[54] Thin-layer chromatography of lipids

Skipski, Vladimir P.; Barclay, Marion

doi:10.1016/s0076-6879(69)14056-2

Cited by 510 publications

(257 citation statements)

References 121 publications

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“…The plates were developed in two dimensions with chloroform-methanol-concentrated NH4OH (18:14:3, v/v) in the first direction, and chloroformacetone-methanol-acetic acid-water (6:8:2:2:1, v/v) in the second (14). The phospholipids were identified by co-chromatography with authentic standards and by group-specific reagents such as molybdenum, ninhydrin, Dragendorf and periodate-Schiff reagents (17). For quantitative determination, the phospholipid spots were localized with iodine vapor, scraped from the plate, and analyzed for phosphorus by the method of Rouser et al (14).…”

Section: Methodsmentioning

confidence: 99%

Membrane Lipids in Senescing Flower Tissue of Ipomoea tricolor

Beutelmann

Kende²

1977

Plant Physiol.

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Rib segments excised from flower buds ofIpomoea tricolor Cav Labelng experiments using 33P as marker showed that the rate at which radioactivity was lost from phospholipids during aging was parallel to the rate at which the level of total phospholipids declined. Exogenously applied ethylene accelerated the loss of phospholipid and the senescence of rib segments while benzyladenine retarded both of these processes.Ag+, which counteracts the effect of ethylene in many plants, inhibited rolling up of the rib segments but did not affect either spontaneous and ethylene-induced ethylene generation, or phospholpid loss.In contrast, Co2+, a purported inhibitor of ethylene synthesis, reduced ethylene production, rolling up, and phosphoUpid loss. The inhibition of ethylene-induced rolling up by Co2+ could not be overcome with exogenous ethylene, however.Our results indicate that phospholipid loss is a marker for membrane degradation in rib segments. Changes in membrane integrity and in cellular compartmentation may be the basis for ethylene synthesis during aging of flower tissue.

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Section: Methodsmentioning

confidence: 99%

Membrane Lipids in Senescing Flower Tissue of Ipomoea tricolor

Beutelmann

Kende²

1977

Plant Physiol.

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“…The organic phase was then evaporated to dryness under a current of nitrogen, and the residue was dissolved in 200 μl of hexane. Phospholipids were separated by thin layer chromatography according to the method of Skipsky & Barclay (1969). Fatty acid methyl esters of phospholipids were prepared by the method of Lepage & Roy (1986) and analyzed in a Hewlett-Packard 5890 series II gas chromatograph equipped with a flame ionization detector.…”

Section: Cell Death Assaymentioning

confidence: 99%

Influence of environmental medium on membrane fatty acid composition of Reuber H35 hepatoma cells

García-Pelayo

Garcia-Peregrin

Martínez-Cayuela

2013

Frontiers in Life Science

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Cell membrane proteins are sensitive to their fatty acid environment; however, the effects of those fatty acids are not completely understood. In this work, we have optimized a model for the study of membrane protein regulation, which allows the investigation of lipid metabolism in cultured tumor cells. The membrane fatty acid composition of Reuber H35 hepatoma cells was modified by enriching phospholipids in individual fatty acids of different length and degree of saturation. Lauric, myristic, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids were supplemented to the culture medium of cells growing in presence or absence of lipoproteins. The four fatty acids were incorporated into membrane phospholipids in a dosedependent manner. Lauric and myristic acids were actively metabolized, increasing percentages of longer and unsaturated chain fatty acids, but they did not modify the unsaturated index. As expected, EPA and DHA significantly decreased the unsaturated index. Incorporation of EPA and DHA was accompanied of a significant decrease of total monounsaturated fatty acids and an increase of total saturated fatty acids. Our results demonstrate that Reuber H35 hepatoma cells constitute a suitable model to manipulate the lipid composition of the cell membranes and perform functionality studies on the membrane proteins in response to their lipid environment.

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“…We separated the phuspholipids and neutral lipids by developing duplicate plates in hexane-ether-acetic acid (80:20:1 [vol/vol]) (45). The chromatographed lipids were visualized by treatment with iodine vapor.…”

Section: Lipid Extraction and Determination Of Radioactivitymentioning

confidence: 99%

Biosynthesis of high density lipoprotein by chicken liver: conjugation of nascent lipids with apoprotein A1.

Banerjee¹,

Redman²

1984

The Journal of Cell Biology

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To study the assembly of newly synthesized lipids with apoprotein A1, we administered [2-3H]glycerol to young chickens and determined the hepatic intracellular sites of lipid synthesis and association of nascent lipids with apoprotein A1. [2-3H]glycerol was rapidly incorporated into hepatic lipids, reaching maximal levels at 5 min, and this preceded the appearance of lipid radioactivity in the plasma. The liver was fractionated into rough and smooth endoplasmic reticulum and Golgi cell fractions. The isolated cell fractions were further subfractionated into membrane and soluble (content) fractions by treatment with 0.1 M Na2CO3, pH 11.3. At various times, the lipid radioactivity was measured in each of the intracellular organelles, in immunoprecipitable apoprotein A1, and in materials that floated at buoyant densities similar to those of plasma lipoproteins. Maximal incorporation occurred at 1 min in the rough endoplasmic reticulum, at 3-5 min in the smooth endoplasmic reticulum, and at 5 min in the Golgi cell fractions. The majority (66-93%) of radioactive glycerol was incorporated into triglycerides with smaller (4-27%) amounts into phospholipids. About 80% of the lipid radioactivity in the endoplasmic reticulum and 70% of that in the Golgi cell fractions was in the membranes. The radioactive lipids in the content subfraction were distributed in various density classes with most nascent lipids floating at a density -<1.063 g/ ml. Apoprotein A1 from the Golgi apparatus, obtained by immunoprecipitation, contained sixfold more nascent lipids than did that from the endoplasmic reticulum. These data indicate that [2-3H]glycerol is quickly incorporated into lipids of the endoplasmic reticulum and the Golgi cell fractions, that most of the nascent lipids are conjugated with apoproteins A1 in the Golgi apparatus, and that very little association of nascent lipid to apoprotein A1 occurs in the endoplasmic reticulum.High density lipoprotein (HDL) t represents one of the major classes of plasma lipoproteins. Plasma HDLs are small, spherical particles, -8-10 nm diam, which float at a density of between 1.063 and 1.21 g/ml and consist of a layer of phospholipids, free cholesterol, and apoproteins surrounding a membrane lipid core composed mainly of cholesterol esters and triglycerides (8,16,20,28,46).In mammals the study of HDL synthesis and secretion is complicated by the fact that there is more than one site of apoprotein and lipid synthesis. In addition, there are several Abbreviations used in this paper. HDL, high density lipoprotein; RER and SER, rough and smooth endoplasmic reticulum, respectively; VLDL, very low density lipoprotein. apoproteins that are shared by both very low density lipoprorein (VLDL) and HDL (8,16,19,20,46). In recent years, the avian species has attracted considerable attention as an animal model for the study of the biosynthesis and secretion of HDL. It has been reported that in young chickens, plasma HDL is 92% of the total serum lipoproteins (33), that 90% of the protein content of H...

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[54] Thin-layer chromatography of lipids

Cited by 510 publications

References 121 publications

Membrane Lipids in Senescing Flower Tissue of Ipomoea tricolor

Membrane Lipids in Senescing Flower Tissue of Ipomoea tricolor

Influence of environmental medium on membrane fatty acid composition of Reuber H35 hepatoma cells

Biosynthesis of high density lipoprotein by chicken liver: conjugation of nascent lipids with apoprotein A1.

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