Development of methods for capillary isoelectric focusing of dairy proteins : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Chemistry at Massey University

Abstract

Capillary Isoelectric Focusing (CIEF) is a high-resolution technique which can be applied to the separation and characterisation of complex biological mixtures such as dairy proteins. Although dairy proteins are commonly analysed by traditional gel electrophoresis techniques including 2-Dimensional PAGE, CIEF offers the advantages of reduced analysis times, the ability to handle smaller sample volumes and increased sensitivity with improved separation efficiencies. Several methods for capillary isoelectric focusing of dairy proteins have been developed herein. For the analysis of soluble whey proteins methods that can be used with either UV or mass spectrometry (MS) detection have been set up. For MS detection a coaxial sheath flow interface in conjunction with electrospray ionisation has been utilised. For analysis of the inherently insoluble casein proteins with UV detection denaturing and reducing agents have been introduced into the system. Results have shown very close similarities to those obtained by IEF gels