Background: A study of the correlation between the particle size of lignocellulosic substrates and ultrasound pretreatment on the efficiency of further enzymatic hydrolysis and fermentation to ethanol. Results: The maximum concentrations of glucose and, to a lesser extent, di- and trisaccharides were obtained in a series of experiments with 48-h enzymatic hydrolysis of pine raw materials ground at 380-100 rpm for 30 min. The highest glucose yield was observed at the end of the hydrolysis with a cellulase dosage of 10 mg of protein (204 ±21 units CMCase per g of sawdust). The greatest enzymatic hydrolysis efficiency was observed in a sample that combined two-stage grinding at 400 rpm with ultrasonic treatment for 5-10 min at a power of 10 W per kg of sawdust. The glucose yield in this case (35.5 g glucose l-1) increased twofold compared to ground substrate without further preparation. Conclusions: Using a mechanical two-stage grinding of lignocellulosic raw materials with ultrasonication increases the efficiency of subsequent enzymatic hydrolysis and fermentation.