Electrophoresis of Milk Proteins II. Some Effects of Methods of Preparation on the Electrophoretic Patterns of Whey Proteins

Paper I of this series (11) reported several small differences in the electrophoretic patterns of whey proteins prepared by two methods. These methods were first, a salt-acid procedure devised by Harland and Ashworth (6) and second, a new salt-lyophylized procedure. Both of these patterns were approximately similar to patterns reported by Smith (10) and Deutsch (3) using whey proteins separated from casein with acid or with rennet. Nitschmann and Lehmann (8) found that a-casein, normally a single component, when treated with rennet produced two electrophoretic peaks. Crystalline fl-lactoglobulin also has been reported to be electrophoretically homogeneous (1) at some pH values, but to contain three peaks at others (7). A more extensive electrophoretic comparison of whey proteins prepared by each of the above four methods seemed justified. A *pH of 6.8 is near the natural pH value of fresh milk. Values of 7.8, 6.8, 5.8 and 3.8, therefore, were chosen for this study. Areas, mobilities and isoelectric points all seemed useful bases for comparison. PROCEDURE To insure that any observed differences would be due to methods of preparation and not to differences in the starting milk, the four procedures were used simultaneously on portions of the same samples of milk. Pooled raw whole milk, cooled as soon as milked, was obtained from the Kansas State College Dairy within 3 to 4 hr. after milking. The cream was separated in a Sharples supercentrifuge. The preparations of whey proteins by the salt-acid method and the salt-lyophilized method were exactly as previously described (11). The acid whey and the rennet whey also were dialyzed, lyophilized and redialyzed in a manner similar to that used for the salt-lyophilized whey. For preparation of the acid whey, a procedure modified from Rowland (9) was used. Five hundred ml. of skimmilk were heated to 40 ° C. and placed in a rectangular fruit jar containing a motor stirrer. Fifty ml. of 10 per cent acetic acid were added dropwise, followed after 10-min. standing by 50 ml. of 1.0 N sodium acetate. The mixture then was cooled to about 5 ° C. and filtered after again standing for 2 hr. Eckles et al. (4) described a commercial method for rennet cheese which was