Cytochrome P450s (CYPs) are involved in the metabolism of various drugs, and may generate toxic metabolites or intermediates that result in drug-induced liver injury (DILI). Consequently, inducers of CYPs may promote DILI. In a draft test guideline, the Organisation for Economic Co-operation and Development (OECD) recommends measurement of the metabolic activity of CYP as an index for assessing CYP-inducing activity. However, change of mRNA level has also been used as a simple parameter to evaluate CYP induction. In this study, therefore, we examined the usefulness and limitations of mRNA expression measurement for evaluation of the induction of CYP1A2, CYP2B6, and CYP3A4 by omeprazole, phenobarbital, and rifampicin (RIF), respectively, in HepaRG cells, a well-established cell line derived from human hepatocellular carcinoma. The results of mRNA measurement correlated well with the results of metabolic activity measurement in the lower concentration ranges for all inducers, even though we observed significant decreases in albumin and urea secretion in the presence of 10 µM RIF, reflecting its known hepatotoxicity. Our results indicate that mRNA measurements and metabolic activity measurements in HepaRG cells generally give comparable results for fold-induction of CYPs.