The aim of the present work was to develop and validate a reversed-phase highperformance liquid chromatography with diode array detection (RP-HPLC-DAD) method for determination of cholesterol and cholesterol oxides extracted from milk samples. The method consisted of direct samples saponification and analytes extraction, followed by their determination using RP-HPLC-DAD. Suitable analytical selectivity, linearity and range of application, intra-assay and intermediary precision, recovery, detection and quantification limits (LOD and LOQ) were achieved and, the results showed that the developed RP-HPLC-DAD method can be applied to routine determination of cholesterol and cholesterol oxides in milk samples.
direct saponification; 7-ketocholesterol; 25-hydroxycholesterol; liquid chromatography; dairy fat
