Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species
In addition to cell membrane phospholipids, <i>Actinobacteria</i> in the order <i>Corynebacteriales</i> possess a waxy cell envelope containing mycolic acids (MA). In optimized culture condition, some species can also accumulate high concentrations of intracellular triacylgly...
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doaj-99ce30f637e043b39b885517aee31dd72020-11-25T00:32:38ZengMDPI AGInternational Journal of Molecular Sciences1422-00672020-02-01215167010.3390/ijms21051670ijms21051670Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> SpeciesAkhikun Nahar0Anthony L. Baker1David S. Nichols2John P. Bowman3Margaret L. Britz4Tasmanian Institute of Agriculture, University of Tasmania, Hobart, TAS 7005, AustraliaTasmanian Institute of Agriculture, University of Tasmania, Hobart, TAS 7005, AustraliaCentral Science Laboratory, Division of Research, University of Tasmania, Hobart, TAS 7005, AustraliaTasmanian Institute of Agriculture, University of Tasmania, Hobart, TAS 7005, AustraliaTasmanian Institute of Agriculture, University of Tasmania, Hobart, TAS 7005, AustraliaIn addition to cell membrane phospholipids, <i>Actinobacteria</i> in the order <i>Corynebacteriales</i> possess a waxy cell envelope containing mycolic acids (MA). In optimized culture condition, some species can also accumulate high concentrations of intracellular triacylglycerols (TAG), which are a potential source of biodiesel. Bacterial lipid classes and composition alter in response to environmental stresses, including nutrient availability, thus understanding carbon flow into different lipid classes is important when optimizing TAG synthesis. Quantitative and qualitative analysis of lipid classes normally requires combinations of different extraction, derivatization, chromatographic and detection methods. In this study, a single-step thin-layer chromatography-flame ionization detection (TLC-FID) technique was applied to quantify lipid classes in six sub-Antarctic <i>Corynebacteriales</i> strains identified as <i>Rhodococcus</i> and <i>Williamsia</i> species. A hexane:diethyl-ether:acetic acid solvent system separated the total cellular lipids extracted from cells lysed by bead beating, which released more bound and unbound MA than sonication. Typical profiles included a major broad non-polar lipid peak, TAG and phospholipids, although trehalose dimycolates, when present, co-eluted with phospholipids. Ultra-performance liquid chromatography-tandem mass-spectrometry and nuclear magnetic resonance spectroscopy detected MA signatures in the non-polar lipid peak and indicated that these lipids were likely bound, at least in part, to sugars from cell wall arabinogalactan. Waxy esters were not detected. The single-solvent TLC-FID procedure provides a useful platform for the quantitation and preliminary screening of cellular lipid classes when testing the impacts of growth conditions on TAG synthesis.https://www.mdpi.com/1422-0067/21/5/1670tlc-fidiatroscan rodsnmrlipid extractionmycolic acids<i>williamsia</i><i>rhodococcus</i> |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Akhikun Nahar Anthony L. Baker David S. Nichols John P. Bowman Margaret L. Britz |
spellingShingle |
Akhikun Nahar Anthony L. Baker David S. Nichols John P. Bowman Margaret L. Britz Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species International Journal of Molecular Sciences tlc-fid iatroscan rods nmr lipid extraction mycolic acids <i>williamsia</i> <i>rhodococcus</i> |
author_facet |
Akhikun Nahar Anthony L. Baker David S. Nichols John P. Bowman Margaret L. Britz |
author_sort |
Akhikun Nahar |
title |
Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species |
title_short |
Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species |
title_full |
Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species |
title_fullStr |
Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species |
title_full_unstemmed |
Application of Thin-Layer Chromatography-Flame Ionization Detection (TLC-FID) to Total Lipid Quantitation in Mycolic-Acid Synthesizing <i>Rhodococcus</i> and <i>Williamsia</i> Species |
title_sort |
application of thin-layer chromatography-flame ionization detection (tlc-fid) to total lipid quantitation in mycolic-acid synthesizing <i>rhodococcus</i> and <i>williamsia</i> species |
publisher |
MDPI AG |
series |
International Journal of Molecular Sciences |
issn |
1422-0067 |
publishDate |
2020-02-01 |
description |
In addition to cell membrane phospholipids, <i>Actinobacteria</i> in the order <i>Corynebacteriales</i> possess a waxy cell envelope containing mycolic acids (MA). In optimized culture condition, some species can also accumulate high concentrations of intracellular triacylglycerols (TAG), which are a potential source of biodiesel. Bacterial lipid classes and composition alter in response to environmental stresses, including nutrient availability, thus understanding carbon flow into different lipid classes is important when optimizing TAG synthesis. Quantitative and qualitative analysis of lipid classes normally requires combinations of different extraction, derivatization, chromatographic and detection methods. In this study, a single-step thin-layer chromatography-flame ionization detection (TLC-FID) technique was applied to quantify lipid classes in six sub-Antarctic <i>Corynebacteriales</i> strains identified as <i>Rhodococcus</i> and <i>Williamsia</i> species. A hexane:diethyl-ether:acetic acid solvent system separated the total cellular lipids extracted from cells lysed by bead beating, which released more bound and unbound MA than sonication. Typical profiles included a major broad non-polar lipid peak, TAG and phospholipids, although trehalose dimycolates, when present, co-eluted with phospholipids. Ultra-performance liquid chromatography-tandem mass-spectrometry and nuclear magnetic resonance spectroscopy detected MA signatures in the non-polar lipid peak and indicated that these lipids were likely bound, at least in part, to sugars from cell wall arabinogalactan. Waxy esters were not detected. The single-solvent TLC-FID procedure provides a useful platform for the quantitation and preliminary screening of cellular lipid classes when testing the impacts of growth conditions on TAG synthesis. |
topic |
tlc-fid iatroscan rods nmr lipid extraction mycolic acids <i>williamsia</i> <i>rhodococcus</i> |
url |
https://www.mdpi.com/1422-0067/21/5/1670 |
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