A facile assay to monitor secretory phospholipase A2 using 8-anilino-1-naphthalenesulfonic acid

Vivek, H. K. and Swamy, S. G. and Priya, B. S. and Sethi, G. and Rangappa, K. S. and Nanjunda Swamy, S. (2014) A facile assay to monitor secretory phospholipase A2 using 8-anilino-1-naphthalenesulfonic acid. Analytical Biochemistry, 461. pp. 27-35. ISSN 0003-2697

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Official URL: https://doi.org/10.1016/j.ab.2014.05.024


Secretory phospholipases A2 (sPLA2s) are present in snake venoms, serum, and biological fluids of patients with various inflammatory, autoimmune and allergic disorders. Lipid mediators in the inflammatory processes have potential value for controlling phospholipid metabolism through sPLA2 inhibition. Thus, it demands the need for screening of potential leads for sPLA2 inhibition. To date, sPLA 2 activity has been assayed using expensive radioactive or chromogenic substrates, thereby limiting a large number of assays. In this study, a simple and sensitive NanoDrop assay was developed using non-fluorogenic and non-chromogenic phospholipid substrate 1,2-dimyristoyl-sn-glycero-3- phosphocholine (DMPC) and 8-anilino-1-naphthalenesulfonic acid (ANS) as interfacial hydrophobic probe. The modified assay required a 10ng concentration of sPLA2. ANS, as a strong anion, binds predominantly to cationic group of choline head of DMPC through ion pair formation, imparting hydrophobicity and lipophilicity and resulting in an increase in fluorescence. Triton X-100 imparts correct geometrical space during sPLA2 catalyzing DMPC, releasing lysophospholipid and acidic myristoyl acid, which in turn alters the hydrophobic environment prevailing around ANS-DMPC, which leads to weakening of the electrostatic ion pair interaction between DMPC and ANS ensuing decrease in fluorescence. These characteristic fluorescence changes between DMPC and ANS in response to sPLA2 catalysis are well documented and validated in this study.

Item Type: Article
Uncontrolled Keywords: controlled study, unclassified drug, animal, Animals, chemistry, antagonists and inhibitors, metabolism, priority journal, procedures, enzyme activity, 1, enzyme assay, article, hydrophobicity, calcium, catalysis, Daboia russellii, Russell's Viper, Phospholipases A2, pH, Calcium, Fluorescence, lipophilicity, quercetin, calibration, Calibration, Quercetin, spectrophotometer, choline, Hydrogen-Ion Concentration, chromogenic substrate, Elapidae, fluorescent dye, Fluorescent Dyes, Spectrometry, 2 dimyristoyl sn glycero 3 phosphocholine, 8 anilino 1 naphthalenesulfonic acid, 8-anilino-1-naphthalenesulfonic acid, Anilino Naphthalenesulfonates, ascites fluid, critical micelle concentration, deoxycholic acid, Deoxycholic Acid, dimyristoylphosphatidylcholine, Dimyristoylphosphatidylcholine, Enzyme Assays, fluorescence, lysophospholipid, micelle, Micelles, octoxinol, Octoxynol, phosphorylcholine, pleura fluid, Secretory, secretory phospholipase A2, spectrofluorometry, triton x 100
Subjects: C Chemical Science > Chemistry
Divisions: Department of > Chemistry
Depositing User: Arshiya Kousar Library Assistant
Date Deposited: 31 Aug 2019 05:20
Last Modified: 31 Aug 2019 05:20
URI: http://eprints.uni-mysore.ac.in/id/eprint/4295

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