Abstract: Royal jelly is a yellowish, creamy and acidic substance secreted by mandibular and hypopharyngeal glands of young worker bees. Fatty acids including (E)-10-hydroxy-2-decenoic acid, 10-hydroxydecanoic acid, 3,10-dihydroxydecanoic acid, 8-hydroxyoctanoic acid, sebacic acid and so on are one of the major components of royal jelly with a variety of biological activities, such as anti-inflammation, anti-bacteria and anti-tumor. Fatty acids are one of the main active components of royal jelly which are considered an important indicator in quality control. This article summarizes the identification methods of the royal jelly fatty acid including the principle of the method, the comparisons of technology and application features of methods, as well as the future research prospects. This review will provide a reference for the development of determining methods for fatty acids and pave the way for quality control of royal jelly.