A rapid and sensitive method was established for the accurate determination of patulin in fresh apples. Experimental parameters including extraction methods, mobile phase composition and flow rate for HPLC which always affect the sensitivity and analysis efficiency of the method were optimized systemically. The results indicated that solid phase extraction (SPE) techniques using functionalized polystyrene-divinylbenzene column (PEP) exhibited satisfactory recoveries comparing with other extraction methods. Under the optimized chromatographic conditions, patulin could be well separated from other impurities within 4 min, and detected with a wide linear range between 10-25 000 μg/L and a detection limit of 10 μg/L (S/N=3) using acetonitrile/water (90:10, v/v) as mobile phase, 0.8 mL/min as flow rate. The proposed PEP-SPE-HPLC method was further applied to the determination of patulin in fresh apples in the range of 82.42%-95.57%, indicating it might serve as an efficient tool for the accurate quantification of patulin in food samples.