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Factors influencing the synthesis of monoester and diester from polyethylene glycol400 and oleic acid by lipase.

Factors influencing the synthesis of mono- and diester of polyethylene glycol400 (PEG400) oleate with immobilized lipase from Candida sp.1619 as biocatalyst were investigated. Mono- and diester were formed with different molar ratios of the substrates after reacting for 6 hours. The amount of monoester to diester formed was in the range of 3.5:1 to 4.1:1 when the molar ratio of acid to PEG400 was 0.25:1 to 2:1. Almost equal amounts of mono- and diesters were produced when the molar ratio of acid to PEG400 was 3:1 to 8:1. When the equilibrium of reaction was reached (22 hours), only diester was found in the reaction mixture with different molar ratio of substrates. The amount of monoester to diester was 1:3.2 in the reaction system containing hexane, even the molar ratio of acid to PEG400 was 2:1.

Candida↗

The effect of phospholipids and fatty acids on tight-junction permeability and bacterial translocation.

The activity of phospholipase A2 (PLA2) is elevated in the intestinal epithelia of patients with inflammatory bowel disease (IBD). We recently reported that PLA2 mediates hydrolysis of phosphatidylcholine (PC) to lysophosphatidylcholine (L-PC) when both are applied to the apical surface of cultured EC monolayers, resulting in increased bacterial translocation (BT) and decreased transepithelial electrical resistance (TEER). Free fatty acids (FFA) are the other products of this reaction, however, their effect on Caco-2 cell permeability has not been reported. In addition to PC, other luminal phospholipids are present at the surface of the enterocyte. PLA2 may also mediate the hydrolysis of luminal phospholipids other than PC. The aim of this study was to examine the effects of phospholipids other than PC and common FFA on intestinal epithelial permeability and BT. Human Caco-2 enterocytes were grown to confluence on porous filters in the apical chamber of a two-chamber cell-culture system. Monolayer integrity and tight-junction permeability were measured as TEER. First, common FFA released by PC hydrolysis were determined using thin-layer chromatography (TLC). In separate experiments, monolayers were treated with phosphatidylethanolamine (PE), lysophosphatidylethanolamine (L-PE), or palmitoleic acid, oleic acids, linoleic acids, and arachidonic acid solubilized in solution with PC. The magnitude of BT was determined 2 h after treatment by adding Escherichia coli C25 to the apical chamber followed by quantitatively culturing basal-chamber samples. Statistical analysis was by the Kurosaki-Wallis test. TLC of PC samples incubated with PLA2 on the apical surface of Caco-2 monolayers demonstrated the production of palmitoleic acid, oleic acids, linoleic acids, and arachidonic acid. L-PE significantly decreased TEER compared to controls, but to a lesser degree than L-PC alone. L-PE had no effects on BT. Palmitoleic acid and oleic acid likewise significantly decreased TEER compared to controls, however, less than L-PC. All FFA tested had no effect on BT. Phospholipids applied to the apical surface of enterocytes, such as those found in vivo in mucus, can be hydrolyzed by the enzyme PLA2 resulting in lysophospholipid and FFA species that can alter enterocyte monolayer permeability. However, FFA and L-PL, other than L-PC, appear to have no effect to stimulate BT. This observation may have clinical implications in the pathogenesis and treatment strategies for IBD patients in whom enterocyte PLA2 activity has been shown to be elevated.

Bacterial Translocation↗

The use of fluorescence resonance energy transfer to study the disintegration kinetics of liposomes containing lysolecithin and oleic acid in rat plasma.

PURPOSE: To validate Fluorescence Resonance Energy Transfer (RET) as method to monitor disintegration of fluorescently labeled liposomes varying in lysolecithin/oleic acid (equimolar) content, lysolecithin fatty acid composition and vesicle size in rat blood plasma and buffer. METHODS: NBD-PE and Rho-PE were used for RET. The measurements were performed on a Perkin Elmer LS-50 spectrofluorimeter. Liposomes were prepared by the extrusion method. RESULTS: Analysis of the RET data was optimised using a fitting procedure to correct for fluorescence interference by plasma. The disintegration patterns of liposomes could be described by a biexponential decay model. Disintegration rate increased at increasing lysolecithin/oleic acid content and decreasing size. In contrast, all liposomes showed no disintegration in buffer. CONCLUSIONS: RET is a suitable method to monitor liposome disintegration in non-diluted plasma. Rate and extent of liposome disintegration increases at decreasing liposome size and increasing lysolecithin/oleic acid content.

Animals↗