The general global trend of transition to a carbon-neutral economy is updating research on deep processing of renewable organic raw materials. The most popular direction in this area is the processing of lignocellulosic biomass (LCB) for the production of valuable chemical products. Hemicelluloses are an important class of plant biopolymers consisting of different monosaccharide units depending on the type of LCB and the method of their extraction. These biopolymers are receiving more attention because they exhibit a wide range of biological and pharmacological activities, such as antitumor, immunomodulatory, antimicrobial, antioxidant, anticoagulant, which makes them one of the most promising targets in the biomedical and pharmaceutical fields. However, hemicelluloses are widely distributed in nature and can be found in various sources, such as plants, microorganisms, algae and animals. This scientific article provides an overview of the structural diversity and isolation methods of hemicelluloses, the understanding of which is critical for their full potential use in various fields, including biomedical fields. A description is also provided of the dependence of the structural differences of polysaccharides on the sources of their content, and the advantages and disadvantages of various isolation procedures are described.