Electrospray lipids

Schneiter, R. Brugger, B. Sandhoff, R. Zellnig, G. Leber, A. Lampl, M. Athenstaedt, K. Hrastnik, C. Eder, S. Daum, G. Paltauf, F. Wieland, F. T. Kohlwein, S. D. Electrospray ionization tandem mass spectrometry (ESI-MS/MS) analysis of the lipid molecular species composition of yeast subcellular membranes reveals acyl chain-based sorting/remodeling of distinct molecular species en route to the plasma membrane. J. Cell Biol. 1999,146,741-754. 

In the present chapter, we first provide some general information concerning the chemistry of waxes and lipids currently encountered in various items from our cultural heritage and we detail the main protocols based on direct mass spectrometry that have been developed so far. We then discuss the mass spectra obtained by EI-MS on a range of reference substances and materials sampled from museum and archaeological artefacts. We then focus on the recent possibilities supplied by electrospray ionisation for the elucidation of the structure of biomarkers of beeswax and animal fats. 

Figure 4.1 Analytical strategy in which direct mass spectrometry analyses using either elec tron ionisation or electrospray are used for detecting and identifying lipid substances in archaeological and museum samples...

Although El MS is an efficient way to provide structural information on several molecular constituents of various lipid substances it only provides partial information and it is particularly not suitable for the study of the low volatile components. High molecular weight and nonvolatile compounds are particularly difficult to analyse in this way and it may therefore be interesting to explore the possibilities of other ionisation modes such as electrospray for an accurate structural study of high molecular constituents such as monoester and diester species of beeswax (Gamier et al., 2002) and TAGs of animal fats... 

Recently, a quantitative electrospray ionization/mass spectrometry method (ESI/MS) has been developed to analyze the molecular profile, or hpidome of different lipid classes in very small samples. In this method, total lipid extracts from tissues or cultured cells can be directly analyzed. By manipulating the ionization method, the mass spectrographs of polar or even non-polar lipids can be obtained [8]. This method and the use of lipid arrays allow precise and quantitative identification of the lipid profile of a given tissue, and map functional changes that occur. 

D1 (10,17S-docosatriene) from DHA using tandem liquid chromatography-photodiode array-electrospray ionization-tandem mass spectrometry (LC-PDA-ESI-MS-MS)-based lipidomic analysis have been documented in ischemic brain [4] and retinal pigment epithelium [5], This new lipid is called neuroprotectin D1 (1) because of its neuro-protectiveproperties in brain ischemia-reperfusion [4] and in oxidative stress-challenged retinal pigment epithelial cells [5] (2) because of its potent ability to inactivate proapoptotic signaling (see apoptosis, Ch. 35) [5] and (3) because it is the first identified neuroprotective mediator derived from DHA. 

Several useful publications describing early direct injection API-electrospray methods appHcable to neurochemical studies include studies on acetylchoHnesterase phosphonylation products (Barak et al., 1997) glycerophosphocholine lipids (Flarrison and Murphy, 1996) receptor antibody characterization (Lewis et al., 1994) neuropeptide Yanalogs (Beck-Sickinger et al., 1994) and large biomolecules (Loo et al.,... 

Atmospheric Pressure Chemical Ionization (APCI)/MS APCI/MS is used to analyze compounds of intermediate molecular weight (100-1,500 da) and intermediate polarity and is particularly useful for the analysis of biochemicals such as triacylglycerides, carotenoids, and lipids (Byrdwell, 2001). For volatile, nonpolar compounds of low molecular weight, GC/MS is preferred to APCl/MS whereas APl-electrospray/ MS provides better results for larger, more polar materials. The selection of APCl/MS over GC/MS or APl-electrospray/MS depends on the compounds to be analyzed. Many LC/MS instruments can be easily switched between APCl/MS and APl-electrospray/MS so that it can be rapidly determined which ionization process is more suitable to a given chemical. Additional manipulations such as pre and postcolumn derivatization reactions (Nagy et al., 2004 Peters et al., 2004) or coulometric oxidation (Diehl et al., 2001) can make the chemicals of interest more amenable to detection by APCI. 

Lieser B, Liebisch G, Drobnik W, Schmitz G. 2003. Quantification of sphingosine and shinganine from crude Hpid extracts by HPLC electrospray ionization tandem mass spectrometry. J Lipid Res 44 2209. 

A newer approach for lipid analysis is electrospray ionization tandem mass spectrometry (ESI-MS/MS) (Welti et al., 2002). This method requires limited sample preparation and sample size to identify and quantify lipids. Fauconnier et al. (2003) used ESI-MS/MS to analyze phospholipid and galactolipid levels during aging of potato tubers. 

SCHEME 21. Fragmentation pattern of the ozonide of an unsaturated glycerophosphocholine lipid in electrospray MS... 

Perwaiz S, Tuchweber B, Mignault D, Gilat T, Yousef IM (2001) Determination ofbile acids in biological fluids by liquid chromatography-electrospray tandem mass spectrometry. J Lipid Res 42 114-119... 

M. Gu, J. L. Kerwin, J. D. Watts, and R. Aebersold, Ceramide profiling of complex lipid mixtures by electrospray ionization mass spectrometry, Anal. Biochem., 244 (1997) 347-356. 

Advanced analytical techniques, particularly mass spectrometry (MS), often combined with liquid chromatography (LC) or gas chromatography (GC), are requisite for lipid analysis and they have played the crucial role in the emergence as well as the progresses of lipidomics. MS is the principal choice for the lipid analysis, particularly using electrospray ionization (ESI) and sometimes also atmospheric pressure chemical ionization or laser-based MS methods for surface analysis. The MS-based techniques are the best choice for lipidomics due to their superior sensitivity and molecular specificity, and because they provide the ability to resolve the extensive compositional and structural diversity of lipids in biological systems. 

In recent years the rapid development of high-sensitivity analytical techniques such as mass spectrometry (MS) and liquid chromatography (LC) supported the investigation of the endocannabinoids as part of a complex lipid network. The identification of lipid components of the endocannabinoid system can be achieved in a single analytical step by state-of-the-art platforms such as tandem mass spectrometry (MS/MS), which provides the detailed structural information necessary for characterization of lipids and increases specificity in complex biological matrices. Furthermore, the implementation of ionization techniques such as electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) allow the coupling of LC to MS, and permits the separation and analysis of endocannabinoids with greater speed and accuracy. 

Dill A, Ifa D, Manicke N, Zheng O, Cooks R (2009) Mass spectrometric imaging of lipids using desorption electrospray ionization. J Chromatogr B 877 2883-2889. doi 10.1016/j. jchromb.2008.12.058... 

Ellis S, Wu C, Deeley J, ZhuX, Truscott R, Panhuis J, Cooks R, Mitchell T, Blanksby S (2010) Imaging of human lens lipids by desorption electrospray ionization mass spectrometry. J Am Soc Mass Spectrom 21 2095-2104. doi 10.1016/j.jasms.2010.09.003... 

Girod M, Shi Y, Cheng J, Cooks G (2010) Desorption electrospray ionization imaging mass spectrometry of lipids in rat spinal cord. J Am Soc Mass Spectrom 21 1177-1189. doi 10.1016/j. jasms.2010.03.028... 

Nemes P, Woods A, Vertes A (2010) Simultaneous imaging of small metabolites and lipids in rat brain tissue at atmospheric pressure by laser ablation electrospray ionization mass spectrometry. Anal Chem 82 982-988. doi 10.1021/ac902245p... 

In an interesting experimental protocol, Silvestro et al. (1993) utilized HPLC-mass spectrometry with an ion spray (electrospray) interface for determination of PAF and lysoPAF in human PMN (neutrophils). Both unstimulated and stimulated (with complement-activated zymosan) cells were used as starting material. The total lipids were isolated in the usual way, and the PAF was isolated and purified by a combination of thin-layer chromatography, HPLC, and silica chromatography. This final PAF preparation was subjected to a bioassay with the inclusion of 3H 16 0 PAF to monitor recoveries. 

Kalo, P., Kemppinen, A., Ollilainen, V., Kuksis, A. 2004. Regiospecific determination of short-chain triacylglycerols in butterfat by normal-phase HPLC with on-line electrospray-tandem mass spectrometry. Lipids, 39, 915-928. 

Kerwin, J.L., Tuininga, A.R., Ericsson, L.H. 1994. Identification of molecular species of glycerophospholipids and sphingomyelin using electrospray mass spectrometry. J. Lipid Res., 35, 1102-1114. 

Electrospray mass spectrometry has developed into a well-established method of wide scope and potential over the past 15 years. The softness of electrospray ionization has made this technique an indispensable tool for biochemical and biomedical research. Electrospray ionization has revolutionized the analysis of labile biopolymers, with applications ranging from the analysis of DNA, RNA, oligonucleotides, proteins as well as glycoproteins to carbohydrates, lipids, gly-colipids, and lipopolysaccharides, often in combination with state-of-the-art separation techniques like liquid chromatography or capillary electrophoresis [1,2]. Beyond mere analytical applications, electrospray ionization mass spectrometry (ESMS) has proven to be a powerful tool for collision-induced dissociation (CID) and multiple-stage mass spectrometric (MSn) analysis, and - beyond the elucidation of primary structures - even for the study of noncovalent macromolecular complexes [3]. 

One limitation of LC-MS/MS methodology is that compounds in the sample extract, typically lipids, can affect the initial ionization of the analyte. This is referred to as matrix effects and can either enhance or suppress the electrospray ionization, leading to considerable inaccuracies in both qualitative and quantitative analyses [96, 97]. As such, the cleanup methodology employed, especially when working with biological samples, is of the utmost importance. In addition, the use of mass-labelled internal standards and matrix-matched standards, can minimize matrix effects [93, 94]. 

Wenk, M.R., Lucast, L., Di Paolo, G., Romanelli, A.J., Suchy, S.F., Nussbaum, R.L., Cline, G.W., Shulman, G.I., McMurray, W., and De Camilli, P., 2003, Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry. Nat. Biotechnol. 21 813-817. 

Analytical methods for tocol analysis have continued to improve, as noted by Abidi (2000), and in the intervening ten years, as noted in this chapter. We predict that advances will continue to be made in the field of the chromatographic analysis of tocols. Also, we believe that lipidomic methods (quantitative analysis via direct injection tandem electrospray ionization mass spectrometry) will be developed for the rapid analysis of tocols, just as these methods have already been used for the profiling of phospholipids and glycolipids (Han, 2011 Welti, 2011). These methods usually involve the direct injection of lipid samples into a... 

The dynamic development of mass spectrometry has had a huge impact on lipid analysis. Currently, a variety of suitable mass spectrometers is available. In principal, a mass spectrometer consists of an ion source, a mass analyzer, and an ion detector. The typical features of each instrument (Fig. 2) result mostly from the types of ion source and mass analyzer. To date, the ionization techniques apphed to lipid analysis include Electrospray Ionization (ESI or nano-ESI), Atmospheric Pressure Chemical Ionization (APCI), Matrix-Assisted Laser Desorption/Ionization... 

DESI is performed by directing electrosprayed-charged droplets onto a surface for analysis under atmospheric conditions. The collision of the charged droplets with the surface leads to the ionization and desorption of the analyte (35). Then, the ions produced in the gas phase are sampled by an atmospheric interfaced mass analyzer. DESI has been used to create two-dimensional images related to the distribution of lipid species in human tissues (36). 

Brtigger B, Erben G, Sandhoff R, Wieland FT, Lehmann WD. Quantitative analysis of biological membrane lipids at the low picomole level by nano electrospray ionization tandem mass spectrometry. Proc. Natl. Acad. Sci. U.S.A. 1997 94 2339-2344. 

Han X. Characterization and direct quantitation of ceramide molecular species from lipid extracts of biological samples by electrospray ionization tandem mass spectrometry. Anal. Biochem 2002 302 199-212. 

Cai SS, Syage JA. Comparison of atmospheric pressure photoionization, atmospheric pressure chemical ionization, and electrospray ionization mass spectrometry for analysis of lipids. Anal. Chem. 2006 78 1191-1199. 

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