Pattern Recognition Analysis for Hepatotoxicity Induced by Acetaminophen Using Plasma and Urinary ¹H NMR-Based Metabolomics in Humans

Abstract

Drug-induced liver injury (DILI) is currently an increasingly relevant health issue. However, available biomarkers do not reliably detect or quantify DILI risk. Therefore, the purpose of this study was to comparatively evaluate plasma and urinary biomarkers obtained from humans treated with acetaminophen (APAP) using a metabolomics approach and a proton nuclear magnetic resonance (NMR) platform. APAP (3 g/day, two 500 mg tablets every 8 h) was administered to 20 healthy Korean males (age, 20-29 years) for 7 days. Urine was collected daily before and during dosing and 6 days after the final dose. NMR spectra of these urine samples were analyzed using principal component analysis (PCA) and partial least-squares-discrimination analysis. Although the activities of aspartate aminotransferase and lactate dehydrogenase were significantly increased 7 days post-APAP treatment, serum biochemical parameters of aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, total bilirubin, γ-glutamyl transpeptidase, and lactate dehydrogenase were within normal range of hepatic function. However, urine and plasma (1)H NMR spectroscopy revealed different clustering between predosing and after APAP treatment for global metabolomic profiling through PCA. Urinary endogenous metabolites of trimethylamine-N-oxide, citrate, 3-chlorotyrosine, phenylalanine, glycine, hippurate, and glutarate as well as plasma endogenous metabolites such as lactate, glucose, 3-hydroxyisovalerate, isoleucine, acetylglycine, acetone, acetate, glutamine, ethanol, and isobutyrate responded significantly to APAP dosing in humans. Urinary and plasma endogenous metabolites were more sensitive than serum biochemical parameters. These results might be applied to predict or screen potential hepatotoxicity caused by other drugs using urinary and plasma (1)H NMR analyses.

Dynamic analysis of the endogenous metabolites in depressed patients treated with TCM formula Xiaoyaosan using urinary 1H NMR-based metabolomics

“…For NMR analysis, urine samples were thawed at 0 1C and then centrifuged at 3000 rpm for 15 min. A 500-μL sample of urine was mixed with 200 μL of phosphate buffer in D 2 O (0.2 M Na 2 HPO 4 Á 12H 2 O/0.2 M NaH 2 PO 4 Á 2H 2 O, 81:19, v/v, pH 7.4), containing 0.015% TSP (used as chemical shift reference) (Carrola et al, 2010;Kim et al, 2013). After centrifugation at 13,000 rpm for 20 min, 600 μL of supernatant was transferred into 5-mm NMR tubes.…”

Method Validation for Preparing Urine Samples for Downstream Proteomic and Metabolomic Applications

“…Urine is proving to be an increasingly useful source of metabolites and protein biomarkers for detecting disease states in both adults and neonates, [4][5][6][7] as well as the effects of drugs and dietary intake on physiologic processes. [8][9][10] Best Practices 11 and Standard Operating Procedures [12][13][14][15][16] for routine processing and storing of biobank samples have been published.…”

Biomarkers in DILI: One More Step Forward

“…Metabolic profiling in urine has also been performed in search for metabolites associated with DILI. Healthy volunteers administered non-toxic levels of APAP presented both urine and plasma metabolic alterations when comparing pre- and post-dosing samples, which were more sensitive than changes in serum biochemical parameter over the study period (Kim et al, 2013). It should be pointed out that while many metabolic studies to date coincide in detecting variations in endogenous metabolites, the identified metabolites often vary between studies.…”