Abstract: The addition of different kinds of acidifiers into animal feed would be effective in decreasing pH value and acid-binding capacity, and thus it brings the advantages such as facilitating the digestion and absorption, and enhancing the immune function. In connection with the demands for food safety and environment-friendly production, acidifiers have received considerable attention as feed additives. Regulating the usage and management of feed acidifiers requires the establishment of a synchronous detection method for different kinds of organic acids with high speed and high accuracy. The research object of this study was to establish the synchronous detection method, and to optimize the detecting condition of 9 kinds of organic acids (citric acid, tartaric acid, malic acid, lactic acid, formic acid, acetic acid, fumaric acid, propionic acid and butyric acid) based on the ion exclusion chromatography coupled with diode array detector (DAD). All the organic acids in this study were extracted by deionized water for further detection. A Bio-Rad Aminex HPX-87H (300 mm × 7.8 mm, 9 μm) ion exclusion chromatography column with DAD was applied and sulfuric acid solution was taken as the flowing phase, and the different effects of chromatographic conditions such as mobile phase, flow rate, column temperature and detecting wave length on the 9 organic acids in feed acidifiers during synchronous detection were compared and analyzed. Then, the analytical methods established for all the above mentioned parameters were investigated for their linearity range, the limit of detection, the recovery rate and precision, followed by the verification using actual samples. The results showed that the best optimized parameter conditions for synchronously detecting the 9 organic acids were 0.025 mol/L sulfuric acid solution as flowing phase, flowing speed of 0.70 mL/min, column temperature of 30 ℃, and 210 nm detecting wave length for DAD. The qualitative analysis of organic acids was executed based on their retention time and ultraviolet absorption spectra, while the quantitative analysis was executed based on the external standard method. According to the results of this study, it could be found that the mass concentration and the peak area had behaved nice linear correlations in specific range for the 9 organic acids, respectively, with the determination coefficients higher than 0.999. All the 9 organic acids could be successfully separated within 20 min. The limit of detection of the synchronous detection method was from 0.01 to 2.4 mg/L and the recovery rate was from 90.7% to 101.1%, while the relative standard deviation was from 0.91% to 3.33%. Bases on these results, the synchronous detection method focused in this study shows to be one of the successful ways for organic acid detecting in different animal feed acidifiers. This study can provide the technical support in further investigation of the acidifiers’ application in the animal feed manufacturing factory as well as in the real market for quality assurance.