In this experiment, glutathione (GSH), oligopeptide (GCG), and three substances including glutamate (Glu), cysteine (Cys) and gamma aminobutyric acid (GABA) were used as treatments, water was used as control (CK), two wheat (Triticum aestivum L.) seedlings Jingdu 40 and Liaochun 17 were taken to study the effects of different treatments on physiological characteristics under high temperature resistance, and furthermore to determine the physiological activity of the oligopeptide GCG. The results show that under the condition of high temperature stress, the treatments with 200μmol/L GSH and GCG are able to maintain higher leaf relative water content of wheat seedlings, reduce the relative electrolyte leakage and malondialdehyde (MDA) content, and conduce to the integrity of cell membrane. In the meantime, GCG treatment could significantly enhance the 1, 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity in wheat seedlings under high temperature stress, and the activities of antioxidative enzymes are also improved to reduce the damage caused by the high temperature. Comparing with the other two kinds of treatments, the effect of GCG treatment is most significant. On the whole, GCG could enhance the high temperature tolerance of wheat through modifying the activities of antioxidative enzymes similar to GSH. Therefore as a kind of synthetic oligopeptide, GCG is expected to develop the applications as an agricultural production regulator.