REVERSED-PHASE LIQUID CHROMATOGRAPHY OF EGG WHITE PROTEINS. OPTIMIZATION OF OVALBUMIN ELUTION

This paper is an experimental contribution to the optimization of ovalbumin elution in RP-HPLC with a C4 support. The effects of TFA concentration, gradient duration, flow rate, and the nature of the solvent were studied using an experimental design. The results highlighted the major role of TFA concentration in the recovery yield of ovalbumin, with an optimum concentration range which is very narrow and much lower (0.025 %) than the levels usually applied (0.1 %). In this respect, ovalbumin seems to behave in a specific way. Among the four solvents tested, i.e. acetonitrile, isopropanol, ethanol, and a mixture of these three, the first one is definitely the most satisfactory in the area of recovery yield and efficiency. By modifying the elution conditions alone, the recovery yield of ovalbumin may range from 50 to 85 %. Finally, this work has enabled us to define the chromatographic conditions for quantitative egg white analysis of its major proteins in only six minutes and with minimal subsequent ghosting effects.