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In situ monitoring of galactolipid digestion by infrared spectroscopy in both model micelles and spinach chloroplasts

Sahaka, Moulay; Mateos-Diaz, Eduardo; Amara, Sawsan; Wattanakul, Jutarat; Gray, David; Lafont, Dominique; Gontero, Brigitte; Launay, Hélène; Carrière, Frédéric

In situ monitoring of galactolipid digestion by infrared spectroscopy in both model micelles and spinach chloroplasts Thumbnail


Authors

Moulay Sahaka

Eduardo Mateos-Diaz

Sawsan Amara

Jutarat Wattanakul

DAVID GRAY david.gray@nottingham.ac.uk
Professor of Applied Lipid Science

Dominique Lafont

Brigitte Gontero

Hélène Launay

Frédéric Carrière



Abstract

Galactolipids are the main lipids from plant photosynthetic membranes and they can be digested by pancreatic lipase related protein 2 (PLRP2), an enzyme found in the pancreatic secretion in many animal species. Here, we used transmission Fourier-transform infrared spectroscopy (FTIR) to monitor continuously the hydrolysis of galactolipids by PLRP2, in situ and in real time. The method was first developed with a model substrate, a synthetic monogalactosyl diacylglycerol with 8-carbon acyl chains (C8-MGDG), in the form of mixed micelles with a bile salt, sodium taurodeoxycholate (NaTDC). The concentrations of the residual substrate and reaction products (monogalactosylmonoglyceride, MGMG; monogalactosylglycerol, MGG; octanoic acid) were estimated from the carbonyl and carboxylate vibration bands after calibration with reference standards. The results were confirmed by thin layer chromatography analysis (TLC) and specific staining of galactosylated compounds with thymol and sulfuric acid. The method was then applied to the lipolysis of more complex substrates, a natural extract of MGDG with long acyl chains, micellized with NaTDC, and intact chloroplasts isolated from spinach leaves. After a calibration performed with α-linolenic acid, the main fatty acid (FA) found in plant galactolipids, FTIR allowed quantitative measurement of chloroplast lipolysis by PLRP2. A full release of FA from membrane galactolipids was observed, that was not dependent on the presence of bile salts. Nevertheless, the evolution of amide vibration band in FTIR spectra suggested the interaction of membrane proteins with NaTDC and lipolysis products.

Citation

Sahaka, M., Mateos-Diaz, E., Amara, S., Wattanakul, J., Gray, D., Lafont, D., …Carrière, F. (2023). In situ monitoring of galactolipid digestion by infrared spectroscopy in both model micelles and spinach chloroplasts. Chemistry and Physics of Lipids, 252, 105291. https://doi.org/10.1016/j.chemphyslip.2023.105291

Journal Article Type Article
Acceptance Date Mar 9, 2023
Online Publication Date Mar 15, 2023
Publication Date 2023-05
Deposit Date Apr 11, 2023
Publicly Available Date Apr 18, 2023
Journal Chemistry and Physics of Lipids
Print ISSN 0009-3084
Electronic ISSN 1873-2941
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 252
Pages 105291
DOI https://doi.org/10.1016/j.chemphyslip.2023.105291
Keywords Enzyme; Galactolipase; Lipid digestion; Pancreatic lipase related protein 2; Chloroplast; Thin layer chromatography
Public URL https://nottingham-repository.worktribe.com/output/18804686
Publisher URL https://www.sciencedirect.com/science/article/pii/S0009308423000130?via%3Dihub
Additional Information This article is maintained by: Elsevier; Article Title: In situ monitoring of galactolipid digestion by infrared spectroscopy in both model micelles and spinach chloroplasts; Journal Title: Chemistry and Physics of Lipids; CrossRef DOI link to publisher maintained version: https://doi.org/10.1016/j.chemphyslip.2023.105291; Content Type: article; Copyright: © 2023 Elsevier B.V. All rights reserved.

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