Microbial biosurfactants are surface-active compounds produced by microorganisms, classified based on molecular weight, physicochemical properties, and mode of action. Low-molecular-weight biosurfactants reduce surface and interfacial tension, while high-molecular-weight bioemulsifiers stabilize oil-in-water emulsions. These compounds are gaining attention due to their advantages over synthetic surfactants in environmental, food, biomedical, and industrial applications. However, their large-scale production is limited by high costs and limited understanding of their interactions with cells and the environment. This review summarizes current knowledge and recent advances in biosurfactant applications and biotechnological strategies for improving production. Biosurfactants are structurally diverse, consisting of hydrophilic and hydrophobic moieties. They are divided into two main classes: biosurfactants (e.g., lipopeptides, glycolipids, proteins) and bioemulsifiers (e.g., polysaccharides, lipopolysaccharides, lipoproteins). Glycolipids, such as rhamnolipids, trehalolipids, sophorolipids, and mannosylerythritol lipids, are the best-studied microbial surfactants. These compounds have various applications, including environmental remediation, food processing, and pharmaceuticals. Future research should focus on improving production efficiency and understanding their interactions with the environment and cells to enhance their potential in industrial applications.Microbial biosurfactants are surface-active compounds produced by microorganisms, classified based on molecular weight, physicochemical properties, and mode of action. Low-molecular-weight biosurfactants reduce surface and interfacial tension, while high-molecular-weight bioemulsifiers stabilize oil-in-water emulsions. These compounds are gaining attention due to their advantages over synthetic surfactants in environmental, food, biomedical, and industrial applications. However, their large-scale production is limited by high costs and limited understanding of their interactions with cells and the environment. This review summarizes current knowledge and recent advances in biosurfactant applications and biotechnological strategies for improving production. Biosurfactants are structurally diverse, consisting of hydrophilic and hydrophobic moieties. They are divided into two main classes: biosurfactants (e.g., lipopeptides, glycolipids, proteins) and bioemulsifiers (e.g., polysaccharides, lipopolysaccharides, lipoproteins). Glycolipids, such as rhamnolipids, trehalolipids, sophorolipids, and mannosylerythritol lipids, are the best-studied microbial surfactants. These compounds have various applications, including environmental remediation, food processing, and pharmaceuticals. Future research should focus on improving production efficiency and understanding their interactions with the environment and cells to enhance their potential in industrial applications.