This study evaluated the effects of different delignification pretreatments (enzymatic and organosolv) on the crystallinity and enzymatic hydrolysis of sugar cane residues. Six substrates were analyzed using Fourier Transform infrared spectroscopy (FTIR), X-ray diffraction, and scanning electron microscopy (SEM). Substrates treated only with organosolv showed good enzymatic hydrolysis performance and high crystallinity, likely due to the removal of amorphous material. The Crystallinity Index (CrI), Relative Number of Intensity (Ir), degree of cellulose mercerization (IIC-%), and Global Index of Saccharification (GIS) were used as measurement parameters. The results showed that substrates with higher CrI and lower IIC-% had better enzymatic hydrolysis. The organosolv pretreatment was effective in removing lignin and facilitating the recovery of glucose. The study concluded that pretreatment significantly affects the crystalline structure and enzymatic hydrolysis of lignocellulosic materials, with organosolv pretreatment being particularly effective in improving hydrolysis efficiency. The findings suggest that the crystallinity of cellulose is a key factor in enzymatic hydrolysis, and that pretreatments can enhance hydrolysis by reducing crystallinity and removing lignin. The study also highlights the importance of considering factors such as lignin content, crystallinity, and porosity in the hydrolysis process.This study evaluated the effects of different delignification pretreatments (enzymatic and organosolv) on the crystallinity and enzymatic hydrolysis of sugar cane residues. Six substrates were analyzed using Fourier Transform infrared spectroscopy (FTIR), X-ray diffraction, and scanning electron microscopy (SEM). Substrates treated only with organosolv showed good enzymatic hydrolysis performance and high crystallinity, likely due to the removal of amorphous material. The Crystallinity Index (CrI), Relative Number of Intensity (Ir), degree of cellulose mercerization (IIC-%), and Global Index of Saccharification (GIS) were used as measurement parameters. The results showed that substrates with higher CrI and lower IIC-% had better enzymatic hydrolysis. The organosolv pretreatment was effective in removing lignin and facilitating the recovery of glucose. The study concluded that pretreatment significantly affects the crystalline structure and enzymatic hydrolysis of lignocellulosic materials, with organosolv pretreatment being particularly effective in improving hydrolysis efficiency. The findings suggest that the crystallinity of cellulose is a key factor in enzymatic hydrolysis, and that pretreatments can enhance hydrolysis by reducing crystallinity and removing lignin. The study also highlights the importance of considering factors such as lignin content, crystallinity, and porosity in the hydrolysis process.