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Journal of Lipid Research, Vol. 45, 561-566, March 2004
Copyright © 2004 by American Society for Biochemistry and Molecular Biology

Separation and quantitative recovery of mouse serum arylesterase and carboxylesterase activity

Philip W. Connelly^1,*, Graham F. Maguire and Dragomir I. Draganov

^* Departments of Medicine, Biochemistry, and Laboratory Medicine and Pathobiology, University of Toronto, and Department of Medicine, St. Michael's Hospital, Toronto, Canada
Department of Medicine, University of Toronto, Toronto, Canada
Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI

¹ To whom correspondence should be addressed. e-mail: connellyp{at}smh.toronto.on.ca

Paraoxonase-1 (PON1) is known to be associated with high density lipoproteins. We optimized buffer conditions to obtain quantitative recovery of PON1 (arylesterase) activity and analyzed the distribution of PON1 in mice using a combination of size-exclusion chromatography and ultracentrifugation. Size-exclusion chromatography of mouse serum separated the esterase activity into two peaks, one overlapping the high density lipoproteins and a second peak of lower molecular weight, consistent with serum carboxylesterase, which accounted for 20% of the total esterase activity of normal mouse serum. Using conditions for the quantitative recovery of arylesterase activity, we fractionated serum by ultracentrifugation into d < 1.21 g/ml, d < 1.25 g/ml, d > 1.21 g/ml, and d > 1.25 g/ml fractions. We observed that PON1 arylesterase activity and mass were isolated in the d < 1.21 g/ml fraction and that serum carboxylesterase was recovered in the d > 1.25 g/ml fraction. The significance of the confounding of PON1 arylesterase activity by serum carboxylesterase was demonstrated by studying mice challenged with a high-fat, high-cholate diet for 14 days. It was shown that all of the decrease in arylesterase activity in response to this diet is attributable to the HDL-associated arylesterase activity (PON1).

We conclude that mouse PON1 is quantitatively associated with high density lipoproteins. The contribution of serum carboxylesterase to the total esterase activity significantly confounds the interpretation of total arylesterase activity in mouse serum.

Abbreviations: CEase, serum carboxylesterase; DTPA, diethylenetriamine penta-acetic acid; FPLC, fast-protein liquid chromatography; PON1, paraoxonase-1

Supplementary key words paraoxonase-1 • high density lipoproteins • diet

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