Determination of Phospholipids in Milk by HPLC with Evaporative Light Scattering Detector: Optimization and Validation - Juniper Publishers

Determination of Phospholipids in Milk by HPLC with Evaporative Light Scattering Detector: Optimization and Validation



Authored by Rodríguez-Otero JL

Phospholipids are part of the milk fat globule membrane. Functional and technological properties make them interesting for the development of functional foods or as ingredients of technological interest. In this study an analytical procedure for determination of major phospholipids in milk (phosphatidyl Ethanolamine, phosphatidylinositol, phosphatidyl Choline, phosphatidyl Serine and sphingomyelin) by HPLC with evaporative light scattering detector was optimized and validated. The best calibrations were achieved when an exponential model was applied; the squared correlation coefficients show a satisfactory linearity ranging from 0.975 to 0.993. The limit of detection for the different phospholipids ranged from 0.17μg for phosphatidyl Serine to 0.61μg for phosphatidyl Ethanolamine; and the limit of quantification from 0.40μg for phosphatidyl Serine to 1.26μg for phosphatidyl Ethanolamine. The intra-day and inter-day precision of the method was below 10% for all the compounds except phosphatidylinositol in the intra-day assay with a value of 12.4%. However, the values of intra-day and inter-day repeatability show that the variations, due to the extraction procedure, do not depend whether the assays were performed after short intervals of time or not. The recovery ranged from 74% for phosphatidil Serine to 112% for phosphatidilcoline. The addition of formic acid to the mobile phase in order to achieve an acidic buffer extends the column life considerably.  

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