HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Chandan, R. C. Articles by Shahani, K. M. Search for Related Content PubMed Articles by Chandan, R. C. Articles by Shahani, K. M.

Purification and Characterization of Milk Lipase. I. Purification^{1, 2, 3,}

Department of Dairy Husbandry, University of Nebraska, Lincoln

ABSTRACT

Milk lipase was isolated successfully from clarifier sediment. The method involves preparation of acetone powder of the sediment, extraction of the enzyme with water, and fractional precipitation with ammonium sulfate and acetone. The enzyme with some minor contaminants selectively precipitates between 35 and 45% saturation of ammonium sulfate and at 45% (v/v) concentration of acetone. Finally, the enzyme is purified by chromatography on a Sephadex G-50 column. The purity and homogeneity of the enzyme was confirmed with the ultracentrifugal and gel electrophoretic techniques. The specific activity of the purest lipase fraction had a purification of about 88-fold on the milk protein basis, with an over-all yield of about 22%. The purification was 250-fold on the basis of milk solids and 2,600-fold on the basis of milk. The purified enzyme gave a typical protein ultraviolet absorption spectrum with maximum absorption at 277 mµ and minimum at 251 mµ, with A₂₇₇/A₂₅₁ ratio being 2.34. Chemical analysis showed that the enzyme contained 49.63% carbon, 7.51% hydrogen, 14.33% nitrogen, 1.04% sulfur, and 0.26% phosphorus.

The purified enzyme seems to be distinctly different in its characteristics from other known milk proteins and enzymes. In milk it probably exists as a physically adsorbed unit on casein particles.

¹ Published with the approval of the Director as paper no. 1206, Journal Series, Nebraska Agricultural Experiment Station, Lincoln.

² Data taken from a thesis presented by R. C. Chandan in partial fulfillment of the requirements of the Doctor of Philosophy degree, University of Nebraska, 1963.

³ The following abbreviations are used: CMC, carboxymethyl cellulose; DEAE cellulose, diethylaminoethyl cellulose; Tris, tris (hydroxymethyl) aminomethane; EDTA, ethylene-diaminetetraacetic acid.