Analysis of Lipid Extracts

Positive-ion MALDI mass spectrum of PE species is characterized by the presence of a specific fragment ion corresponding to the loss of the phosphoethanolamine head group [93]. PE species can also be ionized in the negative-ion mode with a lower sensitivity in comparison to that in the positive-ion mode. Negative-ion MALDI mass spectra of PE species are usually dominated by the matrix adducts of PE. 

Many classes of polar lipids have been characterized by MALDI-MS [94]. MALDI-MS has also been employed for characterization of nonpolar lipid classes such as cholesterol and TAG species. Positive-ion MALDI mass spectra of TAG species exclusively display the sodiated species. MALDI mass spectra of TAG species also display the ions corresponding to the loss of sodium fatty acyl carboxy-late(s) due to postsource decay [93]. An extensive list of lipid classes detected by MALDI-MS and corresponding matrices has been documented [101], which should be consulted for those interested. Previously, postsource decay techniques have been commonly employed for characterization of polar lipids [93]. However, since both MALDI-TOF/TOF and MALDI Q-TOF mass spectrometers are now commercially available, true tandem MS analysis of lipids by MALDI-MS is feasible [102-104]. 

MALDI-MS has already been successfully applied for direct analysis of lipids in tissue samples. This topic is extensive described in Chapter 12. 

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Sacchi, R., Medina, I., Aubourg, S.P., Guidicianni, I., Paolilli, L. and Addeo, F. (1993) Quantitative high resolution 13C NMR analysis of lipids extracted from the white muscle of Atlantic tuna (Thurmus alalumga). J. Agric. Food Chem., 41, 1247-1253. 

Galanos, D.S. and Kapoulas, V.M. (1 965) Preparation and analysis of lipid extracts from milk and other tissues. Biochimica et Biophysica Acta Lipids and Lipid Metabolism 98, 278-292. 

Great advances have been made in the direct analysis of lipid extracts (i.e., those requiring minimal preparative fractionation) by the use of dedicated analytical instrumentation such as TLC-FID and HPLC. Mass spectrometry enables the combination of aspects of lipid separation by molecular weight and structural detail from techniques of ionization-induced fragmentation. Combined with chromatographic procedures, this represents the most powerful but expensive tool available for lipid analysis. 

Holzer et al. employed Curie-point pyrolysis to analyze microbial fatty acid by in situ methylation with trimethylanilinium hydroxide (TMAH). The fatty acid methyl ester profiles produced during pyrolysis of the whole cell agreed well with the analysis of lipid extracts from the same microorganism. Dworzanski et al. also used pyrolytic methylation-GC to profile fatty acids in whole cells of E. coli, Mycobacterium tuberculosis, and B. subtilis. Curie-point pyrolysis coupled to GC/ion trap MS was used by Snyder et al. " to characterize 14 strains representing 9 bacterial species for their lipid biomarker content. A quartz tube Pyroprobe from... 

Methods of the Analysis of Lipids Extracted from Human Arteries and Other Tissues... 

Lipidomics smdy has been broadly applied to explore the metabolic origin of diabetes and obesity eomplieations. For example, lipidomic analysis of lipid extracts from white adipose tissue in young healthy monozygous twins with different body weights was performed [20]. The study led to a finding that obese co-twins compared to... 

Because of the small amounts of sample that are usually obtained, coupled GC-MS is the method of choice for analysis of volatile pheromones. The analysis of the less-volatile lipids and polar pheromone components may require derivatization and microchemical tests, both to improve chromatographic characteristics and to provide information about the structures. It is likely that chromatographic techniques with high separation power and high sensitivity for polar compounds, such as coupled capillary electrophoresis-mass spectrometry, will prove useful for analysis of spider extracts in future studies. 

Numerous studies have indicated that pro-inflammatory mediators (cytokines) are involved in the destruction of the insulin-producing p-cells of the pancreas in the development of type I diabetes. Tabatabaie et al. introduced cytokines and PBN into the pancreas of rats. The analysis of pancreatic extracts revealed that the cytokines stimulate the formation of lipid radicals. Radical generation did not occur in rats treated with streptozotocin, which destroys the P-cells.33 Evidence for the role of radicals in diabetes has also been provided by spin trapping studies in pancreatic homogenates, showing that streptozotocin, which is often used to induce the condition in laboratory animals, stimulates OH production.332 Other workers, using EPR to observe the decay of a spin probe in the abdomen of mice (at 1.2 GHz), have demonstrated that strep-... 

D Holme and H Peck, Analytical Biochemtstiy, 3rd ed (1998), Addison Wi li Longman (New York), pp 406-442 Structure, function, and analysis of lipids C Matthews, K van Holde, and K Ahern, Biochemistry, 3rd ed (2000), Benjamin/ Cummings (San Francisco), pp 315-357 Lipid structure and function G Patton, S Cann, H Brunengraber, and J. Lowenstein, in Methods in Enzymology, Vol 72, J Lowenstein, Editor (1981), Academic Press (New York), pp 8-20 Separation of fatty acid methyl esters by gas chromatography on capillary columns N Radin, in Methods in Enzymology, Vol 72, J M Lowenstein, Editor (1981), Academic Press (New York), pp 5-7 Extraction of lipids with hexane-isopropanol L Stryer, Biochemistry, 4th ed (1995), W H Freeman (New York), pp 263-270, 603-606 Lipid structure and function... 

The two procedures of lipid extraction and saponification can be combined as a prolonged or hot alkaline alcoholic extraction, a method that is satisfactory for certain routine analysis (44). 

Several new methodologies have been developed in recent years for the surface analysis of lipids. These methods allow direct detection of lipids from surfaces such as tissue sections and intact cells without prior extraction and thus enable the determination of spatial distribution of the lipids. There are three basic requirements for the surface analysis. The analytes of interest must be desorbed from the surface by the interaction of a sampling probe such as spray, laser, or plasma. The desorbed analytes must first be ionized and then... 

Analysis of lipid pyrolysates using Py-GC/MS was performed [12], but it lacked a good separation that would have shown the individual compounds. Extracted ion... 

The solubility of the seed material in carbon dioxide was defined as the intial concentration prior to observation of these depletion effects. The solubility was found to range from 0.016 wt.% to 0.6 wt.% (Figure 4). Analysis of the extracts revealed that the monocrotaline content was very low (<0.05 wt.%), and that the extracts were predominantly lipid material as expected. 

It is worth noting that in many cases some of the resorcinolic lipids present in biological material still remain structurally uncharacterised. For instance, chromatographic analysis of acetone extracts from cereal grains shows the presence of at least 4 other components that probably belong to the group of resorcinolic lipids. Two of them have been identified as 5-(2-oxoalkyl) resorcinol and 5-(2-oxoalkenyl) resorcinols [178] and the another as a 5-(2-hydroxyalkyl) resorcinol [181]. 

Major applications of SFE-SFC are somewhat limited at the moment to the analysis of lipids and pesticides from foods and similar matrices and different types of additives used in the production of polymers [79,146,188-194]. The approaches used cover a wide range of sophistication and automation from comprehensive commercial systems to simple laboratory constructed devices based on the solventless injector [172,174,175,188]. Samples usually consist of solid matrices or liquids supported on an inert carrier matrix. Aqueous solutions are often analyzed after solid-phase extraction (SPE-SFE-SFC) to minimize problems with frozen water in the interface [178,190]. The small number of contemporary applications of SFE-SFC reflects a lack of confidence in supercritical fluid chromatography as a separation technique and competition for... 

Liescheski, P.B. 1996. Supercritical fluid extraction coupled directly to infrared spectroscopy for the analysis of lipids in food. Seminars in Food Analysis, 1, 85. 

Where the fat in a meat or meat product sample is to be characterized in terms of the fatty acid profile, extraction of the fat with chloroform/methanol is required. This solvent mixture, while it may not give complete fat extraction, is used to ensure no chemical change to the lipids and the extraction of phospholipids. Fatty acid analysis of the extracted fat is undertaken by formation of volatile methyl esters of the fatty acids (ISO 5509 2000) and determination by gas chromatography (ISO 5508 1990). 

Iodine. The most convenient and commonly used technique is exposure to iodine vapours. Iodine crystals are put in one of the covered tanks and placed under the fume hood, vapours of iodine being toxic. After it is freed from the solvent, the TLC plate is placed in the iodine tank and the spots are visualized. This is a qualitative technique, although some researchers use it for quantitation by densitometry. The theory behind iodine exposure is that the iodine is physically absorbed into the lipid spot. The disadvantage with this technique is that the saturated lipid samples do not take up iodine easily and, moreover, when unsaturated molecules are left in iodine for a sufficiently long time they tend to react with it chemically. Iodine is added at the double bonds and produces artifacts, and this has been proved by GC analysis of material extracted from plates after exposure to iodine (Vioque and Holman, 1962). 

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