In order to investigate the mitigating effect of exogenous salicylic acid (SA) on flue-cured tobacco under low temperature stress, our experiment used an artificial climate incubator to simulate a low temperature of 4 ℃, and set five mass concentrations of exogenous salicylic acid (SA) (10, 25, 50, 100, 150 mg/L) to pretreat potted K326 tobacco seedlings through roots for 3 consecutive days, followed by low temperature stress treatment for 4 consecutive days. Normal temperature (25 ℃) growth was used as a positive control (CK), and a low temperature of 4 ℃without the application of exogenous SA was performed as a negative control (CK0). The relative electrical conductivity, malondialdehyde (MDA) content, osmotic adjustment substance content, antioxidant enzyme activity and non-enzyme antioxidant content of tobacco leaves were measured. The results showed that the application of exogenous SA could significantly increase the activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD). The enzymes were the most active under 50, 10, 25 mg/L exogenous SA treatment, respectively; the glutathione reductase (GR) and ascorbate peroxidase (APX) activities were increased, and GR and APX activities reached the maximum under the treatment of 100 mg/L and 50 mg/L exogenous SA respectively. Under low temperature stress, the soluble sugar (SS) content of tobacco seedlings was increased by 17.46% compared to CK0 under the treatment of 10 mg/L exogenous SA. The contents of soluble protein (SP), proline (Pro), and ascorbic acid (ASA) were the highest at 50 mg/L exogenous, which increased by 30.35%, 316.44% and 31.21% respectively compared with CK0. The content of reduced glutathione (GSH) was significantly increased by 51.38% compared with CK0 under 25 mg/L exogenous SA treatment. The improvement of various indicators has alleviated the rise of REC and MDA content under low temperature stress, and cold resistance membership function analysis shows 50mg /L exogenous SA treatment had the greatest alleviation effect on K326 under 4 ℃ low temperature stress.