Mānuka honey, derived from the nectar of Leptospermum scoparium, is renowned for its unique phytochemical composition and antibacterial properties. The key component responsible for its bactericidal activity is methylglyoxal, which is formed from dihydroxyacetone present in the nectar during storage. Leptosperin, a di-glucoside specific to mānuka, including Leptospermum plants, serves as a certification marker for mānuka honey by the Unique Mānuka Factor Honey Association (UMFHA). In addition, the New Zealand government has established authentication criteria based on the quantification of four specific chemical markers and pollen DNA (Leptospermum scoparium). The bioavailability of leptosperin in both humans and mice has been studied, with methyl syringate (MSYR) and its metabolites identified as the predominant compounds associated with its intake. Recent studies have shed light on the biosynthetic pathway of leptosperin, which involves MSYR as a template for MSYR-glucose biosynthesis, subsequently leading to leptosperin production. The compositional differences between nectar and honey during maturation can be attributed to plant- or honeybee-derived enzymatic activities or microbial hydrolases. Understanding the phytochemical composition and authentication methods of mānuka honey is crucial to ensure its quality, prevent adulteration, and explore its potential health benefits.