Abstract: To study the metabolic response mechanism of Vetiveria zizanioides under prometryne stress, extensive targeting metabolism was adopted to study the changes of metabolites in the leaves of V. zizanioides under 5 mg/L prometryne stress. The results showed that when prometryne was stressed for 5 days, the total amount of differential metabolites was 121 compared with the control; compared with the control and stress for 5 days, the total amount of differential metabolites of 10 days stress was 151 and 48 respectively. In the process of stress, flavonoids, alkaloids and lipids are the most down-regulated metabolites, and nucleotides and their derivatives, alkaloids and others(such as microorganisms, sugars and alcohols) are the most up-regulated metabolites; the estimation of pyruvate,jasmonic acid, serotonin and other substances is critical in the stress resistance process, in which jasmonic acid may play a role in the termination of defense signal in the later stage of stress; the amount of down-regulated free amino acids was higher than that of up-regulated free amino acids. Analysis on metabolic pathway revealed that arginine and proline metabolism and plant hormone signal transduction play a major role in leaf resistance to prometryne stress.