Thermal Gelation of β-Lactoglobulin AB Purified from Cheddar Whey. 1. Effect of pH on Association As Observed by Dynamic Light Scattering

Abstract

Thermal association of β-lactoglobulin AB (β-Lg) purified from Cheddar whey, a major source of commercial whey ingredients, was studied by dynamic light scattering (DLS). The objective was to observe the effect of the process and/or possible micromolecules in this β-Lg source of commercial relevance. Protein solution (8%, w/v) was heated (25−90 °C) at pH 3.5, 7, and 9.0. DLS data were analyzed according to the method of Cumulants for apparent mean diameter and by the Contin method for percentile size distribution. Data indicated gradual change in mean size that started at 35 °C, much below the reported denaturation temperature of 70 °C. Moreover, the percentile distribution of various micrometer- and submicrometer-sized aggregates differed at the different pH values studied, conceivably indicating conformational alteration limiting spatial freedom for intermolecular association. Monomeric/dimeric form (1−9 nm range) was seen only at pH 3.5 at temperatures below 65 °C. The ubiquitous aggregate size range was 100−599 nm (Agg3), and greatest changes in aggregate size and distribution were detected around 70 °C. Micrometer-sized (>1000 nm) aggregates were formed at this temperature and above, concomitant with disappearance of Agg3. On the basis of polydispersity data and rate kinetics dependent on the disappearance of Agg3, the association tendency between 25 and 70 °C was in the order pH 7.0 > pH 3.5 > pH 9.0. Keywords: Thermal; association; gelation; whey; β-lactoglobulin

Influence of Cation Sequestering and pH on Quiescent Thermal Association of .BETA.-Lacto-globulin NB from Fresh Cheddar Whey: An Insight into Gelation Mechanism.

“…In that study and in the present one, exhaustive purification was not carried out to remove trace peptides so as to examine ␤-Lg in its impurity induced conformational state in which it conceivably exists and functions in food systems. Our data (Haque & Sharma, 1997) indicated a greater degree of association at all stages of heating compared to yet another study using chromatographically pure ␤-Lg (Sharma et al, 1996). The starting mean aggregate diameter (MAD) prior to heating (25˚C) was a reflection of the pore size of the membrane (200 nm) used to remove traces of dust from the sample (Haque & Sharma, 1997).…”

“…Furthermore, percentile distribution of various micron (>1000 nm) and submicron-sized (<1000 nm) aggregates differed at the different pH conditions studied, conceivably reflecting conformational alteration that facilitate or limit spatial freedom for intermolecular association. This previous study in this series showed that the smallest level of association, the monomeric/dimeric form (1-9 nm range) was only seen at temperatures below 65˚C at pH 3.5, the ubiquitous aggregate size range was 100-599 nm (Agg3), and greatest changes in aggregate size and distribution were detected around 70˚C (Haque & Sharma, 1997). Large, micron-sized aggregates were formed at this temperature and above, concomitant with disappearance of Agg3.…”

“…Our data (Haque & Sharma, 1997) indicated a greater degree of association at all stages of heating compared to yet another study using chromatographically pure ␤-Lg (Sharma et al, 1996). The starting mean aggregate diameter (MAD) prior to heating (25˚C) was a reflection of the pore size of the membrane (200 nm) used to remove traces of dust from the sample (Haque & Sharma, 1997). It was also noted that gradual stepwise heating changed the MAD starting at 35˚C, much below the reported denaturation temperature of purified ␤-Lg (carried out at a much lower concentrations) of about 70˚C (deWit & Klarenbeek, 1981).…”

“…We reported on the effect of heat and pH on the association properties of ␤-Lg fractionated from fresh Cheddar whey without chelation of cations (Haque & Sharma, 1997). In that study and in the present one, exhaustive purification was not carried out to remove trace peptides so as to examine ␤-Lg in its impurity induced conformational state in which it conceivably exists and functions in food systems.…”

“…Large, micron-sized aggregates were formed at this temperature and above, concomitant with disappearance of Agg3. Based on rate kinetics dependent on the disappearance of Agg3, association tendency at the pre-gelling range of 25-70˚C were 2.45, 3.75, and 2.65¥10 -4 s -1 for pH 3.5, 7, and 9, respectively (Haque & Sharma, 1997).…”