Chenopodium ambrosioides is an invasive species, which has strong allelopathic effect on surrounding plants. In this study, the methods of soil culture and filter paper culture were adopted to simulate the eluviation and volatilization of the volatile oil from C. ambrosioides, respectively, and to investigate the allelopathy of the volatile oil on the lipid peroxidation and antioxidant enzyme activities of Vicia faba root tip cells, with the mechanisms of the induced tip cell apoptosis analyzed. At the early stage (24 h) of soil culture and filter paper culture, the superoxide dismutase, peroxidase and catalase activities of the tip cells decreased after an initial increase with the increasing dose of the volatile oil, and the malondialdehyde content of the tip cells increased with the increasing volatile oil dose and treated time. At the midterm (48 h) and later (72 h) stages of soil culture and filter paper culture, a typical DNA ladder strip appeared, suggesting that the volatile oil from C. ambrosioides could induce the apoptosis of the tip cells, and the apoptosis was dose- and time dependent. This study showed that the volatile oil from C. ambrosioides could act on its surrounding plants via eluviation and volatilization, making the lipid peroxidation of acceptor plants aggravated and the antioxidant enzyme activities of the plants inhibited, resulting in the oxidative damage and apoptosis of the plant root tip cells, and accordingly, the inhibition of the plant growth. Under soil culture, the root tip cells of V. faba had higher antioxidant enzyme activities and lesser DNA damage, suggesting that the volatile oil from C. ambrosioides via volatilization had stronger allelopathy on the growth of surrounding plants than via eluviation.