65
other seaweed. They illustrated that a range of oxylipins (HETEs, di-HETEs, prostaglandins, and leukotrienes) can be extracted from seaweeds using methanol followed by enrichment with Waters OASIS HLB SPE cartridge and separation by
LC-MS using mobile phase A, 0.1% formic acid (w/v) in 2% acetonitrile in ultrapure water (v/v), and mobile phase B, 0.1% formic acid (w/v) in acetonitrile on a
BEH C18 UPLC column (2.1 mm x 1.7 μm; Waters or a similar C18 column).
Different chromatographic methods coupled with MS have invariably been used
for lipid analysis, but it is the development of soft ionization techniques (electrospray ionization mass spectrometry, ESI-MS; atmospheric pressure chemical ionization, APCI; and matrix-assisted laser desorption ionization, MALDI) that has
greatly accelerated the field of lipidomics. In APCI, the sample is nebulized and
heated so that both solvent and analytes are in the gas phase followed by a corona
discharge which ionizes the solvent molecules that subsequently also ionize the analyte molecules. APCI generally yields monocharged ions and is mainly used with
small thermally stable nonpolar molecules (<1500 Da) and is mostly applied for
glycerolipids, fatty acyls and sterols, and fatty acid esters (Li et al. 2014; Vaz et al.
2015). For MALDI-MS-based lipidomic analysis, the sample is mixed with a matrix
that readily forms crystals that aid the ionization process. The fluid mixture of sample and matrix is spotted on a MALDI plate and allowed to dry. To ionize the analyte
molecules, a laser is fired at the matrix crystals in the dried-droplet spot, which
absorbs the laser energy resulting in desorption and ionization. The ionized matrix
molecule then transfers its charge to the analyte, thus ionizing the analyte (Vaz et al.
2015; Pati et al. 2016). ESI-MS, particularly, has emerged as a powerful and indispensable tool for lipidomics, as it allows analysis of thermally labile and nonvolatile
compounds and thus can be applied to almost all lipid categories, discussed hereby
in detail. (Vaz et al. 2015). In ESI-MS, the lipid sample is nebulized through a highly
charged capillary using heated nitrogen gas, producing a fine aerosol that results in
evaporation of the solvent and ionization of the molecules after which the ions enter
the mass spectrometer. ESI-MS analysis of lipids results in decreased molecular ion
decomposition and better reproducibility and has lower detection limits (Gross and
Han 2011; Pati et al. 2016; Vaz et al. 2015; Wang et al. 2016). ESI-MS-based lipidomics is classified into two categories depending on whether the lipidomic analysis is
being accomplished at a constant or varied lipid concentration:
A. LC-MS-based lipidomics: It is also known as CLASS approach (comprehensive
lipidomic analysis by class separation), where the concentration of lipid solution
is constantly changing. The lipid molecules are separated by HPLC/LC and analyzed by online MS.
B. Shotgun lipidomics: It is also known as direct infusion in which the concentration of lipid solution delivered to the ion source is constant. In this, the lipid
sample is infused directly in the mass spectrometer without prior separation, and
lipid species are identified and quantified by specific precursor ion scans (PIS)
or neutral loss scans (NLS) reviewed recently by Wang et al. (2016). The lipid
solution is delivered by a syringe pump (generally a tightly sealed high quality
glass syringe) for direct infusion. Its major disadvantage is clogging of capillary
4 Seaweed Lipidomics in the Era of ‘Omics’ Biology: A Contemporary Perspective
other seaweed. They illustrated that a range of oxylipins (HETEs, di-HETEs, prostaglandins, and leukotrienes) can be extracted from seaweeds using methanol followed by enrichment with Waters OASIS HLB SPE cartridge and separation by
LC-MS using mobile phase A, 0.1% formic acid (w/v) in 2% acetonitrile in ultrapure water (v/v), and mobile phase B, 0.1% formic acid (w/v) in acetonitrile on a
BEH C18 UPLC column (2.1 mm x 1.7 μm; Waters or a similar C18 column).
Different chromatographic methods coupled with MS have invariably been used
for lipid analysis, but it is the development of soft ionization techniques (electrospray ionization mass spectrometry, ESI-MS; atmospheric pressure chemical ionization, APCI; and matrix-assisted laser desorption ionization, MALDI) that has
greatly accelerated the field of lipidomics. In APCI, the sample is nebulized and
heated so that both solvent and analytes are in the gas phase followed by a corona
discharge which ionizes the solvent molecules that subsequently also ionize the analyte molecules. APCI generally yields monocharged ions and is mainly used with
small thermally stable nonpolar molecules (<1500 Da) and is mostly applied for
glycerolipids, fatty acyls and sterols, and fatty acid esters (Li et al. 2014; Vaz et al.
2015). For MALDI-MS-based lipidomic analysis, the sample is mixed with a matrix
that readily forms crystals that aid the ionization process. The fluid mixture of sample and matrix is spotted on a MALDI plate and allowed to dry. To ionize the analyte
molecules, a laser is fired at the matrix crystals in the dried-droplet spot, which
absorbs the laser energy resulting in desorption and ionization. The ionized matrix
molecule then transfers its charge to the analyte, thus ionizing the analyte (Vaz et al.
2015; Pati et al. 2016). ESI-MS, particularly, has emerged as a powerful and indispensable tool for lipidomics, as it allows analysis of thermally labile and nonvolatile
compounds and thus can be applied to almost all lipid categories, discussed hereby
in detail. (Vaz et al. 2015). In ESI-MS, the lipid sample is nebulized through a highly
charged capillary using heated nitrogen gas, producing a fine aerosol that results in
evaporation of the solvent and ionization of the molecules after which the ions enter
the mass spectrometer. ESI-MS analysis of lipids results in decreased molecular ion
decomposition and better reproducibility and has lower detection limits (Gross and
Han 2011; Pati et al. 2016; Vaz et al. 2015; Wang et al. 2016). ESI-MS-based lipidomics is classified into two categories depending on whether the lipidomic analysis is
being accomplished at a constant or varied lipid concentration:
A. LC-MS-based lipidomics: It is also known as CLASS approach (comprehensive
lipidomic analysis by class separation), where the concentration of lipid solution
is constantly changing. The lipid molecules are separated by HPLC/LC and analyzed by online MS.
B. Shotgun lipidomics: It is also known as direct infusion in which the concentration of lipid solution delivered to the ion source is constant. In this, the lipid
sample is infused directly in the mass spectrometer without prior separation, and
lipid species are identified and quantified by specific precursor ion scans (PIS)
or neutral loss scans (NLS) reviewed recently by Wang et al. (2016). The lipid
solution is delivered by a syringe pump (generally a tightly sealed high quality
glass syringe) for direct infusion. Its major disadvantage is clogging of capillary
4 Seaweed Lipidomics in the Era of ‘Omics’ Biology: A Contemporary Perspective