Acetaminophen is the most used analgesic and anti-pyretic drug in the world, but also one of the main sources of drug induced liver injury (DILI). Aminotransferase blood levels, the actual golden-standard for the assessment of liver injury cannot be detected before the onset of liver injury and are poor predictors of disease outcome. Thus, new biomarkers of acetaminophen-induced liver injury are sorely needed and small circulating microRNAs were demonstrated to have a very interesting potential. In this study, we tested and implemented a microRNA detection protocol in a cohort of hospitalized patient receiving a therapeutic 4g/ day dose of acetaminophen. We found the classic phenol-chloroform extraction method to remain the most effective nucleic acid purification procedure but would recommend a more user-friendly column-based protocol for extraction of a large number of samples. During the optimization and validation steps of the procedure, we also confirmed the importance of sample hemolysis for microRNA quantification. We followed evolution of the potential liver injury marker miR122 in the plasma samples of two patients of the CER 13-265 study and observed a stepper increase of miR122 over ALAT / ASAT values over time, suggesting that miR122 could be a more convenient but not necessarily earlier indicator of liver fonction impairment than the classic liver test measurements. We also tested different plasma fraction for enrichment of miR122 and, interestingly, our results suggest different tendencies for exosomal and microparticle preparations. Finally, on-going preparations for a large scale, parallel transcriptomic and proteomic approach for new early acetaminophen-induced hepatotoxcity biomarker discovery experiments are described.