Analysis of calcium phosphate cross-linkage in casein micelles by radioactive isotope Professor

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Although calcium and inorganic phosphate (Pi) in bovine casein micelles were at equilibrium with those in soluble form, their properties have not yet been elucidated sufficiently. In this study, the behavior of calcium and Pi in casein micelles was studied using ^<45>Ca and ^<32>P by ultrafiltration, equilibrium dialysis and gel permeation HPLC in the presence of 6M urea. Approximately 30-40% of the calcium and Pi in bovine casein micelles were estimated to be hard to exchanged with those in soluble phase. Then, artificial casein micelles were prepared at a casein concentration of 2.5% with 30-40 mM calcium, 22-27 mM Pi and 10mM citrate using ^<45>Ca and ^<32>P.About 75 and 65% of micellar calcium and Pi in artificial casein micelles were exchanged after dialysis of artificial casein micelles against a simulated milk ultrafiltrate at 4ﾟC for 72h.The percentage of ^<45>Ca and ^<32>P in the fraction of casein aggregates cross-linked by micellar calcium phosphate were 26.9 and 27.6%, which decreased to about 10% after equilibrium dialysis against a simulated milk ultrafiltrate at 4ﾟC for 72h. The proportion of exchanged calcium and Pi in artificial casein micelles during dialysis for 72h were higher at 25ﾟC than at 4ﾟC,suggesting that the exchange of calcium and Pi between diffusible and micellar phases depended on temperature. The effects of calcium and Pi on the formation and properties of casein micelles were also examined using response surface analyzes. The results suggest that the exchange of calcium and phosphate between diffusible and micellar phases proceeds slowly, and a proportion of calcium and Pi in casein micelles is difficult to exchage.