LAUSR

BACKGROUND Urinary 5-hydroxyindoleacetic acid (5-HIAA) is a first line investigation for gastrointestinal neuroendocrine tumours (NETs) 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-HIAA that incorporates a supported liquid extraction (SLE) and 13C labelled internal standard. METHODS Samples were diluted in ammonium acetate containing a 13C labelled internal standard (5-Hydroxyindole-3a,4,5,6,7,7a-13C6-3-acetic acid). SLE was performed followed by chromatographic separation using the 2.1 x 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-HIAA and 198.2 > 152.4 m/z for 13C-5-HIAA. RESULTS A well-defined 5-HIAA peak was observed at 0.8 min with a run time of 2.4 mins. The assay was linear (r2 >0.99) to 382 µmol/L, with a lower limit of quantification of 5.3 µmol/L (CV <15%). Analysis of 29 external quality assurance (EQA) 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 hrs post-extraction 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 EQA material and is in routine service for clinical samples.