KINETIC-STUDY OF CASEIN MICELLE FORMATION OBSERVED BY A STOPPED-FLOW APPARATUS

The time dependent optical density changes were observed when aqueous solutions (Tris buffer) of sodium casein from milk were mixed with those of calcium chloride by a newly constructed stopped-flow apparatus which was controlled by an 8-bit microcomputer. Two approaches for single exponential decay curve analysis were used, i.e. the Guggenheim analysis and an analysis according to a linear relation between the time and the logarithm of the corresponding absorbance change, at a concentration range between 0.02-0.05 wt% of the casein and in that between 10-30 mmol dm-3 of calcium chloride at 25-degrees-C. The same time constants and reaction amplitudes were obtained from both analyses of the decay curves. The obtained rate parameters were found to be concentration dependent. The result was interpreted by a stepwise reaction mechanism for the formation of a casein aggregate. The rate-determining step was speculated to be the first step of the larger molecular aggregate of the casein according to the concentration dependence of the apparent rate constant and the reaction amplitude. The rate constant for the casein micellization could be approximately estimated as being of the order of 10(4) mol-1 dm3 s-1 at 25-degrees-C. The effects of NaCl, MgCl2, and CaBr2 on casein micelle formation were also examined; it was kinetically confirmed that only calcium cations played an important role in the formation of the casein micelle.