Chaksawat Sangawitayakorn a , Pirom Chenprakhon b , Prapin Wilairat c , * , Rattikan Chantiwas a , *
a Department of Chemistry and Center of Excellence for Innovation in Chemistry, Faculty of Science, Mahidol University, Rama VI Rd., Bangkok 10400, Thailand
b Institute for Innovative Learning, Mahidol University, Nakhon Pathom 73170, Thailand
c Analytical Sciences and National Doping Test Institute, Mahidol University, Rama VI Rd., Bangkok 10400, Thailand
* Corresponding author.E-mail addresses: prapin.wil@mahidol.ac.th (P. Wilairat), rattikan.cha@mahidol.ac.th, rattikan.cha@mahidol.edu (R. Chantiwas).
Abstract:
Introduction: Phytochemical analysis of phenolic acids and flavonols poses a challenge, necessitating This study investigates the unexpected formation of ketoprofen methyl ester (KME) during the routine alkaline liquid–liquid extraction (LLE) process for analyzing basic drugs in horse urine samples using GC–MS analysis. An unidentified peak in the GC–MS chromatogram was observed in certain horse urine samples, identified as KME through mass spectral comparison with in situ synthesized KME. Since the KME is not a metabolite of ketoprofen present in urine, it is proposed that its formation occurs during the LLE process due to the reaction between ketoprofen contained in the urine and methanol used as the solvent of the spiked internal standard. Surprisingly, no artifact was detected when negative quality control horse urine samples (absence of ketoprofen) even when spiked with standard ketoprofen and methanol. Further investigation indicated that the presence of lipase enzymes from bacteria in specific urine samples is the key factor in the formation of the KME artifact. This hypothesis was confirmed when negative quality control horse urines were spiked with ketoprofen, methanol, and a lipase enzyme and the KME artifact was detected. Additionally, the formation of a methyl ester artifact was also detected for flunixin, a carboxylic acid NSAID drug, when negative quality control horse urines were spiked with the drug, methanol, and lipase enzyme. These findings will be valuable for scientists analyzing drugs in urine.
KEYWORDS
Artifacts, Ketoprofen methyl ester, Horse urine, Liquid–liquid extraction, GC–MS and Drug analysis
Reference:
Sangawitayakorn, C.; Chenprakhon, P.; Wilairat, P.; Chantiwas, R. Formation of ketoprofen methyl ester artifact in GC–MS analysis of basic drugs in horse urine using alkaline liquid–liquid extraction. Arabian Journal of Chemistry 2024, 17 (5), 105727.
doi: 10.1016/j.arabjc.2024.105727