Abstract: Ghee is a widely consumed dairy product recognized for its nutritional, therapeutic, and culinary significance. The fatty acid composition of ghee plays a crucial role in determining its quality, oxidative stability, and potential health benefits. In recent years, Gas Chromatography–Mass Spectrometry (GC–MS) has emerged as a powerful and reliable analytical technique for the characterization of fatty acid profiles in dairy lipids. Typically, lipid samples are subjected to transesterification to convert triglycerides into fatty acid methyl esters (FAMEs), which enhances their volatility and facilitates accurate separation and identification. Numerous studies have reported that ghee contains a complex mixture of saturated, monounsaturated, and polyunsaturated fatty acids. Among these, palmitic acid, stearic acid, oleic acid, and linoleic acid are consistently identified as the major constituents. The relative abundance of these fatty acids can vary significantly depending on factors such as the source of milk, animal diet, geographical origin, and processing methods. Such variations directly influence the nutritional profile and functional properties of ghee. Furthermore, the ratio of saturated to unsaturated fatty acids is widely used as an important indicator for evaluating the nutritional quality of ghee. GC–MS-based fatty acid profiling has also been extensively applied in assessing product authenticity, detecting adulteration, and ensuring quality control in commercial samples. Overall, existing literature highlights that GC–MS provides a sensitive, accurate, and comprehensive approach for fatty acid characterization, making it an indispensable tool in dairy chemistry research.