Glyphosate and 2,4-D have been commonly used for control of common and giant ragweed before planting of corn and soybean in the midwestern United States. Because these herbicides are primarily applied in early spring, environmental factors such as temperature may influence their efficacy. The objectives of this study were to (1) evaluate the influence of temperature on the efficacy of 2,4-D or glyphosate for common and giant ragweed control and the level of glyphosate resistance and (2) determine the underlying physiological mechanisms (absorption and translocation). Glyphosate-susceptible (GS) and glyphosate-resistant (GR) common and giant ragweed biotypes from Nebraska were used for glyphosate dose–response studies, and GR biotypes were used for 2,4-D dose–response studies conducted at two temperatures (day/night [d/n]; low temperature [LT]: 20/11 C d/n; high temperature [HT]: 29/17 C d/n). Results indicate improved efficacy of 2,4-D or glyphosate at HT compared with LT for common and giant ragweed control regardless of susceptibility or resistance to glyphosate. The level of glyphosate resistance decreased in both the species at HT compared with LT, primarily due to more translocation at HT. More translocation of 2,4-D in GR common and giant ragweed at HT compared with LT at 96 h after treatment could be the reason for improved efficacy. Similarly, higher translocation in common ragweed and increased absorption and translocation in giant ragweed resulted in greater efficacy of glyphosate at HT compared with LT. It is concluded that the efficacy of 2,4-D or glyphosate for common and giant ragweed control can be improved if applied at warm temperatures (29/17 C d/n) due to increased absorption and/or translocation compared with applications during cooler temperatures (20/11 C d/n).