This paper describes three simple high performance liquid chromatography methods to separate mixtures of free fatty acids, mixtures of different triglycerides and mixtures of all fat classes (monoglycerides, diglycerides, triglycerides and free fatty acids). It is possible with our methods to identify and quantify each peak of the chromatogram. These methods have been designed to monitor lipase reactions. Using a first set of conditions, we have been able to separate five fatty acids: linolenic, linoleic, palmitic, oleic and stearic, without any specific preparation of the samples. With a second set of conditions, we showed that the same mobile phase and the same column could separate both triglyceride species and fat classes. However, in the latter case, a flow gradient was used.