Fractionation of the Lipoproteins of the Fat Globule Membrane from Cream

SUMlVIARYThe membrane material obtained by freezing washed cream has been fractionated from suspensions in 2% sodium desoxyeholate into five sedimentable fractions representing 75% of the total material and one soluble fraction (25%).These fractions were clearly differentiated by their appearance and sedimentation behavior.Fractions A and D contained most of the xanthine oxidase activity and were about 32% lipid.Fraction B contained most of the alkaline phosphatase activity and was about 65% lipid.The other fractions were over 70% lipid.In Fractions A, B, and D, the lipid was 15% phospholipid; in Fractions C and E, it was 5-6% phospholipid; and in the supernatant, it was 50% phospholipid.The literature on the membrane of the bovine fat globule before 1955 has been reviewed comprehensively (11).Extensive fractionation and characterization studies have been carried out on the proteins of membrane material obtained after fat extraction [e.g., Herald and Brunner (7, 9)].However, only limited work has been done on the lipoproteins as such.Enzymic and chemical analyses have been performed on milk microsomes, a small fraction of membrane material that sedimented in 15 rain.at 50,000 × g but not at 10,000 × g (3, 4).Also, material obtained by churning unwashed cream, in which it is, therefore, difficult to determine the origin of the various fractions, has been fractionated with ammonium sulfate, and membrane material from washed cream was divided arbitrarily into two fractions by centrifugation (14).These fractions were analyzed only for alkaline phosphatase activity.The present paper reports the fractionatiou of the lipoproteins of most of the membrane material obtained from washed cream by freezing (8), and characterizes the fractions by enzyme content and chemical analyses. MATERIALS AND ~[ETHODSFresh bulk milk, kept at a temperature higher than 30 ° C,, was separated within 3 hr.after milking.The cream was washed and membrane material prepared essentially as described previously (8).The cream, approximately 50% fat, was frozen and stored at --10 ° C. in test tubes (1.4 cm.i.d.) for about 20 hr. ; stored at --40 ° C. if not required immediately, never longer than 4 wk.After thawing the cream and diluting it fourfold with 0.01% gelatin solution, the membrane material was sedimented at 78,410 × g for 1 hr.