Abstract: This study investigated the improvement effects of beta-glucanase (BGS) on the specific volume, texture, and volatile flavor compounds of bread prepared with 30% and 50% whole highland barley flour. The changes in specific volume, texture, crumb structure and color, and sensory while the BGS levels ranging from 0.02% to 0.10% were analyzed. The volatile flavor compounds in highland barley bread were tested by gas chromatography-ion mobility spectrometry. The results showed that the specific volume, elasticity, and gas cell numbers of highland barley bread all improved significantly (P<0.05) with increasing BGS concentration. When BGS was added at 0.10%, the specific volume of 30% highland barley bread reached up to 3.46 mL/g, close to that of the white bread (3.83 mL/g). Meanwhile, the elasticity of 30% and 50% highland barley bread increased by 10.36% and 43.57%, respectively. The main volatile flavor compounds in the bread core were esters, aldehydes, alcohols, ketones, and acids. The relative content of esters and ketones in 50% barley bread was higher than that in 30% barley bread, and the characteristic aromas of highland barley with fruit flavor, grass flavor, and flower flavor were more prominent. The effects of BGS on the composition of volatile flavor compounds of bread were unobvious. However, it could increase the relative contents of volatile flavor compounds such as n-hexanol, benzaldehyde, and ethyl acetate, and enhance the fruit flavor of highland barley bread.