Glycyrrhetinic Acid (GA) is renowned for its anti-inflammatory and antioxidative properties, particularly in treating various liver diseases. However, its effects on normal hepatocytes, like human LO2 cells, are not well-documented. This study aimed to explore these effects and extrapolate potential systemic impacts on physical fitness and mental health. In this research, LO2 cells were exposed to varying concentrations of GA. We assessed cell viability, levels of reactive oxygen species (ROS), superoxide dismutase (SOD), glutathione (GSH), and malondialdehyde (MDA). Additionally, whole-transcriptome sequencing (RNA[1]Seq) was employed to identify changes in gene expression post-GA treatment. The results showed that GA significantly inhibited LO2 cell viability and led to increased intracellular ROS and MDA levels, alongside decreased SOD and GSH contents. Post-treatment, pro-inflammatory cytokines (TNF-α, IL-1, and IL-6) were notably increased. A total of 2856 differentially expressed genes were identified in the GA-treated group compared to controls, predominantly enriched in inflammation-related gene ontology (GO) terms and pathways, with the NF-κB pathway being notably activated at both mRNA and protein levels. These findings indicate that GA induces oxidative stress and inflammatory responses in LO2 cells through the activation of the NF-κB pathway. Given the critical role of oxidative stress and inflammation in systemic health, these cellular-level changes might have broader implications. Specifically, they could influence physical fitness and mental health, considering the interconnectedness of liver function with overall physiological and psychological wellbeing. This study lays the groundwork for further exploration into how GA’s effects on hepatocytes could translate into systemic health outcomes, emphasizing the need for a holistic understanding of its impacts beyond liver-specific applications.