Identification of fatty acids by electrospray mass spectrometry and tandem mass spectrometry.

Low-energy negative-ion electrospray mass spectrometry (ESI-MS) and ESI-MS/MS were used to characterize saturated and unsaturated fatty acids. The carbon number and degree of unsaturation of fatty acids were determined using ESI-MS, and MS/MS was used to localize some double bond positions of mono-and polyunsaturated fatty acids. For compounds with up to two unsaturated bonds, fragmentation was dominated by loss of H2O from the carboxyl moiety and very low-intensity peaks generated from bonds cleaved at carbons alpha and/or beta to sites of unsaturation. Fragmentation of monounsaturated fatty acids was minimal using this soft method of mass spectrometric analysis, but increased with progressively greater degrees of fatty acid unsaturation. There was extensive hydride migration during ESI-MS/MS of compounds with three or more double bonds. Although this behavior complicated localization of double and triple bonds, the spectra were reproducible. Many peaks could not be definitively assigned to specific product ions, but the spectra of standards and complementary natural products were similar and isobaric compounds could be differentiated. The utility of this technique to examine biological samples was shown by analysis of the fatty acid composition of cod liver oil. Detection limits for negative-ion ESI-MS/MS were at or below 1 pg.

Sterol and steryl ester regulation of phospholipase A2 from the mosquito parasite Lagenidium giganteum.

Lagenidium giganteum, a facultative parasite of mosquito larvae, cannot synthesize sterols, and requires an exogenous source of these lipids in order to enter its reproductive cycle. This parasite grows vegetatively in the absence of sterols, but requires cholesterol or structurally related compounds to produce motile zoospores, which are the only stage capable of infecting mosquitoes. Sterols structurally related to cholesterol and some steryl esters inhibited the activity of L. giganteum phospholipase A2 (PLA2), an enzyme that hydrolyzes fatty acids from the sn-2 position of glycerophospholipids. Sterols that induce reproduction inhibited L. giganteum PLA2 activity, while sterols and steroids that do not support sporulation had minimal effect. Most steryl esters had no effect on enzyme activity, but cholesteryl arachidonate (CA) was a potent inhibitor of parasite PLA2. Not all enzymes partly purified using a DEAE-Sephacel column were affected by these lipids, demonstrating selective inhibition of specific enzymes. Potency was enhanced by up to several orders of magnitude if epoxy fatty acids were esterified to the cholesterol nucleus. The steryl ester pool was dynamic during morphogenesis, and the fatty acid composition of the steryl esters did not mimic total cell or membrane (glycerophospholipid) fatty acid composition as L. giganteum proceeded through its growth cycle. Synthesis of CA and monoepoxy CA by the parasite was confirmed using electrospray mass spectrometry and collision-induced dissociation. Steryl derivatives selectively inhibited PLA2 enzymes from bovine pancreas, snake venom, and human cytoplasmic 85-kDa PLA2.

Isoprenoid-mediated changes in the glycerophospholipid molecular species of the sterol auxotrophic fungus Lagenidium giganteum.

The mosquito pathogenic fungus Lagenidium giganteum (Oomycetes: Lagenidiales) is a sterol auxotroph that can grow vegetatively in the absence of these compounds, but requires an exogenous source of sterols to enter its sexual and asexual reproductive cycles. Electrospray mass spectrometry (MS) and electrospray MS/MS were used to examine three major glycerophospholipid molecular species--glycerophosphocholine (GPC), glycerophosphoethanolamine (GPE) and glycerophosphoinositol (GPI)--from fungal mycelium and nuclei grown in defined medium with and without isoprenoids which induce (cholesterol and ergosterol) or do not induce (squalene, cholestane) reproduction. Testosterone supplementation of defined media inhibited growth of L. giganteum, so the effect of this steroid on phospholipid metabolism could not be assessed. Mycelium grown in defined media supplemented with these isoprenoids produced significantly different quantities of total phospholipid relative to unsupplemented media and to each other, ranging from a mean of 292 micrograms phosphate per g wet weight for cholesterol-supplemented media to 56 micrograms phosphate per g wet weight for mycelium grown in the presence of squalene. A very large percentage of the GPC (69-80 mol%) and GPI (74-79 mol%) molecular species from mycelia and nuclei contained ether linkages. GPE molecular species had 13-20 mol% ether-containing moieties. The elevated levels of ether lipids may be related to the sterol auxotrophic nature of the fungus. Isoprenoid supplementation of defined growth media resulted in many significant changes in molecular species for all three lipid classes. Significant differences (P < 0.05) in the percentage of total cell ether lipids in GPC and GPE were generated by isoprenoid supplements to culture media. Mycelium grown in the presence of the two sterols which induce asexual and sexual reproduction in L. giganteum, cholesterol and ergosterol, had a significantly greater percentage of ether-containing GPE moieties. The glycerolipid species from nuclei isolated from cultures grown with cholesterol and ergosterol were similar to the composition of nuclei isolated from fungus cultured in defined medium without any supplement or supplemented with squalene. The nuclear membrane from mycelia grown in cholestane-supplemented media, however, had a very different glycerophospholipid composition relative to either whole cells or nuclei from cells grown on other media. It appears that one of the reasons that cyclic isoprenoids such as cholestane do not induce fungal reproduction is that they drastically alter the nuclear membrane glycerophospholipid composition.

Complicacoes pos-operatorias imediatas em gastrectomias parciais eletivas. / Immediate post-operative complication of elective partial gastrectomies

Os autores fazem uma analise das complicacoes pos-operatorias imediatas em gastrectomias parciais eletivas. A complicacao mais frequente e a infeccao respiratoria que e favorecida pelo fumo, sonda nasogastrica, anestesia endotraqueal, intervencao em abdome superior. A gastrectomia e uma cirurgia de grande porte e o paciente deve ser preparado para tal, principalmente em sua funcao respiratoria. A boa tecnica cirurgica contribui para diminuir o numero de complicacoes pos-operatorias