The invention discloses application of phthalic acid in amino acid analysis. The application comprises adding the phthalic acid in a mobile phase for performing amino acid liquid chromatography-mass spectrum combined analysis, wherein concentration of the added phthalic acid is 0.005-0.1mmol/L. According to the application of phthalic acid in amino acid analysis, detection sensitivity of the amino acid can be significantly improved, and meanwhile chromatography separating effect can be improved; the application is convenient that only adding proper amount of the phthalic acid into the mobile phase is needed, and thereby analysis result of the amino acid can be significantly improved without changing other analysis conditions; in addition, experiments show that a chromatographic column and a mass spectrum ion source are prevented from being seriously damaged by using the mobile phase added with the phthalic acid for a long time, reproducibility is good, and quantitative analysis is easy to implement, and good application and promotion values are provided.