (Waters Xevo TQ-XS) was performed in electrospray positive mode using the transitions 192.3 > 146.4 m/z (quantifier) and 192.3 > 118.4 m/z (qualifier) for 5-hydroxyindoleacetic acid and 198.2 > 152.4 m/z for 13 C-5-HIAA. Results: A well-defined 5-hydroxyindoleacetic acid peak was observed at 0.8 min with a run time of 2.4 min. The assay was linear (r 2 > 0.99) to 382 µ mol/L, with a lower limit of quantification of 5.3 µ mol/L (CV <15%). Analysis of 29 external quality assurance samples showed good agreement between our method and the UKNEQAS method mean (4.7% positive bias). The intra- and inter-assay precision was within acceptable limits, and the assay was stable up to 96 h postextraction with minimal carryover. Conclusion: We have developed a robust LC-MS/MS method with semi-automated extraction that offers an improved run time and performance over the existing, labour-intensive, HPLC method. The method was quick, precise, showed good agreement with UKNEQAS external quality assurance material and is in routine service for clinical samples.