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

Abstract

Abstract:Casein is the major protein fraction in milk, and its cross-linking has been a topic of scientific interest for many years. Enzymatic cross-linking has huge potential to modify relevant techno-functional properties of casein, whereas non-enzymatic cross-linking occurs naturally during the storage and processing of milk and dairy products. Two size separation techniques were applied for characterisation of these reactions: gel electrophoresis and size exclusion chromatography. This review summarises their separation principles and discusses the outcome of studies on cross-linked casein from the last~20 years. Both methods, however, show limitations concerning separation range and are applied mainly under denaturing and reducing conditions. In contrast, field flow fractionation has a broad separation range and can be easily applied under native conditions. Although this method has become a powerful tool in polymer and nanoparticle analysis and was used in few studies on casein micelles, it has not yet been applied to investigate cross-linked casein. Finally, the principles and requirements for absolute molar mass determination are reviewed, which will be of increased interest in the future since suitable calibration substances for casein polymers are scarce.

Partial Purification and Characterisation of Pectinase Produced by Aspergillus niger LFP-1 Grown on Pomelo Peels as a Substrate

“…Gel filtration chromatography or sometimes referred to as size exclusion chromatography is a chromatography technique that allows the separation of macromolecules based on their hydrodynamic size whereby the larger molecules can only penetrate larger pores (elute earlier) whereas the smaller molecules can access the higher number of pores and retain longer in the column ( Raak et al 2018 ). The present study revealed that the pectinase was purified up to 76 folds with a specific activity of 61.54 U/mg through gel filtration chromatography ( Table 1 ).…”

Bean phenolic compound changes during processing: Chemical interactions and identification

“…Another chromatographic method used is size exclusion chromatography (SEC) and affinity chromatography, which are useful and straightforward to demonstrate the associations between polyphenols and soluble proteins of a matrix by studying the size of the complex formed (Raak et al., 2018; Zhu, 2018). Isothermal titration calorimetry (ITC) (Callies & Daranas, 2016; Zhu, 2018) is an easy and versatile technique to study interactions of macromolecules.…”

Preparation and potential applications of casein–polysaccharide conjugates: a review

“…Direct monitoring of SCN glycation by gel electrophoresis is complicated by the difficulty to resolve the conjugates mixture. Therefore, glycation of SCN with polysaccharides has been followed indirectly by the disappearance of the casein bands . Differences in the glycation degree of casein fractions were evident from two‐dimentional gel electrophoresis profile where α‐CNs and κ‐CN demonstrated the highest and the lowest susceptibility to glycation respectively.…”

“…Size exclusion chromatography has been also used to monitor the changes in the molecular size of casein during glycation …”

“…Size exclusion chromatography has been also used to monitor the changes in the molecular size of casein during glycation. 39,40,61 Fourier-transform infrared (FTIR) spectroscopy was used to differentiate between glycated and non-glycated SCN 63 and to provide complementary information about changes in the protein structure of glycated samples.…”