The invention provides a method for detecting retinol and a precursor thereof. The method comprises the following steps: 1, extracting retinol and/or the precursor thereof from a sample; 2, separating retinol and/or the precursor thereof through HPLC; and 3, carrying out MS/MS quantitative detection, wherein the sample is a dry blood spot card sample, and an extractant adopted in the invention is trichloromethane. A filter paper card is adopted as a carrier to adsorb a tiny amount of blood in order to make the dry blood stop card, so the difficulties of much blood induced by venous blood collection, bad population compliance, death induced by animal model blood drawing, and inconvenient long-distance conveying of samples in previous serum collection and detection process are overcome. The method has the advantages of small matrix effect, good linear relation of analysis in the concentration range of a standard curve, low detection limit, high sensitivity, good accuracy and high precision, is related with the result of detection of retinol in serum through HPLC, allows the contents of the retinol and the precursor thereof in the sample to be simultaneously and synchronously analyzed, and provides a method for comprehensive assessment of the in vivo retinol level.