Quantification of distinct molecular species of the 2-lyso metabolite of platelet-activating factor by gas chromatography—negative-ion chemical ionization mass spectrometry

Turk, John; Bohrer, Alan; Stump, W. Thomas; Ramanadham, Sasanka; Mangino, Martin J.

doi:10.1016/0378-4347(92)80145-g

Cited by 5 publications

(7 citation statements)

References 27 publications

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“…After centrifugation (800g, 5 min), the ether phase containing the diglyceride product was aspirated into a separate vial and concentrated to dryness. The diglyceride was then saponified with LiOH in dimethoxyethane as described elsewhere (Turk et al, 1992b). The product was then converted to the bispentafluorobenzoate derivative and analyzed by GC-NICI-MS as described above.…”

Section: Methodsmentioning

confidence: 99%

“…Determination of Acid Lability of Phospholipids. Two aliquots of [3H] arachidonate-containing islet phospholipid samples were prepared, spiked with 10 pg of (C16:0a/C18: 2)-PC as internal standard, concentrated, treated with acid or with vehicle, and analyzed by RP-HPLC as described elsehwere (DaTorre & Creer, 1991;Ramanadham et al, 1992b). Plasmalogens are destroyed by acid treatment and appear only in the chromatogram of the vehicle-treated sample, but diacyl-and alkylacylphospholipids are stable under these conditions and appear in the chromatograms of both acidtreated and vehicle-treated samples.…”

Section: Methodsmentioning

confidence: 99%

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Mass spectrometric characterization of arachidonate-containing plasmalogens in human pancreatic islets and in rat islet .beta.-cells and subcellular membranes

Ramanadham

Bohrer

Gross³

et al. 1993

Biochemistry

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Pancreatic islets, when stimulated with D-glucose, secrete insulin by processes requiring glycolytic metabolism and generation of ATP. Hydrolysis of membrane phospholipids also occurs in glucose-stimulated islets, resulting in accumulation of nonesterified arachidonate, which facilitates Ca2+ entry and the rise in beta-cell [Ca2+] that triggers insulin secretion. Glucose-induced hydrolysis of arachidonate from islet phospholipids is mediated in part by an ATP-stimulated, Ca(2+)-independent (ASCI) phospholipase A2 (PLA2) which prefers plasmenyl over diacyl phospholipid substrates. Here we characterize the endogenous plasmalogen content of islet cells and subcellular membranes. Fast atom bombardment mass spectrometric analyses demonstrated that three of the most abundant molecular species of ethanolamine phospholipids in rat pancreatic islets were plasmalogens with sn-2 arachidonate residues and palmitic, oleic, or stearic aldehyde residues, respectively, in the sn-1 position. Purified populations of beta-cells prepared by fluorescence-activated cell sorting were also found to contain these plasmenylethanolamine molecular species in abundance similar to that in intact islets and greater than that in islet alpha-cells. Both islet plasma membranes (PM) and endoplasmic reticulum (ER) also contained these plasmenylethanolamine species, which accounted for 42% (PM) to 64% (ER) of the ethanolamine phospholipid arachidonate content of these membranes, as measured by stable isotope dilution mass spectrometry. Plasmenylethanolamine species were also abundant constituents of human pancreatic islets (accounting for 58% of their ethanolamine phospholipid arachidonate content) and were hydrolyzed more rapidly than diacyl ethanolamine phospholipid by human islet cytosolic ASCI-PLA2. Both secretagogue-induced eicosanoid release and insulin secretion from human islets were attenuated by an ASCI-PLA2 suicide substrate which sterically resembles plasmalogens. These observations are consistent with the hypotheses that islet beta-cell ASCI-PLA2-catalyzed hydrolysis of arachidonate from endogenous plasmenylethanolamine substrates may occur in membrane compartments which participate in regulation of the beta-cell cytosolic [Ca2+] and that this may be an intermediary biochemical event in the induction of insulin secretion.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Mass spectrometric characterization of arachidonate-containing plasmalogens in human pancreatic islets and in rat islet .beta.-cells and subcellular membranes

Ramanadham

Bohrer

Gross³

et al. 1993

Biochemistry

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“…The samples were extracted for total lipids by the method of Bligh and Dyer. For quantitation of any lyso-PAF in the samples, the lipid extract was treated with propionylchloride to convert lyso-PAF to propinoyl PAF (11). The extract was concentrated to dryness under N 2 and derivatized with 100 ~tL propionic acid and 50 ~tL propionyl chloride for 45 min at 60~ This material was dried (N2) and reconstituted in 1 mL chloroform.…”

Section: Methodsmentioning

confidence: 99%

“…Gas chromatography (GC) was performed on a Hewlett Packard 5890 gas chromatograph with a fused silica capillary column 8m Hewlett Packard Ultraperformance (methysilicone, 0.17 micron film thickness, 0.32 mm i.d.) interfaced with a Hewlett Packard model 5988 quadrapole mass spectrometer (11). Injections were made in the splitless mode with helium as the carrier gas (total flow 10 mL/min, head pressure 4 psi).…”

Section: Methodsmentioning

confidence: 99%

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Quantification of distinct molecular species of platelet activating factor in ulcerative colitis

Thyssen¹,

Turk²,

Bohrer³

et al. 1996

Lipids

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The aim of this study was to characterize the synthesis and metabolism of platelet activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by colonic mucosa from patients with ulcerative colitis and healthy individuals. Tissue was obtained by endoscopic biopsy and by scraping the mucosa from surgical resections. Tissue was assayed for the various molecular species of PAF and its biologically inactive metabolite lyso-PAF using gas chromatography/mass spectrometry. Mucosa from surgical resections for ulcerative colitis contained C16:0 PAF (mean = 156 ng/g of mucosa), but not C18:0 PAF, PAF was not identified in mucosa from normal surgical resections or in endoscopic biopsies from either patients with ulcerative colitis or normal individuals. Both C16:0 lyso-PAF and C18:0 lyso-PAF were found in mucosa from normal and ulcerative colitis surgical resections and in endoscopic biopsies from ulcerative colitis and normal tissue. Levels of lyso-PAF were similar in ulcerative colitis and normal mucosa. Incubation of mucosa from areas of active inflammation in ulcerative colitis with the calcium ionophore A23187 increased the levels of C16:0 PAF by 2-3 fold and also increased the levels of C16:0 lyso-PAF. Addition of 3H-PAF to endoscopic biopsies from either normal individuals or patients with ulcerative colitis resulted in hydrolysis to 3H-lyso-PAF. The data on colonic mucosal levels of PAF are consistent with the results of earlier studies measuring PAF in patients with ulcerative colitis by bioassay. This study examines the synthesis and metabolism of specific molecular species of PAF in ulcerative colitis for the first time.

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Identification and Quantitation of Changes in the Platelet Activating Factor Family of Glycerophospholipids over the Course of Neuronal Differentiation by High-Performance Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry

et al. 2007

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Glycerophospholipids are important structural lipids in membranes with changes associated with progressive neurodegenerative disorders such as Alzheimer disease. Synthesis of the platelet activating factor (PAF) glycerophospholipid subclass is implicated in the control of neuronal differentiation and death. In this article, we combine nanoflow HPLC and mass spectrometry to screen, identify, and quantitate changes in glycerophospholipid subspecies, specifically PAF family members, over the course of neuronal differentiation. Furthermore, precursor ion scans for fragments characteristic of PAF phosphocholine family members and the standard additions of PAF subspecies were combined to perform absolute quantitation of PAF lipids in undifferentiated and differentiated PC12 cells. Surprisingly, a marked asymmetry was detected in the two predominant PAF species (C16:0, C18:0) over the course of differentiation. These results describe a new technique for the sensitive analysis of lipids combining nanoflow HPLC, ESI-MS, and precursor ion scan. Limits of detection of as little as 2 pg of PAF and LPC were obtained, and analysis of the lipidome of as little as 70,000 cells was performed on this system. Furthermore, application to the PC12 model identified a quantifiable difference between PAF molecular species produced over the course of neuronal differentiation.

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Quantification of distinct molecular species of the 2-lyso metabolite of platelet-activating factor by gas chromatography—negative-ion chemical ionization mass spectrometry

Cited by 5 publications

References 27 publications

Mass spectrometric characterization of arachidonate-containing plasmalogens in human pancreatic islets and in rat islet .beta.-cells and subcellular membranes

Mass spectrometric characterization of arachidonate-containing plasmalogens in human pancreatic islets and in rat islet .beta.-cells and subcellular membranes

Quantification of distinct molecular species of platelet activating factor in ulcerative colitis

Identification and Quantitation of Changes in the Platelet Activating Factor Family of Glycerophospholipids over the Course of Neuronal Differentiation by High-Performance Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry

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