Factors affecting paraoxonase1 activity: standardization of arylesterase assay

Kuruvatti, Sowmya P. and Frank, Elizabeth A. and D'Sourza, Cletus J. M. (2015) Factors affecting paraoxonase1 activity: standardization of arylesterase assay. Asian Journal Of Biochemical and Pharmaceutical Research, 5 (2). pp. 153-161. ISSN 2249-622X

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Abstract

Paraoxonase1 (PON1) is an enzyme associated with High Density Lipoprotein (HDL) and contributes to its cardioprotective properties. Measurement of PON1 activity is an index of quality of HDL and hence its functionality. Because of the promiscuous nature of PON1 and lack of knowledge about the natural substrate of the enzyme, diverse assay methods with different conditions of buffer, cofactors, enzyme dilutions and solvents are used for activity measurements, making comparison between different assays difficult. Our study aims at investigating the effects of various solvents and assay conditions on PON1 activity. Methanol, isopropanol, N, N-dimethyl formamide, acetone and dimethyl sulphoxide were used to dissolve phenylacetate and paraoxon, substrates for PON1 assay. Arylesterase activity of PON1 at various conditions of buffer pH, buffer concentration, Ca2+ concentration and enzyme dilution was measured to determine optimum reaction conditions. Paraoxon and phenylacetate dissolved in methanol gave the highest PON1 activity followed by dimethyl sulphoxide. 10 mM to 100 mM tris-HCl buffer, pH 7.5 to 8.5 containing 1.5 mM to 2 mM calcium chloride, 2 mM phenylacetate, and enzyme dilution of 1:1000 in the final reaction mixture was optimum for arylesterase activity. Non-specific esterase activity was lowest at pH 8.0. Most of the published PON1 assays within the parameters set in this study can be compared with each other since the PON1 activity remained constant.

Item Type: Article
Uncontrolled Keywords: PON1 Assay and Arylesterase and Phenylacetate and Solvent effect.
Subjects: C Chemical Science > Biochemistry
Divisions: Department of > Biochemistry
Depositing User: Shrirekha N
Date Deposited: 18 Jul 2019 05:28
Last Modified: 18 Jul 2019 05:28
URI: http://eprints.uni-mysore.ac.in/id/eprint/5329

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