Abstract
We studied the effect in vitro of various concentrations of Triton WR-1339 on normolipidemic canine plasma and on the high density lipoproteins (HDL) isolated from this plasma by ultracentrifugation. As a preamble to this study, we established that Triton WR-1339 has a unimer molecular weight of 4,500, a micellar molecular weight of 180,000, and a critical micellar concentration (CMC) of 0.018 mM or 0.008 g/dl. Above its CMC, Triton WR-1339 in concentrations between 2 and 10 mg/ml induced concentration-dependent structural changes in HDL which were characterized by a progressive displacement of apoA-I from the HDL surface without loss of lipids. The addition of Triton WR-1339 to the HDL particles modified their electrophoresis mobility and caused an increase in size (95 +/- 5 A to 114 +/- 7 A). At the extreme Triton WR-1339 concentrations utilized in these studies (10 mg/ml) disruption of the HDL particles occurred; at this stage, the original, relatively homogeneous, spherical HDL particles were replaced by a heterogeneous population ranging in size between 50 and 250 A, representing complexes of Triton WR-1339 with lipids essentially free of apoA-I which could be sedimented by ultracentrifugation. The effects of Triton WR-1339 on whole plasma or isolated HDL were comparable. These studies indicate that Triton WR-1339 in vitro alters HDL in a concentration-dependent manner and that these changes vary from a displacement of apoA-I from the HDL surface to a state where all lipids are solubilized into the Triton WR-1339 micellar phase and are driven away from the protein moiety.(ABSTRACT TRUNCATED AT 250 WORDS)
Highlights
A h & We studied the effect in vitro of various concentrations the HDLparticles were recognizedto be senof Triton WR-1339 on normolipidemic canine plasma and on the high density lipoproteins (HDL) isolated from this plasma by ultracentrifugation.As a preambleto this studyw, e established that Triton WR-1339 has a unimer molecular weight of4,500, a micellar molecular weight of 180,000, and a critical micellar concentration (CMC)of 0.018 mM or 0.008 g/dl
At the extreme Triton WR-1339 concentrations utilized in these studies (10 mg/ml) disruption of the high density lipoproteinsd l .063-1.2 l g/ml (HDL) particlesoccurred;at this staget,he original, relatively homogeneous, spherical HDL particles were replacebdy a heterogenous population ranging in size between 50 and 250 A, representing complexesofTriton WR-1339 with lipidsessentially sitive to the action of the detergent bothin vitro and in vivo (1 1)
The effectsof Triton WR-1339 onwholeplasma or isolated HDL were comparab1e.M These studies indicate that Triton WR-1339 in vitroalters HDLin a concentration-dependent manner and that these changes vary from a displacement of apoA-Ifrom the HDLsurfaceto a state wherealllipids are amined someof the physicochemical properties ofTriton WR-1339 in solution
Summary
After incubation with Triton WR-I339, theplasma was separated by the singlestep density gradient ultracentrifugation as previously described [19]. The effluentswere monitored at 280 nm and collected as 400-p1 fractions. Gel filtration was conducted in glass columns (2.5 X 70 cm) packed with Sepharose 4B (Pharmacia Fine ChemicalUs,ppsala, Sweden). The columns were eluted with 0.05 M phosphate, pH 7.2, at a flow rate of 20 ml/hr at 6°C. Eluates were monitored at 280 nm or at 278 nm
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