Insoluble casein

It has been shown that the major part of Ca and Mg are found in the soluble fraction (whey) of human milk, whereas only small amounts are present in insoluble caseins or in fat [11-13]. Calcium and Mg speciation studies in milk whey using size exclusion chromatography (SEC) combined with inductively coupled plasma atomic emission spectrometry (ICP-AES) [14] indicated that they were preferably associated with the milk nonprotein fraction. A further work [15] showed similar chromatographic profiles by SEC hyphenated with inductively coupled plasma mass spectrometry (ICP-MS), with Ca and Mg eluting in the lowmolecular-weight region (LMW) (<1.4 kDa). Thus, there seem to be weak associations, if any, of Ca2+ or Mg2+ with higher molecular mass biocompounds of milk. 

Separation of Caseinogen by Bennin.— To 5 ml. of milk add 5-10 drops of an active preparation of the enzyme rennin, obtained from the gastric mucosa of calves. Mix and incubate at 40°. In a few minutes the milk will solidify owing to the conversion of caseinogen into insoluble casein (paracasein), and the tube may be inverted without spilling the contents. 

In the presence of calcium it converts soluble caseinogen into insoluble casein (paracasein), the optimal pH being 6-0-6 5. This causes the coagulation of milk. 

Minerals, particularly Bentonite, ate used to remove proteins that tend to cause haze in white wines. The natural tannin of ted wines usually removes unstable proteins from them. Excess tannin and related phenols can be removed and haze from them prevented by addition of proteins or adsorbents such as polyvinylpyttohdone. Addition of protein such as gelatin along with tannic acid can even be used to remove other proteins from white wines. Egg whites or albumen ate often used to fine ted wines. Casein can be used for either process, because it becomes insoluble in acidic solutions like wines. 

Soybean Protein Isolates. Soybean protein isolates, having a protein content of >90 wt%, are the only vegetable proteins that are widely used in imitation dairy products (1). Most isolates are derived from isoelectric precipitation, so that the soybean protein isolates have properties that are similar to those of casein. They are insoluble at thek isoelectric point, have a relatively high proportion of hydrophobic amino acid residues, and are calcium-sensitive. They differ from casein in that they are heat-denaturable and thus heat-labile. The proteins have relatively good nutritional properties and have been increasingly used as a principal source of protein. A main deterrent to use has been the beany flavor associated with the product. Use is expected to increase in part because of lower cost as compared to caseinates. There has been much research to develop improved soybean protein isolates. 

In the rennet coagulation process fresh skimmed milk is adjusted to a pH of six and about 40 ounces of a 10% solution of rennet are added per 100 gallons of milk. The initial reaction temperature is about 35°C and this is subsequently raised to about 60°C. The coagulation appears to take place in two stages. Firstly the calcium caseinate is converted to the insoluble calcium paracaseinate and this then coagulates. 

Following research by Tossavainen et al. (1986), extrusion is often used to produce insoluble acid casein from skim milk powder and to convert acid casein into sodium caseinate. The procedure is faster than batch mixing (Akdogan, 1999). 

Kobayashi S, Koga K, Hayashida O, Nakano Y and Hasegawa Y (1990) Specific inhibition of insoluble glucan synthase (GTF-1) by Maillard reaction products from casein and albumins. Agric Biol Chem 54, 1417-1424. 

In addition, gelatin peptides have shown to accelerate absorption of dietary calcium in animal models increasing calcium bioavailability (Kim et al., 1998). Jung et al. (2006) reported that fish bone peptides (FBP) could inhibit the formation of insoluble Ca salts in neutral pFI. During the experimental period, Ca retention was increased and loss of bone mineral was decreased by FBP II supplementation in ovariectomized rats. The levels of femoral total Ca, bone mineral density, and strength were also significantly increased by the FBP diet to levels similar to those of the casein phosphopeptide diet group. 

It was reported by Jourdain et al. (2008) that it was impossible to make a proper emulsion with sodium caseinate alone at pH = 4 because of the very low protein solubility close to pI. Nevertheless, a fine stable emulsion (c/43 1.0 pm) could easily be prepared when dextran sulfate was present in a modest amount. In contrast, the monodisperse bilayer emulsion could not be prepared at low pH because the primary caseinate-stabilized emulsion was already either partially aggregated (pH = 2) or completely insoluble (pH = 4) under these conditions. 

Co-preclpltate is an insoluble milk protein product that is produced by heating skinimllk to high temperatures ( > 90 C) to denature the whey proteins and complex them with the casein micelles. The heated system is subsequently adjusted to isoelectric point conditions of pH 4.5-5 to precipitate the complexed whey protein-casein micelles, centrifuged or filtered to recover the precipitate, washed and dryed. The resulting product, which is virtually insoluble, exhibits only minor functionality in most typical emulsification applications. 

Casein may be coagulated and recovered as rennet casein by treatment of milk with selected proteinases (rennets). However, one of the caseins, K-casein, is hydrolysed during renneting and therefore the properties of rennet casein differ fundamentally from those of acid casein. Rennet casein, which contains the colloidal calcium phosphate of milk, is insoluble in water at pH 7 but can be dissolved by adding calcium sequestering agents, usually citrates or polyphosphates. It has desirable functional properties for certain food applications, e.g. in the production of cheese analogues. 

At all temperatures, asl-CN B and C are insoluble in calcium-containing solutions and form a coarse precipitate at Ca2+ concentrations greater than about 4 mM. asl-CN A, from which the very hydrophobic sequence, residues 13-26, is deleted, is soluble at [Ca2+] up to 0.4 M in the temperature range 1-33°C. Above 33°C, it precipitates but redissolves on cooling to 28°C. The presence of asl-CN A modifies the behaviour of asl-CN B so that an equimolar mixture of the two is soluble in 0.4 M Ca2+ at 1°C asl-CN B precipitates from the mixture at 18°C and both asl-CN A and B precipitate at 33°C. aBl-CN A does not form normal micelles with K-casein. Since asl-CN A occurs at very low frequency, these abnormalities are of little consequence in dairy processing but may become important if the frequency of asl-CN A increases as a result of breeding practices. 

The as2-caseins are also insoluble in Ca2+ (above about 4mM) at all temperatures, but their behaviour has not been studied in detail. 

Casein is soluble at high concentrations of Ca2+ (0.4 M) at temperatures below 18°C, but above 18°C /9-casein is very insoluble, even in the presence of low concentrations of Ca2 + (4 mM). Ca-precipitated /9-casein redissolves readily on cooling to below 18°C. About 20°C is also the critical temperature for the temperature-dependent polymerization of /9-casein and the two phenomena may be related. 

Certain of the milk salts (e.g. chlorides, and the salts of sodium and potassium) are sufficiently soluble to be present almost entirely in the dissolved phase. The concentration of others, in particular calcium phosphate, is higher than can be maintained in solution at the normal pH of milk. Consequently, these exist partly in soluble form and partly in an insoluble or colloidal form associated with casein. The state and distribution of these salts has been extensively reviewed by Pyne (1962) and Holt (1985). 

The dividing line between soluble and colloidal is somewhat arbitrary, its exact position depending very much on the method used to achieve separation. However, a fairly sharp separation between the two phases is not difficult since the insoluble salts occur mainly associated with the colloidal casein micelles. 

Figure 10.23 Water-insoluble and water-soluble peptides derived from asl-casein (A), as2-casein (B) or -casein (C) isolated from Cheddar cheese DF = diafiltration. The principal chymosin, plasmin and lactococcal cell-envelope proteinase cleavage sites are indicated by arrows (data from T.K. Singh...

Minerals found in milk which are insoluble remain in water in the curd and are more concentrated in the cheese than in milk. About two-thirds of the calcium and one-half of the phosphorus of milk remains in cheese. A major portion of the milk calcium is retained in the curd of cheese made with coagulating enzymes. Acid coagulation alone results in the loss of portions of both calcium and phosphorus salts in the acid whey, since these minerals are more soluble in the acidic medium. Most milk fat and fat-soluble vitamins are retained in the curd, but a considerable amount of water-soluble vitamins is lost during cheese manufacture. Retention of part of some B-complex vitamins in curd is due to their extended association with casein in the original milk. 

The configuration and association of the minor /3-caseins have not been investigated to any extent, but from their primary structure one can conclude that /3-casein X-4P-(f 1-28), -(f 1-105), and -(f 1-107) and /3-casein X-lP-(f 29-105) and -(f 29-107) are more hydrophilic than the main /3-casein component and that /3-casein X-lP-(f 29-209) and /3-casein-(f 106-209) and -(f 108-209) will be more hydrophobic. The hy-drophobicities of the last three caseins are the highest of all the caseins, with calculated Bigelow hydrophobicities of 1386 to 1511 (Swais-good 1973). Therefore, we would expect these to demonstrate a strong temperature-dependent association /3-casein B-(f 106-209) and -(108— 209) are insoluble at pH 8.0 and 25°C but soluble at 3°C, while 0-casein A-(f 108-209) is soluble at both temperatures. Sedimentation studies... 

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