Ann Lab Med 2018; 38(3): 255-260  https://doi.org/10.3343/alm.2018.38.3.255
Evaluation of Stability of Thiopurine Metabolites Using a Validated LC-MS/MS Method
In-Young Yoo, M.D.1, Kyunghoon Lee, M.D.1, Ok-Ja Ji, M.T.1, Hye In Woo, M.D.2, and Soo-Youn Lee, M.D.1
Department of Laboratory Medicine and Genetics1, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul; Department of Laboratory Medicine2, Samsung Changwon Hospital, Sungkyunkwan University School of Medicine, Changwon, Korea
Corresponding author: Soo-Youn Lee
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-ro, Gangnam-gu, Seoul 06351, Korea
Tel: +82-2-3410-1834
Fax: +82-2-3410-2719
E-mail: suddenbz@skku.edu
Received: August 10, 2017; Revised: October 24, 2017; Accepted: January 5, 2018; Published online: May 1, 2018.
© Korean Society for Laboratory Medicine. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Measurement of thiopurine metabolites is helpful to monitor adverse effects and assess compliance in patients on thiopurine treatment. The purpose of this study was to develop and validate an analytical method for measurement of thiopurine metabolites, thioguanine nucleotides (6-TGN) and 6-methylmercaptopurine nucleotide (6-MMPN), in RBCs. We developed and validated a liquid chromatography–tandem mass spectrometry (LC-MS/MS) method for the quantification of 6-TGN and 6-MMPN and evaluated the stability of the thiopurine metabolites in RBC and whole blood states without any preprocessing at various storage conditions. The linear range was 0.1–10 μmol/L and 0.5–100 μmol/L for 6-TGN and 6-MMPN, respectively. The mean extraction recovery at the two concentrations was 71.0% and 75.0% for 6-TGN, and 102.2% and 96.4% for 6-MMPN. Thiopurine metabolites in preprocessed RBC samples were stable at 25°C and 4°C after storage for 4 hours and at –70°C for up to 6 months. However, 6-TGN decreased by 30% compared with the initial concentration when stored at –20°C for 180 days. In whole blood states, 6-TGN decreased by about 20% at four days after storage at 4°C. We validated a reliable LC-MS/MS method and recommend that the patient’s whole blood sample be preprocessed as soon as possible.
Keywords: Thiopurine metabolites, 6-Thioguanine nucleotide, 6-Methylmercaptopurine nucleotide, Mass spectrometry, Stability



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