High temperature and accompanying high vapor pressure deficit often stress plants without causing distinctive changes in plant canopy structure and consequential spectral signatures. Sun‐induced chlorophyll fluorescence (SIF), because of its mechanistic link with photosynthesis, may better detect such stress than remote sensing techniques relying on spectral reflectance signatures of canopy structural changes. However, our understanding about physiological mechanisms of SIF and its unique potential for physiological stress detection remains less clear. In this study, we measured SIF at a high‐temperature experiment, Temperature Free‐Air Controlled Enhancement, to explore the potential of SIF for physiological investigations. The experiment provided a gradient of soybean canopy temperature with 1.5, 3.0, 4.5, and 6.0°C above the ambient canopy temperature in the open field environments. SIF yield, which is normalized by incident radiation and the fraction of absorbed photosynthetically active radiation, showed a high correlation with photosynthetic light use efficiency (r = 0.89) and captured dynamic plant responses to high‐temperature conditions. SIF yield was affected by canopy structural and plant physiological changes associated with high‐temperature stress (partial correlation r = 0.60 and −0.23). Near‐infrared reflectance of vegetation, only affected by canopy structural changes, was used to minimize the canopy structural impact on SIF yield and to retrieve physiological SIF yield (ΦF) signals. ΦF further excludes the canopy structural impact than SIF yield and indicates plant physiological variability, and we found that ΦF outperformed SIF yield in responding to physiological stress (r = −0.37). Our findings highlight that ΦF sensitively responded to the physiological downregulation of soybean gross primary productivity under high temperature. ΦF, if reliably derived from satellite SIF, can support monitoring regional crop growth and different ecosystems' vegetation productivity under environmental stress and climate change. The study investigated soybean responses to high‐temperature stress, which is a major threat to crop yield in the U.S. Corn Belt. We utilized an unprecedented experiment, Temperature Free‐Air‐Controlled Enhancement experiment, which enabled detailed investigation by providing four levels of canopy temperature increments. Using advanced hyperspectral remote sensing technique, we collected sun‐induced chlorophyll fluorescence (SIF) data and demonstrated that SIF has the unique capability of quantifying crop physiological variability under high‐temperature stress (i.e., depressions in instantaneous photosynthetic rates per unit leaf area). The unique strength of SIF for quantifying physiological status is expected to contribute to improving vegetation productivity quantification.