The global health care company Fresenius Kabi uses high performance liquid chromatography (HPLC) and ultra-high-performance liquid chromatography (UHPLC) with an evaporative light scattering detector (ELSD) for the identification, quantification and determination of triglycerides (TG) in structured triglycerides (STG) oil. The existing detector is no longer available in the market, thus a new ELSD detector was chosen to be used instead. The purpose of this project was to optimize and validate the existing analytical method with the new detector, by optimizing the parameters that affect the separation of triglycerides in the analyzed oil on the UHPLC- and HPLC-system. In addition, investigation of the impact of diglycerides (DG) during the determination of STG was one of the main goals of this project. This was done by observing if the DG were detected in the same assay chromatogram and if their peaks were separated from the peaks derived from TG. The parameters that were optimized were; the column type, flow rate, gradient program of the elution of the sample, different suppliers of the acetonitrile used as a solvent and the “gain” of the signal for the detector.

A C30 column “Develosil” and a flow rate 0.8 ml/min with a specific gradient program was selected to be used in the validation stage. DGs were separated from TGs and then analyzed. Peaks of the DGs were detected at the beginning of the chromatogram of the STG. The variation of temperatures, “gain” of detector as well as different suppliers of acetonitrile did not show any major effect on the results of the analysis. The “Develosil” column chosen after optimization on the UHPLC did not present similar results on HPLC. This indicates that further optimization work is required on HPLC. The “Develosil” column with the selected parameters was planned to be validated on the UHPLC system, while the same parameters used previously in the analytical method were planned to be verified with the new detector on the HPLC system. Only selectivity and three occasions of intermediate precision were performed. The results have shown that the method is selective. The outcomes presented in this thesis needs further work in order to get approval for the use of the analytical method with the new detector as a valid routine method used in the company.