Abstract: Background and Objective: Lettuce seed germination at high temperatures is a problem faced by farmers and seed producers in Brazil. At high temperatures, germination is reduced or null and changes in physiology and the patterns of germination-related enzymes, such as endo-β-mannanase, can occur. The objective of this study was to determine the relationship between endo-β-mannanase activity and tolerance to thermoinhibition in lettuce seeds, as well as to determine the relationship between seeds and antioxidant enzymes. Materials and Methods: Seeds from two lettuce cultivars, Everglades (thermotolerant) and Veronica (thermosensitive), were used. The seeds were assessed for first count, germination and germination speed index at seven temperatures (20, 23, 25, 27, 30, 33 and 35°C). Additionally, the activity of the enzymes endo-β-mannanase, catalase, ascorbate peroxidase and superoxide dismutase and the levels of hydrogen peroxide were quantified in each cultivar at the different temperatures. The experimental design was completely randomized in a 2×7 factorial arrangement, with two cultivars and seven temperatures. Results: At temperatures above 30°C, the Everglades cultivar showed higher values than the Veronica cultivar, including for endo-β-mannanase activity, in all treatments. Additionally, the oxidation levels, as confirmed by the analysis of oxidative pathway enzymes, were higher in the Veronica cultivar, thus confirming that the cultivar is thermosensitive to temperatures above 25°C. Conclusion: It is concluded that high temperature directly affects the germination parameters and consequently the activity of endo-β-mannanase in lettuce seeds, especially in the thermosensitive cultivar Veronica.