Coix lacryma-jobi L. is an Asian plant widely used in traditional medicine. Drought conditions can lead to an impairment of photosynthesis which can be softened by the exogenous application of proline and γ-aminobutyric acid (GABA) to maintain the electron transport chain (ETC) and avoid reactive oxygen species-induced oxidative damage by inducing antioxidative enzymes. The aim of this study was to evaluate the effects of water deficit on C. lacryma-jobi leaves under the application of proline and GABA. Chlorophyll (Chl) a fluorescence emission, antioxidative enzyme activity (catalase, superoxide dismutase, ascorbate peroxidase and guaiacol peroxidase) and proline and GABA contents in leaf tissues were performed in plants subjected to 12 days of drought. Chl a fluorescence indicated impairments on photosynthetic apparatus due to water deficit that resulted into increased activity of antioxidant enzymes. The application of proline but not GABA was more efficient in alleviating the effects of water deficit, as evidenced by the maintenance of the dynamic dissipation of the photosynthetic energy, besides the greater accumulation of proline under these conditions. Water deficit induces alterations in the ETC, which is softened by the application of proline to cope with the deleterious effects of drought in C. lacryma-jobi plant leaves.