The fatty acid composition of milk is of considerable interest due to their nutritional and functional properties. Although rapid milk fat separation and transesterification procedures have been developed, the overall procedure remains time consuming, specially, for the analysis of a large number of samples. In this work, a fast and simple method for direct profiling of fatty acids from milk using thermochemolysis has been developed. This method has the capability of directly analyse fatty acids from one drop of milk without fat extraction or cleanup. Our approach for thermochemolysis is based on thermal desorption integrated with a cold trap inlet. The optimized method does not present isomerisation/degradation of polyunsaturated fatty acid and shows milk fatty acid profiles comparable to the conventional method based on fat extraction and alkaline transesterification. Overall, this method has demonstrated significant potential for high throughput analysis of fatty acids in milk.