High-performance liquid chromatography (HPLC) analysis of the α-tocopherol deposition in the egg yolk: Effect of dl-α-tocopheryl acetate application in feed of breeding hens

A method to quickly measure the content of alpha-tocopherol (alpha-T) in egg yolk was studied by high-performance liquid chromatography and then to further analyze effects of dl-alpha-tocopheryl acetate (alpha-TAC) of different doses in breeding hens' feed on the deposition of alpha-T in the egg yolk. A total of 288 Hyline brown breeding hens, 39 weeks old, were used and divided into six groups. Diets were based on standard layer rations with 0, 20, 50, 80, 160 or 320 mg of alpha-TAC/kg. At the forth weekend, the eggs were selected to measure the content of alpha-T of egg yolk. The following three methods were adopted: dealing with samples of the egg yolk by traditional saponification, ultrasonic processing after methanol extraction and keeping them for 12 hours over night after methanol extraction; methanol as mobile phase at flow rate of 1 ml/min, and using UV-detector to analyze the content of alpha-T in the egg yolk by HPLC at 292 nm wavelength. The method of ultrasonic processing 20 min after methanol extraction was much better than the other two methods in terms of extraction efficiency, with simplicity of operation and less loss of alpha-T. By using this method, a good correlation was shown (r=0.9999) in the observed concentration range. The egg yolk should be extracted at the temperature of 20 C for 10 hours, with a recovery rate of samples from 100.8% to 109%. The deposition of alpha-T in the egg yolk increased significantly when the amount of alpha-TAC was more than 40 mg/kg (P<0.01) and the highest concentration was reached when 320 mg/kg was added. In conclusion, the HPLC method of ultrasonic processing 20 min after methanol extraction is simple, fast and quick-response. It can be used effectively to measure the content of alpha-T in the egg yolk; through adding a heavy close of alpha-TAC into the daily ration, alpha-T egg yolk concentration will be increased and hatching egg rich in alpha-T will be produced, thus improving the hatching efficiency of breeding hens.