title: Excessive centrifugal fields damage high density lipoprotein.
abstract: HDL is typically isolated ultracentrifugally at 40,000 rpm or greater, however, such high centrifugal forces are responsible for altering the recovered HDL particle. We demonstrate that this damage to HDL begins at approximately 30,000 rpm and the magnitude of loss increases in a rotor speed-dependent manner. The HDL is affected by elevated ultracentrifugal fields resulting in a lower particle density due to the shedding of associated proteins. To circumvent the alteration of the recovered HDL, we utilize a KBr-containing density gradient and a lowered rotor speed of 15,000 rpm to separate the lipoproteins using a single 96 h centrifugation step. This recovers the HDL at two density ranges; the bulk of the material has a density of about 1.115 g/ml, while lessor amounts of material are recovered at >1.2 g/ml. Thus, demonstrating the isolation of intact HDL is possible utilizing lower centrifuge rotor speeds.
authors.affiliation: Department of Chemistry and Biochemistry University of California, Los Angeles, Los Angeles, CA 90095.
authors.forename: William H
authors.initials: WH
authors.lastname: Munroe
authors.affiliation: Department of Chemistry and Biochemistry University of California, Los Angeles, Los Angeles, CA 90095.
authors.forename: Martin L
authors.initials: ML
authors.lastname: Phillips
authors.affiliation: Department of Chemistry and Biochemistry University of California, Los Angeles, Los Angeles, CA 90095 Molecular Biology Institute, University of California, Los Angeles, Los Angeles, CA 90095.
authors.forename: Verne N
authors.initials: VN
authors.lastname: Schumaker
mesh_terms: D002499:Centrifugation, Density Gradient / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D007700:Kinetics; D008075:Lipoproteins, HDL / Q000737:chemistry / Q000302:isolation & purification*; D014461:Ultracentrifugation / Q000379:methods*
chemical_list: D008075:Lipoproteins, HDL
keywords: analytical ultracentrifugation; density gradient ultracentrifugation; high density lipoprotein isolation; high density lipoprotein/structure; lipoproteins/assembly; lipoproteins/kinetics; shedding; ultracentrifugation