Determination of α-tocopherol by reversed-phase HPLC in feed and animal-derived foods without saponification

Background: The analysis of α-tocopherol in feed and animal-derived foods usually involves a saponification step. However, since saponification often leads to losses of α-tocopherol, a method for the determination of α-tocopherol in feed and in animal-derived foods was developed without a saponification step.

Results: In this method, α-tocopherol is extracted with hot ethanol and the co-extracted fat is removed by centrifugation. Removal of the fat fraction is made possible by the addition of water, to achieve an ethanol:water ratio of 40:7, followed by cooling on ice before centrifugation. This procedure allows removal of the fat fraction, while α-tocopherol is retained. Matrices differing in gross composition and α-tocopherol content were analyzed: fresh pork, cooked ham, subcutaneous fat, liver, egg yolk, milk and a compound pig feed. Higher α-tocopherol concentrations were found for this novel method compared to a conventional method with saponification, particularly for subcutaneous fat (P < 0.05). Recoveries were higher (P < 0.05) for the novel method (82-103%), compared to the saponification method (66-90%; for subcutaneous fat < 25%).

Conclusion: Determining α-tocopherol in feed and animal-derived foods using pure ethanol without saponification results in higher extraction yields and recoveries compared to the saponification method.