Fatty acid ethyl esters in hair as markers of alcohol consumption. Segmental hair analysis of alcoholics, social drinkers, and teetotalers.
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BACKGROUND: Fatty acid ethyl esters (FAEEs) are products of nonoxidative ethanol metabolism. After incorporation in hair, they should be suitable long-term markers of alcohol abuse. METHODS: Hair samples from 19 alcoholics in a treatment program, 10 fatalities with verified excessive alcohol consumption, 13 moderate social drinkers who consumed up to 20 g ethanol/day, and 5 strict teetotalers were analyzed in 1-12 segments for four FAEEs (ethyl myristate, ethyl palmitate, ethyl oleate, and ethyl stearate) by external degreasing with n-heptane, extraction with a dimethyl sulfoxide-n-heptane mixture, headspace solid-phase microextraction of the extracts, and gas chromatography-mass spectrometry with deuterated internal standards. The n-heptane washings were analyzed in the same way for FAEEs from the hair surface. RESULTS: The sum of the four ester concentrations in hair calculated for the proximal 0-6 cm segment was 2.5-13.5 ng/mg (mean, 6.8 ng/mg) for the fatalities, 0.92-11.6 ng/mg (mean, 4.0 ng/mg) for 17 of the alcoholics in treatment, 0.20-0.85 ng/mg (mean, 0.41 ng/mg) for the moderate social drinkers, and 0.06-0.37 ng/mg (mean, 0.16 ng/mg) for the teetotalers. In almost all cases the segmental concentrations increased from proximal to distal. There was no agreement between the self-reported drinking histories of the participants and the FAEE concentrations along the hair length. Ethyl oleate was the dominant ester in all samples. CONCLUSIONS: FAEEs are deposited in hair mainly from sebum. Despite large individual differences, FAEE hair concentrations can be used as markers for excessive alcohol consumption with relatively high accuracy. (+info)
Interaction of phospholipid vesicles with cells. Endocytosis and fusion as alternate mechanisms for the uptake of lipid-soluble and water-soluble molecules.
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Depending on their phospholipid composition, liposomes are endocytosed by, or fuse with, the plasma membrane, of Acanthamoeba castellanii. Unilamellar egg lecithin vesicles are endocytosed by amoeba at 28 degrees C with equal uptake of the phospholipid bilayer and the contents of the internal aqueous space of the vesicles. Uptake is inhibited almost completely by incubation at 4 degrees C or in the presence of dinitrophenol. After uptake at 28 degrees C, the vesicle phospholipid can be visualized by electron microscope autoradiography within cytoplasmic vacuoles. In contrast, uptake of unilamellar dipalmitoyl lecithin vesicles and multilamellar dipalmitoyl lecithin liposomes is only partially inhibited at 4 degrees C, by dinitrophenol and by prior fixation of the amoebae with glutaraldehyde, each of which inhibits pinocytosis. Vesicle contents are taken up only about 40% as well as the phospholipid bilayer. Electron micrographs are compatible with the interpretation that dipalmitoyl lecithin vesicles fuse with the amoeba plasma membrane, adding their phospholipid to the cell surface, while their contents enter the cell cytoplasm. Dimyristoyl lecithin vesicles behave like egg lecithin vesicles while distearoyl lecithin vesicles behave like dipalmitoyl lecithin vesicles. (+info)
The mycocidal, membrane-active complex of Cryptococcus humicola is a new type of cellobiose lipid with detergent features.
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The chemical composition of the mycocidal complex (formerly known as microcin) secreted by Cryptococcus humicola was investigated by chemical, mass spectrometric and nuclear magnetic resonance methods. The results indicate that the mycocidal complex is composed of glycolipids with a highly acetylated (up to five acetyl groups) cellobiose backbone [beta-D-Glcp-(1'-->4)-beta-D-Glcp] linked to the omega-hydroxyl group of alpha,omega-dihydroxy palmitate [16:0-alpha,omega-di-OH] with an unsubstituted carboxyl group. The acyl chain forming aglycon can be replaced by [18:0-(alpha,omega-di-OH)], [18:0-(alpha,omega-1,omega-tri-OH)], and [18:0-(alpha,omega-2,omega-tri-OH)]. The complex has a comparatively high surface activity; 0.5 mg/ml of it reduced the surface tension of 0.1 M NaHCO(3) from 71 mN/m to 37 mN/m and interfacial tension against n-hexadecane from 39 mN/m to 10 mN/m. The critical micelle concentration of the complex at pH 4.0, determined by the fluorometric method with N-phenyl-1-naphthylamine as fluorescent probe and by the De Nouy ring method, was 2 x 10(-5) M (taking the average molecular mass of the complex to be 750); it did not depend on the presence of 100 mM KCl and was an order of magnitude higher at pH 7.0. By fluorescence resonance energy transfer spectroscopy with N-(7-nitro-2,1,3-benzoxadiazol-4-yl)-phosphatidylethanolamine as energy donor and N-(rhodamine B sulfonyl)-phosphatidylethanolamine as energy acceptor the complex was shown to intercalate into the liposomal lipid matrix. Primary lesions caused by the complex in planar lipid bilayers were revealed as short-living current fluctuations of a broad spectrum of amplitudes. The mycocidal effect of the complex is suggested to be associated with its detergent-like properties. (+info)
Acute effects of fatty acids on insulin secretion from rat and human islets of Langerhans.
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Fatty acids have both stimulatory and inhibitory effects on insulin secretion. Long-term exposure to fatty acids results in impaired insulin secretion whilst acute exposure has generally been found to enhance insulin release. However, there are conflicting data in the literature as to the relative efficacy of various fatty acids and on the glucose dependency of the stimulatory effect. Moreover, there is little information on the responses of human islets in vitro to fatty acids. We have therefore studied the acute effects of a range of fatty acids on insulin secretion from rat and human islets of Langerhans at different glucose concentrations. Fatty acids (0.5 mM) acutely stimulated insulin release from rat islets of Langerhans in static incubations in a glucose-dependent manner. The greatest effect was seen at high glucose concentration (16.7 mM) and little or no response was elicited at 3.3 or 8.7 mM glucose. Long-chain fatty acids (palmitate and stearate) were more effective than medium-chain (octanoate). Saturated fatty acids (palmitate, stearate) were more effective than unsaturated (palmitoleate, linoleate, elaidate). Stimulation of insulin secretion by fatty acids was also studied in perifused rat islets. No effects were observed at 3.3 mM glucose but fatty acids markedly potentiated the effect of 16.7 mM glucose. The combination of fatty acid plus glucose was less effective when islets had been first challenged with glucose alone. The insulin secretory responses to fatty acids of human islets in static incubations were similar to those of rat islets. In order to examine whether the responses to glucose and to fatty acids could be varied independently we used an animal model in which lactating rats are fed a low-protein diet during early lactation. Islets from rats whose mothers had been malnourished during lactation were still able to respond effectively to fatty acids despite a lowered secretory response to glucose. These data emphasise the complex interrelationships between nutrients in the control of insulin release and support the view that fatty acids play an important role in glucose homeostasis during undernutrition. (+info)
Mass spectrometric analysis of GAP-43/neuromodulin reveals the presence of a variety of fatty acylated species.
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GAP-43 (neuromodulin) is a protein kinase C substrate that is abundant in developing and regenerating neurons. Thioester-linked palmitoylation at two cysteines near the GAP-43 N terminus has been implicated in directing membrane binding. Here, we use mass spectrometry to examine the stoichiometry of palmitoylation and the molecular identity of the fatty acid(s) attached to GAP-43 in vivo. GAP-43 expressed in either PC12 or COS-1 cells was acetylated at the N-terminal methionine. Approximately 35% of the N-terminal GAP-43 peptides were also modified by palmitate and/or stearate on Cys residues. Interestingly, a variety of acylated species was detected, in which one of the Cys residues was acylated by either palmitate or stearate, or both Cys residues were acylated by palmitates or stearates or a combination of palmitate and stearate. Depalmitoylation of membrane-bound GAP-43 did not release the protein from the membrane, implying that additional forces function to maintain membrane binding. Indeed, mutation of four basic residues within the N-terminal domain of GAP-43 dramatically reduced membrane localization of GAP-43 without affecting palmitoylation. These data reveal the heterogeneous nature of S-acylation in vivo and illustrate the power of mass spectrometry for identification of key regulatory protein modifications. (+info)
Fatty Acid ethyl esters in scalp, pubic, axillary, beard and body hair as markers for alcohol misuse.
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AIMS: This study examined the possibility of using hair samples other than scalp hair for analysis of fatty acid ethyl esters (FAEE) as markers for alcohol misuse. METHODS: Samples of scalp and pubic hair and, if available, axillary, beard and body hair of one teetotaller, five moderate social drinkers and 22 fatalities were analysed for FAEE by head-space solid-phase microextraction and gas chromatography-mass spectrometry. The sum of the concentrations of ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate (C(FAEE)) in the hair samples was compared with information about the drinking behaviour of the individuals. RESULTS: Although there were large differences in the analyte concentrations in hair from different sites in the same individual, cases of chronic excessive alcohol consumption were characterized by C(FAEE) > 1.0 ng/mg in almost all samples. CONCLUSION: FAEE concentrations in hair other than scalp hair can be used as markers of chronic alcohol misuse. (+info)
Ruminal biohydrogenation of fatty acids from high-oleate sunflower seeds.
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The objective of these experiments was to examine methods of modifying the fatty acid composition of bovine tissues. In the first experiment, four steers were fitted with duodenal fistulas and were assigned to four diets in a Latin square design. The steers were fed a control diet or the same diet containing 10% high-oleate partially crushed sunflower seeds, serum-coated sunflower seeds, and heat-treated, serum-coated sunflower seeds for 5 d. Samples of digesta and feces were collected on d 5. The inclusion of sunflower seeds (plain or serum-coated) in the diet increased (P less than .05) the digesta concentration of stearate. The percentage of stearate in the digesta and feces was increased (P less than .05) from 51 to 67% and from 64 to 74%, respectively, when steers were fed the untreated sunflower seed. The fecal concentration of oleate was increased (P less than .05) by dietary sunflower seeds in steers that were fed the serum-coated, unheated sunflower seeds. In a second experiment, heifers (four per group) were fed a corn-based control diet or diets containing 10% of high-oleate sunflower oil encapsulated with calcium alginate, either plain, coated with blood meal, or with blood meal integrated into the pellet. After 50 d on treatment, samples of perianal adipose tissue were obtained by biopsy. The fatty acid composition of the adipose tissue was not modified by the inclusion of the encapsulated oleate in the diet. In summary, limited ruminal bypass of sunflower seed oleate was accomplished with sunflower seed but not with encapsulated oleate. (+info)
Modulation of P-glycoprotein-mediated drug transport by alterations in lipid fluidity of rat liver canalicular membrane vesicles.
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P-glycoprotein (P-gp) is believed to function as an ATP-dependent efflux pump for natural product anti-cancer drugs in multidrug-resistant (MDR) tumor cells and in certain normal tissues. P-gp has been localized to the apical plasma membrane of the bile canaliculus where it has been shown to transport [3H]daunomycin. In this study, we investigated whether alterations in membrane lipid fluidity of canalicular membrane vesicles (CMV) could modulate the P-gp-mediated accumulation of [3H]daunomycin and [3H]vinblastine. Accumulation of both cytotoxic agents was stimulated by ATP, exhibited temperature dependence and osmotic sensitivity, and followed Michaelis-Menten kinetics. Alterations in CMV lipid fluidity were induced by the known fluidizers, 2-(2-methoxyethoxy)ethyl 8-(cis-2-n-octylcyclopropyl)octanoate (A2C) and benzyl alcohol, and were assessed by fluorescence polarization techniques using the fluorescent probe, 1,6-diphenyl-1,3,5-hexatriene (DPH). Both A2C (2.5-5.0 microM) and benzyl alcohol (10-20 mM) produced a dose-dependent increase in CMV lipid fluidity. Moreover, both fluidizers, at the above doses, significantly inhibited (p < 0.05) the ATP-dependent accumulation of [3H]daunomycin. [3H]Vinblastine accumulation was also inhibited by A2C (p < 0.05). Lower doses of A2C (0.6 microM) and benzyl alcohol (1 mM) failed to influence either lipid fluidity or P-gp-mediated drug accumulation. Kinetic analysis revealed that A2C (5.0 microM) noncompetitively inhibited [3H]daunomycin accumulation and uncompetitively inhibited [3H]vinblastine accumulation with apparent Ki values of approximately 1.5 and approximately 1.2 microM, respectively. Verapamil competitively inhibited P-gp-mediated accumulation of [3H]daunomycin but failed to alter the fluidity of CMV. Taken together, the present results demonstrate that while increases in membrane fluidity of CMV are not necessarily required to inhibit P-gp-mediated drug accumulation, they can inhibit these processes, at least in CMV. Alterations in the physical state of CMV, therefore, appear to be at least one important modulator of P-gp function. (+info)