The pretreatment process has gained renewed interest in recent years and is considered the most essential stage in efficiently refining lignocellulosic biomass (LCB). LCB is a valuable alternative source for biochemicals and biofuels. Comprising cellulose, lignin, and hemicellulose, LCB has a recalcitrant and complex structure that remains challenging to process. However, other chemical and physical pretreatment methods often face significant issues such as solvent toxicity and high energy consumption. Deep eutectic solvents (DESs) have emerged as environmentally friendly and green solvents for pretreating LCB. They are characterized by low toxicity, ease of recycling, affordability, biodegradability, and reusability; as a result, DESs are recognized as sustainable and eco-friendly solvents for LCB pretreatment. This chapter discusses DESs, biofuels, and LCBs, covering their components, detailed LCB composition, and pretreatment methods for biofuel production. It highlights DESs as effective solvent alternatives, describing their types, properties, and recent advancements in biomass pretreatment with DESs. The chapter details the removal of hemicellulose and lignin using DESs and explores how parameters such as temperature, duration, and solid-to-liquid ratios are optimized during pretreatment. It also addresses existing challenges and future perspectives, concluding with final remarks.