Heat-Induced Denaturation/Aggregation of -Lactoglobulin as Affected by N-Ethylmaleimide, NaCl, CaCl₂ and pH

The degree of denaturation/unfolding and subsequent aggregation dictate the functional attributes of -lactoglobulin ( -lg), such as increased water-binding, gelation or general structure/texture development. The heat-induced denaturation/aggregation behaviour of -lg was examined in the presence of water, NaCl, CaCl2 and/or N-ethylmaleimide in the pH range 5.0-7.0. It was shown that rates of unfolding and aggregation of -lg, were pH dependent, with aggregation being rate limiting at pH 7.0 and unfolding rate limiting at pH 5.0. Results suggested that heat-induced -lg aggregation at pH 5 under low ionic strength conditions did not initially involve thiol-disulphide interchange reactions. However, secondary formation of thiol-disulphide interchange linkages was time dependent. The addition of NaCl to -lg solutions prior to heating inhibited denaturation/aggregation on heating at pH 5.0-6.0 but promoted these reactions at pH 6.0-7.0. Over a certain pH range, the presence of N-ethylmaleimide promoted the interaction between unfolded -lg molecules by non-specific forces. The addition of N-ethylmaleimide to -lg solutions prior to heating in the presence of CaCl2, resulted in similar denaturation/aggregation levels at pH values between 5.0 and 5.75 but slightly reduced levels at pH values between 6.0 and 7.0. Results showed the significance of non-specific forces in driving the heat-induced aggregation of -lg over a range of pH values, which has relevance to acidified food systems.