Abstract: Selected the pepper seedlings as the experimental material, different concentration of spermidine, chitosan, salicylic acid, CaCl2, betaine solution were sprayed respectively, and then relative conductivity, chilling injury index, soluble sugar content, proline content, malondialdehyde (MDA) content were measured and analyzed after 5 days under the day temperature/night temperature 10℃/5℃ low temperature stress, this paper discussed the effect of exogenous substances on pepper seedlings in low temperature stress. The results indicated that appropriate concentration of exogenous substances produce positive influence on improving cold resistance of pepper seedlings by reducing relative conductivity, chilling injury index, accumulation of MDA and increasing the osmotic regulation substances content such as soluble sugar and proline. The optimal concentrations of Spermidine, chitosan, salicylic acid, CaCl2, betaine to improve chilling resistance were 0.5 mmol/L, 50 mg/L, 2 mmol/L, 10 mmol/L, 10 mmol/L.