The γ-linolenic acid (GLA), a polyunsaturated fatty acid with three double bonds, has attracted much attention due to its numerous biological properties. Borage, evening primrose, and blackcurrant are the primary sources of GLA, while algae and fungi constitute the majority of microbial sources. Existing GLA separation and purification technologies have their own advantages and disadvantages. A single enrichment method can not meet the requirements for high-purity GLA. The comprehensive use of different methods and the combination of modern separation and extraction technology could be the methods for preparing GLA in the future. Simultaneously, numerous studies have demonstrated the crucial involvement of GLA in the body’s metabolism and physiological control, as well as its efficacy in blood lipids, antithrombotic cardiovascular diseases, cancer and diabetes. Further research is necessary to gain a better understanding of the mechanism of action. This review of GLA sources, enrichment methods,metabolic pathways and physiological efficacy in humans could provide theoretical support for GLA as a functional health food for the prevention of multiple chronic diseases.