Of polycyclic aromatic hydrocarbons (PAHs) present in food, benzo(a)pyrene (B(a)P) is the most often determined compound as a surrogate for all PAHs. Various chromatographic procedures, including gas, liquid and thin layer chromatography have been applied to isolate these substances from contaminated samples. Because of the complexity of food matrices additional clean up of extract prior chromatographic analysis is necessary. This paper describes extraction of B(a)P from special kind of Slovakian smoked cheese by accelerated solvent extraction (ASE), clean up by gel permeation chromatography (GPC) and finally determination of B(a)P by high performance liquid chromatography (HPLC) with fluorescence detector in 430 nm. The influence of parameters such as: sample preparation as well as smoking time and temperature on concentration of B(a)P in smoked cheese was investigated. The method allows determination of B(a)P in smoked cheese on μg/kg level. The content of B(a)P in the analysed smoked cheese samples ranged from <0.1 µg/kg to 3.8 µg/kg and strongly depended on time and temperature of processing. In most cases, the concentration of B(a)P in samples of cheese from home-made ovens was over acceptable limit.