The total polyphenol content in barley spent gains were analyzed by Folin-Ciocalteu method. The optimum conditions were investigated using gallic acid as standard. and were as followed: Folin-Ciocalteu reagent 1.0 mL, 8% Na2CO3 3.0 mL, developing temperature 25 ℃, reaction time 60 min, in darkness. Then the solution was detected at the wavelength of 760 nm. When the gallic acid was in range of 0～8 μg/mL, the concentration and the absorbance value had a good linear relationship. The regression equation was found as followed: y=0.121x+0.017 1, R2=0.999 2. This method had advantages of conciseness, good stability and high accuracy, which is suitable for analysis of polyphenol in barley spent gains.