Dietary plant sterols have received increasing attention in recent years due to their favorable health benefits. The present research focused on quantification of phytosterols as free, esterified and total forms in different tetraploid (5 cultivars of Triticum durum Desf., 9 cultivars of Triticum dicoccon Schrank) and hexaploid (5 cultivars of T. aestivum L., 12 cultivars of Triticum spelta L.) wheats. Tetraploid wheats showed the highest content of total sterol (79.4 and 79.5 mg of sterols /100 g dry weight for T. durum and T. dicoccon, respectively). Hexaploid cultivars were the best source of esterified sterols (40.7% and 37.3% of total sterols for Triticum aestivum and T. spelta, respectively). Significant amounts of free sterols (65.5% and 60.7% of total sterols for T. durum and T. dicoccon, respectively) were found in the tetraploid cultivars. The most abundant phytosterol in all wheat samples was sitosterol accounting for 45.1-59.1, 46.6-57.4 and 38.6-59.5% of total, free and esterified sterol fraction, respectively. These results demonstrate that although the sterol profile present in tetraploid and hexaploid wheat species is the same, differences in their relative amounts and distribution allow statistical differentiation between hexaploids and tetraploids, and between soft and durum wheats.