Phosphatidylcholines mass spectrometry

Cool the tubes and add 75 (jlI of 1M KOH in methanol, sonicate and incubate for 2 hr at 37°C. This step removes most of the interfering glycerolipids, in particular phosphatidylcholines, that can mask sphingomyelins in a simple Mass spectrometry scan. 

Petkovic, M., Muller, J., Muller, M., Schiller, J., Arnold, K., Arnold, J. Application of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for monitoring the digestion of phosphatidylcholine by pancreatic phospholipase A2. Anal. Biochem. 2002, 308, 61-70. 

Koizumi S, Yamamoto S, Hayasaka T, Konishi Y, Yamaguchi-Okada M, Goto-Inoue N, Sugiura Y, Setou M, Namba H (2010) Imaging mass spectrometry revealed the production of lyso-phosphatidylcholine in the injured ischemic rat brain. Neuroscience 168 219-225. doi 10.1016/j.neuroscience.2010.03.056... 

Zheng L, McQuaw C, Ewing A, Winograd N (2007) Sphingomyelin/phosphatidylcholine and cholesterol interactions studied by imaging mass spectrometry. J Am Chem Soc 129(51) 15730-15731. doi 10.1021/ja0741675... 

Phospholipase A2 Action. As in the case of phosphatidylcholine, the above-mentioned phospholipases will attack only the sn-3 form of naturally occurring (as well as synthetic) phosphatidylethanolamine. The products are, of course, lysophosphatidylethanolamine (1 -6>-acyl-2-lyso-.rn-glycero-3-phosphoethanolamine) and the fatty acids (liberated from the sn-2 position). The latter can be analyzed for composition and structure, as the methyl esters, by gas-liquid chromatography coupled with mass spectrometry. Usually these acyl groups are largely the unsaturated types. 

A phospholipid monolayer in the surface is consistent with the current model that LD are formed by TAG deposition between the two leaflets of the ER membrane and may remain connected to it [144, 145 see below]. Distribution of acyl-CoA cholesterol acyltransferase-1, a major enzyme that synthesizes cholesterylester, in the entire ER [148] seems to indicate that LD may bud anywhere along the membrane. However, Cap-LC/ESI mass spectrometry showed that FA moieties of phosphatidylcholine and lyso-phosphatidylchohne in LD are distinct from those in the rough ER [149]. The results do rule out the generation of the LD surface generated from the ER membrane but indicate that the former is a highly differentiated domain. Mature LD might be independent of the ER. Alternatively, the LD may be connected to the ER, but some molecular mechanism may demarcate the LD surface from the bulk ER membrane as postulated for other ER domains [150]. Whatever is true, TAG synthesized in wide areas of the ER do not deposit indiscriminately but are concentrated to loci specialized to make LD. ADRP or other LD-associated proteins may be involved (see below). 

Liebisch G, Lieser B, Rathenberg J, et al. (2004) High-throughput quantification of phosphatidylcholine and sphingomyelin by electrospray ionization tandem mass spectrometry coupled with isotope correction algorithm. Biochim Biophys Acta 1686 108-117... 

Enjalbal, C., Roggero, R., Cerdan, R., Martinez, J., Vial, H., and Aubagnac, J. L. (2004). Automated monitoring of phosphatidylcholine biosyntheses in Plasmodium falciparum by electrospray ionization mass spectrometry through stable isotope labeling experiments. Anal. Chem. 76,4515—4521. 

Al-Saad, K.A., Siems, W.F., Hill, H.H., Zabrouskov, V. and Knowles, N.R., Structural analysis of phosphatidylcholines by post-source decay matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry, J Am Soc Mass Spectrom, 14 (2003a) 373-382. 

Facino, R.M., Carini, M., Aldini, G., and Colombo, L., Characterization of the intermediate products of lipid peroxidation in phosphatidylcholine liposomes by fast-atom bombardment mass spectrometry and tandem mass spectrometry techniques, Rapid Commun. Mass Spec., 10, 1148, 1996. 

N. J. Jensen, K. B. Tomer, and M. L. Gross, Fast atom bombardment and tandem mass spectrometry of phosphatidylserine and phosphatidylcholine. Lipids 21, 580-588 (1986). 

O Donnell, V. B., Mass spectrometry analysis of oxidized phosphatidylcholine and nhosnhatidvlethanolamine. Biochim Biovhvs Acta 2011. 1811(11), 818-26. 

Hyphenated TLC techniques. TLC has been coupled with other instrumental techniques to aid in the detection, qualitative identification and, occasionally, quantitation of separated samples, and these include the coupling of TLC with high-pressure liquid chromatography (HPLC/TLC), with Fourier transform infra-red (TLC/FTIR), with mass spectrometry (TLC/ MS), with nuclear magnetic resonance (TLC/NMR) and with Raman spectroscopy (TLC/RS). These techniques have been extensively reviewed by Busch (1996) and by Somsen, Morden and Wilson (1995). The chemistry of oils and fats and their TLC separation has been so well established that they seldom necessitate the use of these coupling techniques for their identification, although these techniques have been used for phospholipid detection. Kushi and Handa (1985) have used TLC in combination with secondary ion mass spectrometry for the analysis of lipids. Fast atom bombardment (FAB) has been used to detect the molecular species of phosphatidylcholine on silica based on the molecular ion obtained by mass spectrometry (Busch et al, 1990). 

Gasser, H., Strohmaier, W., Schlag, G. et al. (1991) Characterization of phosphatidylcholines in rabbit lung lavage fluid by positive and negative ion fast-atom bombardment mass spectrometry. J. Chromatogr. Biomed. Applic., 562, 257-66. 

Ponchaut, S., Veitch, K., Libert, R. et al. (1996) Analysis by fast-atom bombardment tandem mass spectrometry of phosphatidylcholine isolated from heart mitochondrial fractions evidence of incorporation of monohydroxylated fatty acyl moieties. J. Am. Soc. Mass Spectrom., 1, 50-8. 

Zhang, J. Y., Nobes, B.J., Wang, J. and Blair, LA. (1994) Characterization of hydroxyeicosa-tetraenoic acids and hydroxyeicosatetraenoic acid phosphatidylcholines by liquid secondary ion tandem mass spectrometry. Biol Mass Spectrom., 23, 399 05. 

Figure 9.4 High-pressure liquid chromatography separation of 50 pg of a natural phosphatidylcholine mixture from egg yolk. The reconstructed ion chromatograms of diglyceride ions were selected from data acquired by full mass scanning from 120 amu to 820 amu. The relative intensity is shown based on the peak height. Column 3 pm Ultrasphere-ODS (4.6 mm x 7.5 cm). Mobile phase MeOH/hexane/0.1 m NH4OAC (71 5 7), 1 mlmin . Reprinted with permission from Kim, H. Y. and Salem, N. Jr, Phospholipid molecular species analysis by thermospray liquid chromatography/mass spectrometry. Anal. Chem., 58 (1), 9-14, 1986.

Figure 6.19. Coordination (Ag+) ion spray mass spectrometry (CIS-MS) of oxidized 1-palmitoyl -2-linoleoyl-5n-glycero-3-phosphatidylcholine and 1-stearoyl -2-arachidonyl-5n-glycero-3-phosphatidylcholine showing heterolytic cleavage of silver adduct of linoleoyl and arachidonyl hydroperoxides (CID = collision induced dissociation). Adapted from Milne and Porter (2001). See Figure 6.18.

Jackson, S.N.,Wang, H.Y., Woods, A.S. (2005) In situ structural characterization of phosphatidylcholines in brain tissue using MALDI-MS/MS. Journal of the American Society for Mass Spectrometry, 16, 2052-2056. 

Sugiura, Y., et al. (2009) Visualization of the cell-selective distribution of PUFA-containing phosphatidylcholines in mouse brain by imaging mass spectrometry. Journal of Lipid Research, 50,1776-1788. 

Schiller, J., Siiss, R., Petkovic, M., Hilbert, N., MiiUer, M., Zschornig, O., Arnhold, J., and Arnold, K. 2001. CsCl as an auxiliary reagent for the analysis of phosphatidylcholine mixtures by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDl-TOF MS), Chem. Phys. Lipids, 113 123-131. 

We describe the utility of intermediate-pressure MALDI and tandem mass spectrometry (MS/MS and MS ) for the characterization and imaging of phospholipids in brain tissue sections. The use of both MS/MS spectra and MS/MS images allows for identification of isobaric compounds. The structural characterization of phosphatidylcholines, phosphatidylserines, phosphatidylethanolamines, and sphingomyelins directly fi om tissue sections is described. 

Domingues P, Domingues MR, Amado FM, Ferrer-Correia AJ. Characterization of sodiated glycerol phosphatidylcholine phospholipids by mass spectrometry. Rapid Commun Mass Spectrom. 2001 15 799-804. 

Nakanishi, H., lida, Y., Shimizu, T. and Taguchi, R. (2009) Analysis of oxidized phosphatidylcholines as markers for oxidative stress, using multiple reaction monitoring with theoretically expanded data sets with reversed-phase liquid chromatography/tandem mass spectrometry. J. Chromatogr. B 877, 1366-1374. 

Lehmann, W.D., Koester, M., Erben, G. and Keppler, D. (1997) Characterization and quantification of rat bile phosphatidylcholine by electrospray-tandem mass spectrometry. Anal. Biochem. 246, 102-110. 

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