Isolation of Casein from Milk

1. Isolation of Casein from Milk ■ Kinds of proteins in milk: • Casein • Lactalbumin • Lactoglobulin They are complete proteins.

2. Casein • The main protein in milk • Present at a concentration of about 35g/l. • It is a phosphoprotein. • Not a single compound, it is a heterogeneous mixture of αS1 , αS2 , β, κ. • Casein exists in milk as the calcium salt, calcium caseinate. Molecular surface model of K-Casein.

3. * Principle ■ The isoelectric point of a protein (I.E.P.) : is the PH at which the molecule is electrically neutral (net charge is zero). • When a protein is charged e.g. with a –ve charge, the –ve molecules repel each other and thus remain in the solution and do not precipitate. • When a protein is neutral at the I.E.P. they tend to precipitate due to the absence of the repelling forces and, the high molecular weight of protein. • Most proteins show minimum solubility at their isoelectric point • PH of milk is 4.5-4.8 (its isoelectric point). • Casein is also insoluble in ethanol and this property is used to remove unwanted fat from the preparation. • Milk is present at a high PH than the isoelectric point of casein

4. 60 60 5 5 55 55 10 10 50 50 45 45 15 15 40 40 20 20 35 35 25 25 30 30 Add 2 ml of CH3COONa Add 2 ml of CH3COOH 10% Measure the pH of the milk 40 ºC 25 ºC 30 ml Distilled Water 30 ml Milk pH=4.5

5. Precipitate ph.ph. Shake well 10 ml 0.1N KOH 80 ml D.w. 10 ml H2O 10 ml 95% alcohol 10 ml ether 10 ml ether-alcohol mix.

6. 0.1N HCl VB VT Blank 10 ml 0.1N KOH 80 ml D.W. 3drops ph.ph.

7. * Calculations 1L (1 N) KOH = Mwt of casein 1 ml (0.1N) KOH = Mwt. Of Casein 10×1000 1ml (0.1N) KOH = 0.105 g casein = 1 ml HCl Volume of Casine = VB - VT (VB-VT) x 0.105 = ? g casein/30ml milk Y = ? g casine % Y = (VB-VT) x 0.105 x 100 30 Samar A. Damiati