Objective The purpose of this paper was to establish a detection method of polyethylene microplastics during the use of disposable paper cup with polyethylene inner coating. Methods The sample pretreatment processes such as digestion, filtration and dispersion were fully studied and well optimized. It was found that 23% (v/v) of hydrogen peroxide digestion can eliminate the interference in tap water. 5.0 g/L of sodium bicarbonate coupled with ultrasound could effectively decompose the residual hydrogen peroxide in the digestion solution, so as to protect the silver filter membrane from corruption during filtrating. Before filtrating, adding 25% (v/v) ethanol to the digestion solution could effectively disperse the microplastics in the system, so that the microplastics would not stick to the wall and agglomerate during filtration. Results Based on the above pretreatment methods, we could quantitatively analyze the polyethylene microplastics in the disposable paper cups. During use, the cups could release 5000-200000 p/L with the size greater than 2 μ M microplastics, posing potential hazards to human health. Conclusion The results showed that the inner coating of disposable paper cup falls off a large number of polyethylene microplastics, which mainly occurs at the early stage of pouring hot water. Furthermore, this study revealed that the inner coating’s stability decreases during long-term storage. The use of disposable paper cup with long time storage would generate more microplastics, therefore brought more health risk.