A method for determining the content of polyglutamic acid in water-soluble fertilizer using an ultra-high speed fully automatic amino acid analyzer is proposed. Water-soluble fertilizer containing polyglutamic acid is respectively subjected to hydrolysis treatment and extraction with ultrapure water. Under the selected instrument operating conditions, a single point external standard method is used to measure the content of hydrolyzed glutamic acid and free glutamic acid using a ultrahigh speed fully automatic amino acid analyzer. The difference between the two is the content of polyglutamic acid. The detection limit of the method is 0.004 8 nmol/mL, the relative standard deviation (n=10) of the measurement results is 0.68% to 3.73%, and the ratio of the measured value to the actual value is 96.2% to 106.0%.