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Shotgun lipidomics identifies a paired rule for the presence of isomeric ether phospholipid molecular species.

Yang K, Zhao Z, Gross RW, Han X - PLoS ONE (2007)

Bottom Line: The biochemical basis of this rule results from the fact that the enzymes which participate in either the sequential oxidation of aliphatic alcohols to fatty acids, or the reduction of long chain fatty acids to aliphatic alcohols (metabolic precursors of ether lipid synthesis), are not entirely selective with respect to acyl chain length or degree of unsaturation.Application of this rule to mass spectrometric analyses provides predictive clues to the presence of specific molecular species and greatly expands the number of identifiable and quantifiable ether lipid species present in biological samples.Through appropriate alterations in the database, use of the paired rule increases the number of identifiable metabolites in metabolic networks, thereby facilitating identification of biomarkers presaging disease states.

View Article: PubMed Central - PubMed

Affiliation: Division of Bioorganic Chemistry and Molecular Pharmacology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri, United States of America.

ABSTRACT

Background: Ether phospholipids are abundant membrane constituents present in electrically active tissues (e.g., heart and the brain) that play important roles in cellular function. Alterations of ether phospholipid molecular species contents are associated with a number of genetic disorders and human diseases.

Methodology/principal findings: Herein, the power of shotgun lipidomics, in combination with high mass accuracy/high resolution mass spectrometry, was explored to identify a paired rule for the presence of isomeric ether phospholipid molecular species in cellular lipidomes. The rule predicts that if an ether phospholipid A'-B is present in a lipidome, its isomeric counterpart B'-A is also present (where the ' represents an ether linkage). The biochemical basis of this rule results from the fact that the enzymes which participate in either the sequential oxidation of aliphatic alcohols to fatty acids, or the reduction of long chain fatty acids to aliphatic alcohols (metabolic precursors of ether lipid synthesis), are not entirely selective with respect to acyl chain length or degree of unsaturation. Moreover, the enzymatic selectivity for the incorporation of different aliphatic chains into the obligatory precursor of ether lipids (i.e., 1-O-alkyl-glycero-3-phosphate) is also limited.

Conclusions/significance: This intrinsic amplification of the number of lipid molecular species present in biological membranes predicted by this rule and demonstrated in this study greatly expands the number of ether lipid molecular species present in cellular lipidomes. Application of this rule to mass spectrometric analyses provides predictive clues to the presence of specific molecular species and greatly expands the number of identifiable and quantifiable ether lipid species present in biological samples. Through appropriate alterations in the database, use of the paired rule increases the number of identifiable metabolites in metabolic networks, thereby facilitating identification of biomarkers presaging disease states.

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Product ion analyses of synthetic 18∶0-20∶4 plasmenylethanolamine molecular species in the negative-ion mode.Product ion ESI/MS analysis of deprotonated 18∶0-20∶4 plasmenylethanolamine at m/z 750.54 was performed on an LTQ-Orbitrap mass spectrometer with a C-trap using an ion selective window of 1 Th by LTQ. Collision activation in C-trap was carried out with normalized collision energy of 55% and gas pressure of 1 mT. The resultant fragment ions were analyzed in the Orbitrap. The arrow indicates the absence of the 18:0 FA carboxylate in the spectrum after amplifying the position greater than1,000 fold.
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pone-0001368-g004: Product ion analyses of synthetic 18∶0-20∶4 plasmenylethanolamine molecular species in the negative-ion mode.Product ion ESI/MS analysis of deprotonated 18∶0-20∶4 plasmenylethanolamine at m/z 750.54 was performed on an LTQ-Orbitrap mass spectrometer with a C-trap using an ion selective window of 1 Th by LTQ. Collision activation in C-trap was carried out with normalized collision energy of 55% and gas pressure of 1 mT. The resultant fragment ions were analyzed in the Orbitrap. The arrow indicates the absence of the 18:0 FA carboxylate in the spectrum after amplifying the position greater than1,000 fold.

Mentions: To exclude the possibility that one of the paired pPtdEtn molecular species which generates FA carboxylate (usually in low or very low abundance after CID) was due to the presence of an unknown fragmentation pathway of pPtdEtn molecular ions, additional experiments on the analyses of synthetic pPtdEtn molecular species were conducted. For example, tandem MS analysis of 18∶0-20∶4 pPtdEtn after CID demonstrates an abundant fragment ion at m/z 303.23 (i.e., 20∶4 FA carboxylate), a modest fragment ion at m/z 259.24 (corresponding to the aliphatic anion resulting from the loss of a CO2 molecule from arachidonate as mentioned above), two fragment ions at m/z 446.30 and 464.31 (i.e., lyso-pPtdEtn derivatives), and a very low abundance ion at m/z 267.27 (corresponding to octadecan-1′-enolate which indicates that the pPtdEtn molecular species containing 18∶0 vinyl ether is the most abundant one in all isomeric species) (Figure 4). No detectable fragment ion at m/z 283.26 (i.e., 18∶0 FA carboxylate) was present in the mass spectrum (Figure 4 inset). Identical results were also obtained from the analyses of other synthetic plasmalogen molecular species (data not shown). Accordingly, the very low abundance FA carboxylate fragments present in the tandem MS analyses of pPtdEtn in bovine heart PtdEtn did not result from an unknown CID pathway, further supporting the presence of the paired pPtdEtn molecular species in biological lipid extracts.


Shotgun lipidomics identifies a paired rule for the presence of isomeric ether phospholipid molecular species.

Yang K, Zhao Z, Gross RW, Han X - PLoS ONE (2007)

Product ion analyses of synthetic 18∶0-20∶4 plasmenylethanolamine molecular species in the negative-ion mode.Product ion ESI/MS analysis of deprotonated 18∶0-20∶4 plasmenylethanolamine at m/z 750.54 was performed on an LTQ-Orbitrap mass spectrometer with a C-trap using an ion selective window of 1 Th by LTQ. Collision activation in C-trap was carried out with normalized collision energy of 55% and gas pressure of 1 mT. The resultant fragment ions were analyzed in the Orbitrap. The arrow indicates the absence of the 18:0 FA carboxylate in the spectrum after amplifying the position greater than1,000 fold.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2147047&req=5

pone-0001368-g004: Product ion analyses of synthetic 18∶0-20∶4 plasmenylethanolamine molecular species in the negative-ion mode.Product ion ESI/MS analysis of deprotonated 18∶0-20∶4 plasmenylethanolamine at m/z 750.54 was performed on an LTQ-Orbitrap mass spectrometer with a C-trap using an ion selective window of 1 Th by LTQ. Collision activation in C-trap was carried out with normalized collision energy of 55% and gas pressure of 1 mT. The resultant fragment ions were analyzed in the Orbitrap. The arrow indicates the absence of the 18:0 FA carboxylate in the spectrum after amplifying the position greater than1,000 fold.
Mentions: To exclude the possibility that one of the paired pPtdEtn molecular species which generates FA carboxylate (usually in low or very low abundance after CID) was due to the presence of an unknown fragmentation pathway of pPtdEtn molecular ions, additional experiments on the analyses of synthetic pPtdEtn molecular species were conducted. For example, tandem MS analysis of 18∶0-20∶4 pPtdEtn after CID demonstrates an abundant fragment ion at m/z 303.23 (i.e., 20∶4 FA carboxylate), a modest fragment ion at m/z 259.24 (corresponding to the aliphatic anion resulting from the loss of a CO2 molecule from arachidonate as mentioned above), two fragment ions at m/z 446.30 and 464.31 (i.e., lyso-pPtdEtn derivatives), and a very low abundance ion at m/z 267.27 (corresponding to octadecan-1′-enolate which indicates that the pPtdEtn molecular species containing 18∶0 vinyl ether is the most abundant one in all isomeric species) (Figure 4). No detectable fragment ion at m/z 283.26 (i.e., 18∶0 FA carboxylate) was present in the mass spectrum (Figure 4 inset). Identical results were also obtained from the analyses of other synthetic plasmalogen molecular species (data not shown). Accordingly, the very low abundance FA carboxylate fragments present in the tandem MS analyses of pPtdEtn in bovine heart PtdEtn did not result from an unknown CID pathway, further supporting the presence of the paired pPtdEtn molecular species in biological lipid extracts.

Bottom Line: The biochemical basis of this rule results from the fact that the enzymes which participate in either the sequential oxidation of aliphatic alcohols to fatty acids, or the reduction of long chain fatty acids to aliphatic alcohols (metabolic precursors of ether lipid synthesis), are not entirely selective with respect to acyl chain length or degree of unsaturation.Application of this rule to mass spectrometric analyses provides predictive clues to the presence of specific molecular species and greatly expands the number of identifiable and quantifiable ether lipid species present in biological samples.Through appropriate alterations in the database, use of the paired rule increases the number of identifiable metabolites in metabolic networks, thereby facilitating identification of biomarkers presaging disease states.

View Article: PubMed Central - PubMed

Affiliation: Division of Bioorganic Chemistry and Molecular Pharmacology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri, United States of America.

ABSTRACT

Background: Ether phospholipids are abundant membrane constituents present in electrically active tissues (e.g., heart and the brain) that play important roles in cellular function. Alterations of ether phospholipid molecular species contents are associated with a number of genetic disorders and human diseases.

Methodology/principal findings: Herein, the power of shotgun lipidomics, in combination with high mass accuracy/high resolution mass spectrometry, was explored to identify a paired rule for the presence of isomeric ether phospholipid molecular species in cellular lipidomes. The rule predicts that if an ether phospholipid A'-B is present in a lipidome, its isomeric counterpart B'-A is also present (where the ' represents an ether linkage). The biochemical basis of this rule results from the fact that the enzymes which participate in either the sequential oxidation of aliphatic alcohols to fatty acids, or the reduction of long chain fatty acids to aliphatic alcohols (metabolic precursors of ether lipid synthesis), are not entirely selective with respect to acyl chain length or degree of unsaturation. Moreover, the enzymatic selectivity for the incorporation of different aliphatic chains into the obligatory precursor of ether lipids (i.e., 1-O-alkyl-glycero-3-phosphate) is also limited.

Conclusions/significance: This intrinsic amplification of the number of lipid molecular species present in biological membranes predicted by this rule and demonstrated in this study greatly expands the number of ether lipid molecular species present in cellular lipidomes. Application of this rule to mass spectrometric analyses provides predictive clues to the presence of specific molecular species and greatly expands the number of identifiable and quantifiable ether lipid species present in biological samples. Through appropriate alterations in the database, use of the paired rule increases the number of identifiable metabolites in metabolic networks, thereby facilitating identification of biomarkers presaging disease states.

Show MeSH
Related in: MedlinePlus