The differentiation of human induced pluripotent stem cells (hiPSCs) into functional hepatocytes has the potential to solve the shortage of human primary liver cells and would be of use in drug screening. In this frame, we developed a hiPSCs maturation strategy in microfluidic biochips, using a liver-on-chip approach, a promising technology mimicking in vivo physiology. Hepato-like tissues differentiated in biochips presented advanced liver features, including ALB and CYP3A4 expressing cells. The metabolomics and transcriptomics profiles of hepato-likes cells differentiated either in biochips or Petri dishes were integrated to compare their functionalities. The multi-omics analysis revealed 41 metabolites and 302 genes differentially expressed. Overall, biochip environment lead to higher degree of hepatic differentiation demonstrated by an increase in the metabolic production of lipids, fatty acids and biliary acids …