Micellization of Bovine β-Casein Studied by Isothermal Titration Microcalorimetry and Cryogenic Transmission Electron Microscopy

Abstract

The association behavior, critical micellization concentration (CMC), and enthalpy of demicellization (DeltaHdemic) of bovine beta-casein were studied, for the first time by isothermal titration calorimetry, in a pH 7.0 phosphate buffer with 0.1 ionic strength and in pure water. In the buffer solutions, the CMC decreased asymptotically from 0.15 to 0.006 mM as the temperature was raised from 16 to 45 degrees C. DeltaHdemic decreased with increasing temperature between 16 and 28 degrees C but increased from 28 to 45 degrees C. Thermodynamic analysis below 30 degrees C is consistent with the Kegeles shell model, which suggests a stepwise association process. At higher temperatures, this model exhibits limitations, and the micellization becomes much more cooperative. The CMC values in water, measured between 17 and 28 degrees C, decreased with increasing temperature and, expectedly, were higher than those found in the buffer solutions. beta-Casein micelles were visualized and characterized, for the first time in their hydrated state, using advanced digital-imaging cryogenic transmission electron microscopy. The images revealed small, oblate micelles, about approximately 13 nm in diameter. The micelles shape and dimensions remained nearly constant in the temperature range of 24-35 degrees C.

Invited review: Caseins and the casein micelle: Their biological functions, structures, and behavior in foods

“…The association behavior of β-CN shows these characteristics only in part (see Portnaya et al, 2006, and references therein). Thus, association is endothermic but poorly cooperative and the specific heat of micellization changes sign with temperature.…”

What can isothermal titration microcalorimetry experiments tell us about the self‐organization of surfactants into micelles?

“…This is the case of many surfactants such as alkyltrimethylammonium bromides (Beyer et al, 2006), 1-methyl-4-alkylpyridiniuhmalide surfactants (Bijma and Engberts, 1994), alkyl phenol ethoxylates (Dai and Tam, 2003), pluronic F127 , sodium dodecylsulfate (Paula et al, 1995;Dai and Tam, 2004;Liu and Guo, 2007), b-casein (Portnaya et al, 2006), tetronic 304 (Roques et al, 2009), amino-acid-type surfactants (Ohta et al, 2008), oligo(ethylene oxide) alkyl ethers (Heerklotz et al, 1996), the steroid CHAPS (Heerklotz and Seelig, 2000) and bile salts (Hildebrand et al, 2003;Garidel and Hildebrand, 2005). However, some other amphiphilic molecules clearly show the appearance of two extrema, indicating two CMC values.…”

Beta-casein nanocarriers of celecoxib for improved oral bioavailability

“…The N-terminal region of bCN is highly charged while the remainder of the molecule is neutral and hydrophobic (16). At concentrations above its critical micellization concentration (CMC, 0.5-2 mg/mL at pH∼7, 25°C), bCN self-assembles in water into nanometric micelles (∼20-25 nm in diameter) having a hydrophobic core and a hydrophilic corona (17). The bCN structural and physicochemical properties facilitate its application in drug delivery (18)(19)(20), and bCN has been used for encapsulation of hydrophobic chemotherapeutics (21)(22)(23), However, our formulation is quite different from these, and also from the Cx and casein-based formulation described recently (11).…”