The purpose of the present work was accessing the influence of benz[a]pyrene added into substrate on composition of polyarenes in plant and on growth and development of common barley (Hordeum vulgare) in in vitro conditions. We found that contamination of substrate with benz[a]pyrene (BP) decreased seed germination of Hordeum vulgare L. Larger BP doses produced the trend for a linear decrease in height and biomass of the aboveground part of Hordeum vulgare. The change of root biomass is not linear. Using the method of highly-efficient liquid chromatography, we identified 14 structures of polyaromatic hydrocarbons (PAHs). Significant amounts of light polyarenes were found in the control plants. Concentration of polyarenes in the plants grown on polluted substrate varied depending on benz[a]pyrene doses. The pollutant in doses of 10–20 ng/g increased content of polyarenes in the plants. The dose of 30 ng/g started the mechanisms of polyarene decomposition and so concentration decreased. Higher doses disturbed the protection mechanisms of plants and so increased content of polyarenes in plants and decreased biomass. Light polyarenes preferably accumulated in aboveground plant part whereby heavy polyarenes – in roots.