Arsenic trioxide (As(2)O(3)) has been shown to inhibit invasion/migration in cancer cells. However, the underlying mechanism is poorly understood. To identify the role of As(2)O(3) in regulating invasion/migration activity in human gastric cancer SGC-7901 cells, the effects of As(2)O(3) on cell invasion/migration activity, the expression of cyclooxygenase-2 (Cox-2), prostaglandin E2 (PGE2), thromboxane B2 (TXB2), leukotriene B4 (LTB4), and matrix metalloproteinase-2 (MMP-2) and intracellular reactive oxygen species (ROS) were examined. Furthermore, N-acetyl-l-cysteine (NAC, a radical scavenger) and celecoxib (a Cox-2 inhibitor) were used to explore the molecular mechanism. The results demonstrated that As(2)O(3) (1 and 2 μmol/L) inhibited invasion/migration activity in SGC-7901 cells at 24 h and suppressed the expression of Cox-2, PGE2 and MMP-2 (P < 0.05), whereas the same treatment had no significant effect on TXB2 and LTB4 expression. In contrast, intracellular ROS were increased (P < 0.05). Moreover, NAC eliminated the excessive ROS and restored the expression of Cox-2 and MMP-2 and invasion/migration activity in As(2)O(3)-treated cells (P < 0.05). These results suggest that ROS may be a critical factor in regulating the invasion/migration process. Moreover, celecoxib significantly decreased Cox-2, MMP-2 and PGE2 expression and inhibited invasion/migration activity in As(2)O(3)-treated cells (P < 0.05), indicating that As(2)O(3) inhibits invasion/migration by regulating the expression of Cox-2/PGE2/MMP-2. In conclusion, these results suggest that increased ROS play a critical role in inhibiting invasion/migration by suppressing the Cox-2/MMP-2 pathway in As(2)O(3)-treated SGC-7901 cells and regulating intracellular ROS levels may be a promising strategy in gastric cancer therapy.