Acid-induced gelation of enzymatically cross-linked caseinate in different ionic milieus

Size Separation Techniques for the Characterisation of Cross-Linked Casein: A Review of Methods and Their Applications

“…However, when injecting standards of different casein types slightly different elution following the order β-casein < αS1-casein < κ-casein can be seen (Figure 10b), which is in accordance to their respective molar mass (Table 1) and allows at least rough hints on their different polymerisation velocity [148]. Using Superdex 200, the monomer peak is clearly shifted to higher elution volumes with higher cross-linking of non-micellar casein by TGase (Figure 10a), which suggests the presence of remaining κ-casein, as also observed by SDS-PAGE (see Section 3.3).…”

“…SEC column materials differ in composition and pore volume. The column most frequently used for studying cross-linked casein is Superdex 200, e.g., [30,44,47,48,54,100,115,131,[134][135][136][137][138][139][140][141][142][143][144][145][146][147][148], which is provided by GE Healthcare (Uppsala, Sweden). The column material is based on agarose/dextran with a ratio of pore volume to void volume of V i /V o = 1.7 and exhibits a fractionation range from 10 to 600 kg/mol for globular proteins [128].…”

“…For instance, the highest number of casein dimers in TGase-catalysed polymerisation was observed in moderately cross-linked samples, i.e., at high levels of residual monomers (>50%), indicating that dimers as well as smaller oligomers are good substrates for further cross-linking by TGase [32,78,146]. Moreover, the polymerisation degree (sometimes also referred to as oligomerisation degree) was calculated in several studies from size exclusion chromatograms by relating peak areas of cross-linked caseins to the entire sample area [30,32,44,47,48,53,78,[134][135][136][146][147][148]. Because reference casein samples often reveal a low level of polymerised casein (see Figures 9 and 10), the polymerisation degree after cross-linking was also expressed as the difference between cross-linked samples and reference [137][138][139]144,145].…”

“…Without calculating [CLAA] min , Pellegrino et al [43] suggested that intramolecularly formed lysinoalaine caused conformational changes of β-casein monomers as the monomer peak in SEC had a different shape compared to the reference. Similarly, little changes in the polymer peak while the content of N-ε-(γ-glutamyl)-lysine still increased during extended treatment of non-micellar casein with TGase was taken as an indicator for extensive internal cross-linking in polymers that had reached a maximum size [30,31,148].…”

Addition of crude tiger nut protein extract affects stiffness of enzymatically cross‐linked dairy proteins

“…Nevertheless, it can be seen from earlier elution and higher peak intensities (elution time of about 15 min) that larger casein polymers were formed in the presence of tiger nut protein extracts. A similar observation was made in a previous study where the formation of larger casein polymers was triggered by the addition of ions (Raak et al ., ). The presence of ions during enzymatic cross‐linking is known to screen protein net charge, favouring casein self‐association and thus, the formation of larger polymers as a consequence of cross‐linking molecules within particular aggregates (O'Connell & de Kruif, ; Raak et al ., ).…”