Microalgae have fascinating properties that qualify them as a potential feedstock for biorefinery applications by producing various biomolecules such as lipids, PUFAs, carbohydrates, pigments, biotoxins, sterols, enzymes, and edible proteins. Most of these compounds are produced intracellularly necessitating effective extraction, recovery, and characterization processes. The development of viable and scalable technologies to produce desired microalgal by-products with greater production and purity has attracted a lot of interest recently. This chapter provide an overview of the existing methods that are used for the extraction, separation, and characterization of biomolecules from microalgal biomass to generate value added products. These valuable compounds have diverse applications in the field of pharmaceuticals, nutraceuticals, cosmetic, textile and food industry also; which is reviewed in detail in this book chapter.