A method and a kit for detecting folate are disclosed. The method includes the following steps: (a) mixing a sample and an extraction buffer to form a mixture, heating and then cooling the mixture, and separating a supernatant from the mixture by centrifugation; (b) adding a recombinant γ-glutamyl hydrolase (GGH) and a folate conversion enzyme to the supernatant to drive catalysis; (c) stopping the catalysis; and (d) analyzing the supernatant by high performance liquid chromatography.