Technical and Biological Variation in UPLC−MS-Based Untargeted Metabolic Profiling of Liver Extracts: Application in an Experimental Toxicity Study on Galactosamine

摘要

ABSTRACT: The relative importance of technical versus bio-nlogical variation inUPLC-MS livermetabolic profiling studies wasnassessed on liver samples collected as part of an in vivo hepato-ntoxicity study. Biological variability within and between two treat-nment groups (three rats treated with galactosamine and three withngalactosamineþuridine) was compared with sampling/extractionnvariability (three portions extracted fromeach rat liver section) andnUPLC-MS platform variability (triplicate injections of eachnextract) for aqueous and organic extracts. The impact of scalingnon error measurement was investigated on replicate injections of a quality control sample, and consequently started log-transformation wasnused to stabilize the variance across the ion intensity range. For aqueous extracts, technical variability was two to four times lower than withinngroup interanimal variability. Similar results were obtained for organic extracts for the galactosamine group, sampling/extraction variabilitynbeing more elevated in the galactosamineþuridine group. For both extract types, differences between treatment groups were the principalnsource of observed variation, and triplicate injections clustered closely in PCA plots and in HCA dendrograms, indicating small instrumentnvariability compared to observed biological variation. This protocol can be applied to investigate differences in liver metabolic profilesnbetween animal groups in toxicology studies and clinical investigations of liver disease.