Development and validation of a novel 7α-hydroxy-4-cholesten-3-one (C4) liquid chromatography tandem mass spectrometry method and its utility to assess pre-analytical stability.
Clin Chem Lab Med
; 2024 Aug 05.
Article
in En
| MEDLINE
| ID: mdl-39097844
ABSTRACT
OBJECTIVES:
7α-Hydroxy-4-cholesten-3-one (C4) is the common intermediary of both primary bile acids. C4 is recommended by the British Society of Gastroenterology for the investigation of bile acid diarrhoea (BAD) in patients with chronic diarrhoea. This project aimed to develop and validate an assay to quantitate C4 in serum and assess the stability of C4 in unseparated blood.METHODS:
Accuracy was underpinned by calibrating to quantitative nuclear magnetic resonance analysis. C4 was analysed in a 96-well plate format with a deuterated C4 internal standard and liquid-liquid extraction. Validation followed the 2018 Food and Drug Administration guidelines. To assess C4 stability, healthy volunteers (n=12) donated 8 fasted samples each. Samples were incubated at 20⯰C for up to 72â¯h and retrieved, centrifuged, aliquoted and frozen for storage at different time points prior to C4 analysis.RESULTS:
The C4 method demonstrated excellent analytical performance and passed all validation criteria. The method was found to be accurate, precise, free from matrix effects and interference. After 72â¯h of delayed sample separation, C4 concentration gradually declined by up to 14â¯% from baseline. However, the change was not significant for up to 12â¯h.CONCLUSIONS:
We present a robust method of analysing serum C4, offering a convenient alternative to 75SeHCAT for BAD investigation. C4 was found to decline in unseparated blood over time; however, after 12â¯h the mean change was <5â¯% from baseline. Our results suggest C4 is suitable for collection from both primary and secondary care prior to gastroenterology referral.
Full text:
1
Database:
MEDLINE
Language:
En
Journal:
Clin Chem Lab Med
Journal subject:
QUIMICA CLINICA
/
TECNICAS E PROCEDIMENTOS DE LABORATORIO
Year:
2024
Type:
Article
Affiliation country:
United kingdom