Recent research on non-thermal plasma (NTP, an ionized gas) has identified it as a novel cancer therapeutic tool. However, the molecular mechanism remains unclear. In this study, we demonstrated NTP induced cell death of head and neck cancer (HNC) through the AKT ubiquitin–proteasome system. NTP increased the gene expression of mitochondrial E3 ubiquitin protein ligase 1 (MUL1), an E3 ligase for AKT, and NTP-induced HNC cell death was prevented by MUL1 siRNA. We also showed that MUL1 inhibited the level of AKT and p-AKT and MUL1 expression was increased by NTP-induced ROS. Furthermore, we optimized and manufactured a new type of NTP, a liquid type of NTP (LTP). In syngeneic and xenograft in vivo tumor models, LTP inhibited tumor progression by increasing the MUL1 level and reducing p-AKT levels, indicating that LTP also has an anti-cancer effect through the same mechanism as that of NTP. Taken together, our results suggest that NTP and LTP have great potential for HNC therapy.