Economic biofuel production requires high sugar yields during biomass pretreatment, however, the chemical and structural features of biomass can be obstructive toward efficient xylose hydrolysis. Here, we tested the hindrance imposed by the multiscale structure of biomass on the hydrolysis of xylan during dilute acid pretreatment by studying the effects of both the chemical nature of xylan and physical structure of biomass. Dilute acid pretreatment of poplar wood at particle sizes ranging from 10 um to 10 mm was conducted, however, no significant differences in the rates of xylan hydrolysis were observed over more than 2 orders of magnitude in particle size. A significant reduction in the rate of xylan hydrolysis was observed when compared to the intrinsic rate of hydrolysis for isolated xylan. Thus, it appears likely that the chemical structure of xylan and/or the interaction of xylan with other polymers in the cell wall matrix have greater effects on xylan hydrolysis rates than mass transfer limitations.