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Bovine Milk Lipase. I. Isolation from Skimmilk¹

Department of Food Science and Technology, University of California, Davis

ABSTRACT

A procedure was developed for the isolation of milk lipase from skimmilk: Fresh skimmilk is coagulated with rennet, separating curd, and whey by centrifugation, with most of the lipase accompanying the curd. The lipase is solubilized from the curd by maceration in 1 M NaCl and centrifugation. The supernatant is half-saturated with (NH₄)₂SO₄, the precipitate collected, dissolved, dialyzed, and fractionated on DEAE-cellulose. The lipase-rich fraction is made to 30% (v/v) with dimethylformamide and then to half-saturation with (NH₄)₂SO₄. Under these conditions, the lipase remains soluble and is finally purified by filtration through Sephadex G-200.

The homogeneity of the preparation was established by ultraeentrifugation and starch gel electrophoresis. Specific activity was about 500 times that of skimmilk, with an over-all yield of 10%. The sedimentation coefficient was calculated to be 7.5 S, and the molecular weight about 210,000. With milk fat as substrate, pH optimum was 9.2 and temperature optimum 37 C.

¹ This investigation was supported in part by Public Health Service Research Grant no. UI-00593-04 from the National Center for Urban and Industrial Health.

² Present address: The Agricultural Institute, Moorepark, Fermoy, County Cork, Ireland.