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Effects of feeding ethyl-dihomo-gamma-linolenate on rabbit renomedullary lipid composition and prostaglandin production in vitro.

Feeding the ethyl ester of dihomo-gamma-linolenic acid for 25 days to rabbits resulted in increased PGE1 (20 to 30-fold) and PGE2 (1.5-fold) output by a hormone responsive, in vitro, renal papilla preparation. The relative amount of PGE1 increased from less than 5% of PGE2 in controls to 25-35% of PGE2 in the papillae of 20 : 3 omega 6-supplemented animals. During the study renomedullary tri glycerides in the 20 : 3 omega 6-supplemented animals increased 2.8-fold compared to animals fed an equal amount of a control fatty acid mixture, and in addition to a marked enrichment in 20 : 3 omega 6, also contained increased proportions of 20 : 4 omega 6 and longer chain polyenes. The increase in triglyceride content found in the renal medulla was not seen in the renal cortex or liver. There was no increase in renomedullary phospholipid content during the study, and phopholipids of treated animals contained increased proportions of 20 : 3 omega 6 and 20 : 4 omega 6, but not longer chain polyenes. The results indicate that enriching the prostaglandin precursor pool by feeding 20 : 3 omega 6 can alter the type and amount of prostaglandin released by the renal papilla, at least in vitro. Also, the selective changes in amount and long chain polyene content of renomedullary triglycerides during the study suggest some special functions for this lipid class in prostaglandin precursor metabolism.

8,11,14-Eicosatrienoic Acid↗

Lipid composition of Morris hepatoma 5123c, and of livers and blood plasma from host and normal rats.

The lipid composition of Morris hepatoma 5123c was analyzed together with that of liver and blood plasma from both normal and tumor-bearing rats. The results showed that the liver of tumor-bearing rats contained higher amounts of glycerides, choelsteryl esters, free fatty acids and phospholipids than the liver of normal rats. In the blood plasma of tumor-bearing rats, there was an increase of free cholesterol and triglycerides; this latter difference, however, was not statistically significant. Acyl chain changes in the liver of tumor-bearing rats consisted of an increase of palmitic and oleic acids and a decrease of stearic and arachidonic acids in phosphatidylinositol. Morris hepatoma 5123c contained a lower amount of triglycerides than the livers (both host and normal) and showed a significant decrease of total phospholipids when compared to the host liver. The major acyl chain changes found in Morris hepatoma 5123c compared with both normal and host rat livers were: a) a higher percentage of arachidonic acid together with a lower proportion of palmitic acid in cholesteryl esters; b) an increase of stearic and arachidonic acids and a decrease of palmitic acid in triglycerides; and c) a higher level of palmitic and oleic acids associated with a lower percentage of stearic and C22 polyunsaturated acids in phosphatidylcholine.

Animals↗

Lipase-catalyzed monoesterification of 1-O-hexadecylglycerol in organic solvents.

A simple method is presented to esterify 1-O-hexadecyl-rac-glycerol using lipases in different organic solvents. The following fatty acids were used: C14:0, C16:0, C18:0, C18:1, and C18:2. Monoesterification was achieved by using a limiting amount of fatty acid. Both the 1-O-hexadecyl-3-O-acylglycerol and the 2-O-acylglycerol were obtained in a total yield of 75% and a ratio 7:1 in dichloromethane after 3 d. Chromatographic data for the monoesters, useful for the identification of the natural products, are given (gas-liquid chromatography, thin-layer chromatography, reverse-phase thin-layer chromatography). The structure was confirmed by a chemical synthesis of 1-O-hexadecyl-2-O-hexadecanoylglycerol. The 3-O-glyceride was also formed by acyl migration, as the minor component. The monoesters were separated by column chromatography and characterized by 1H and 13C nuclear magnetic resonance spectra.

Catalysis↗

Lipid composition and viscosity of parotid saliva in Sjögren syndrome in man.

Extraction of the dialysed and lyophilized saliva of patients with this syndrome by chloroform-methanol yielded 15.9 +/- 2.4 mg of lipid/100 ml of secretion, a level 2-times higher than in normal individuals. The saliva of such patients also had 3-times more glycolipid and 20-times more phospholipid than normal, but differences in the content of neutral lipids were less apparent. The neutral lipids, however, had higher proportions of glycerides, and lower proportions of cholesterol and cholesteryl esters than normal. Viscosity measurements, made with a cone/plate viscometer at shear rates between 1.15 and 230 s-1, revealed similarities between the saliva of normal individuals and Sjögren's syndrome.

Cholesterol↗

Lipids in human saliva.

A simple and reproducible method of determining the quality and quantity of neutral lipids in human saliva was tested. Parotid, submandibular and whole stimulated saliva were collected from 10 healthy adults. The lipids were extracted by the Folch method. A special method for extraction of glycolipids was also tested but gave no additional recovery. Thin-layer chromatography was used for separating the different lipid classes. The concentrations of total lipids in parotid, submandibular and whole stimulated saliva were 0.2, 0.9 and 1.3 mg/dl, respectively. Cholesteryl esters, cholesterol, triglycerides, diglycerides, monoglycerides and free fatty acids accounted for 96-99 percent of the total salivary lipids. Thus, polar lipids such as phospholipids contributed only a minor fraction, indicating that the lipids are not primarily of membrane origin. Ultracentrifugation of saliva samples at d = 1.21 g ml(-1) showed that the salivary lipids did not float like blood plasma lipoproteins. Therefore, they must be in a different state of aggregation from lipids in blood or lymph. No significant lipase activity of the type that acts on plasma lipoproteins was found in parotid or submandibular saliva. The content of free fatty acids and partial glycerides was high.

Adult↗

Topographic distribution of enzymes involved in glycerolipid synthesis in rat small, intestinal epithelium.

1. The enzymes involved in glycerolphosphate and monoacylglycerol acylation of rat small intestine were more active in villi than in crypts. Monoglyceride acyltransferase (EC 2.3.1.22) was found to be absent from crypts. 2. In the villi, the enzymes are mainly localized in microsomes, although low activities of palmitoyl-CoA synthetase (EC 6.2.1.3), glycerolphosphate acyltransferase (EC 2.3.1.15) and cholinephosphotransferase (EC 2.7.8.2) are found in mitochondria. Mitochondria lack monoglyceride acyltransferase and lysolecithin acyltransferase (EC 2.3.1.23), both of which are involved in the reacylation of alimentary partial glycerides. Therefore, this process is confined to microsomes. 3. The monoacylglycerol and lysolecithin acyltransferases, as well as choline-phosphotransferase, are probably localized within the endoplasmic reticulum, since these enzymes are relatively Nagerse resistant (subtilisin; EC 3.4.2.1, compared with palmitoyl-CoA synthetase and glycerolphosphate acyltransferase, which are highly Nagarse-sensitive and therefore probably localized on the outside of the microsomes (and mitochondria). 4. The physical separation of alimentary product reacylation from de novo synthetic processes provides the basis of metabolic compartmentation observed by other workers. 5. The use of sucrose instead of a salt medium for the isolation and homogenization of small intestinal epithelial cells allowed the separation of mitochondria and microsomes by differential centrifugation without mutual contamination. 6. Phospholipids were found to stimulate glycerolphosphate acylation in vitro. 7. The glycerolphosphate and monoacylglycerol acylation pathways are not competitive.

1-Acylglycerophosphocholine O-Acyltransferase↗

Synthesis of trinitrophenylaminolauric acid and the use of its glyceryl esters for assaying lipase by a spectrophotometric procedure.

Trinitrophenylaminolauric acid was synthesized from omega-aminolauric acid and trinitrobenzenesulfonic acid and then condensed with glycerol to yield mono-, di- and triacylglyceryl esters of this acid. Hydrolysis of these glycerides was followed by isolation of the yellow fatty acid with the aid of one solvent extraction step and estimating its content by spectrophotometry. This procedure was used to assay the activities of lipases from hog pancreas, rat bile, microsomes of rat brain and Rhizopus arrhizus delamar.

Animals↗

Diacylglycerols enhance human neutrophil degranulation responses: relevancy to a multiple mediator hypothesis of cell function.

At 10 microM, 1-0-oleoyl-, 1-0-palmitoyl-, and 1-0-myristoyl-2-0-acetyl-glycerol weakly stimulated neutrophils to release lysozyme, an enzyme in secondary granules, but had no such effect on the release of a primary granule enzyme, beta-glucuronidase. The glycerides (1-10 microM) had a second effect on both granule populations: they enhanced the degranulating potencies of leukotriene B4, platelet-activating factor, a formylated oligopeptide, and C5a by 10- to 30-fold. In contrast, they were much less effective in enhancing responses to ionophore A23187 and partially inhibited responses to phorbol myristate acetate. The diether analogue, 1-0-hexadecyl-2-0-ethylglycerol was inactive in these regards. We suggest that diacylglycerols are a novel class of bioactive products mobilized from phosphoglycerides in stimulated neutrophils; as co-products of this mobilization, platelet-activating factor and leukotriene B4 may interact with diacylglycerols to promote cell function.

Animals↗

Some aspects of metabolic adaptations in lipid metabolism during starvation are mimicked by epinephrine in rat adipocytes.

1. The effects of fasting on the neutral lipid synthesis to insulin and/or epinephrine in isolated fat cells have been examined using [1-14C]glucose. 2. The ability of adipocytes from starved rats to synthesize fatty acids from both labeled substrates was markedly diminished compared to adipocytes from control rats. 3. The response of lipogenic stimulation to insulin at all concentrations tested was greatly diminished in adipocytes from 24 hr starved rats. 4. [1-14C]glucose utilization rates in the absence or in the presence of insulin were not significantly different in adipocytes from 24 hr starved rats as compared with control adipocytes, although basal and insulin stimulated glyceride-glycerol synthesis were significantly higher in starved adipocytes. 5. Epinephrine acutely inhibited [1-14C]acetate incorporation into fatty acids for insulin-stimulated lipogenesis in control adipocytes, in contrast, this lipolytic agent strongly increased [1-14C]glucose conversion to triacylglycerols. 6. In both cases, the differences in lipid synthesis capacities found in both nutritional states were abolished by epinephrine.

Acetates↗

Lipogenic potential of liver from morbidly obese patients with and without non-insulin-dependent diabetes.

Intra-abdominal liver biopsies were obtained during surgery from fasted obese patients with non-insulin-dependent diabetes mellitus (NIDDM), obese normoglycemic controls, and lean controls. Lipid synthesis was studied in freshly isolated hepatocytes and liver homogenates from the three groups of subjects. Incorporation of 3H2O into the lipids of hepatocytes was determined in the absence and presence of insulin (0.1 mumol/L). The activities of five enzymes involved in fatty acid synthesis, and the incorporation of 14C-glycerol-3-phosphate into lipids were determined in liver homogenates. Basal lipid synthesis by hepatocytes was not different in the three groups of patients. Insulin stimulated lipogenesis by 8% +/- 30% in the lean controls, 33% +/- 8% in the obese controls and 17% +/- 6% in the NIDDM patients. No significant differences in the activities of the five enzymes that are involved in de novo fatty acid synthesis among the three groups of patients were observed. Similarly, incorporation of 14C-glycerol-3-phosphate by liver homogenates, in the presence of saturating or submaximal concentrations of fatty acids, did not differ among the three groups. These results show that under the experimental conditions of this study, including the fasted state of the patients, the basal capacity of liver of NIDDM patients to synthesize fatty acids or glycerides is the same as that of liver from obese and lean controls. Thus, it is likely that an increase in fatty acid flux into a liver with normal lipogenic potential may contribute to the increased synthesis of triglycerides by the liver of these patients in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Age related changes in the lipids of the microsomal and the mitochondrial membranes of rat liver and kidney.

The lipid contents of the microsomal and mitochondrial membrane fractions of liver and kidney were determined in 6 and 24 month old rats. A significant age related increase in the molar ratio of cholesterol/phospholipid was observed in all membrane fractions. A significant age related reduction of phospholipid was noted in the microsomal fractions of liver and kidney. The relative amount of phosphatidylethanolamine was found to decrease in all membrane fractions during aging. Membrane glyceride content, however, remained relatively constant with age. Significant increase in oleic acid was seen in the neutral lipid of both liver and kidney and in the polar lipid of kidney. Significant increase in docosahexaenoic acid and significant decrease in linoleic acid were seen in the polar lipid of the liver membrane fractions. Possible alterations in membrane physiochemical properties and in membrane function due to these age related lipid changes are discussed.

Aging↗

Impact of pesticides on lipid metabolism in the freshwater catfish, Clarias batrachus, during the vitellogenic phase of its annual reproductive cycle.

Specimens of either sex of the freshwater catfish Clarias batrachus were exposed to safe and sublethal concentrations of an organochlorine, gamma-BHC (2 and 8 ppm), and an organophosphorus compound, malathion (1 and 4 ppm), for 4 weeks during the vitellogenic phase of their annual reproductive cycle. The effects on total lipid and its various fractions, viz., free fatty-acids, monoglycerides, diglycerides, triglycerides, phospholipids, free cholesterol, and esterified cholesterol, were studied in the liver, plasma, gonads, and muscle. Except for elevated liver lipid in the male in response to malathion, no significant change in total lipid could be observed following pesticide exposure. However, various lipid fractions responded differently to two concentrations of the pesticides. Both pesticides affected the metabolism of nonpolar and less polar lipids alike. Malathion inhibited only mobilization of hepatic phospholipid to gonads but not its hepatic biosynthesis, whereas gamma-BHC reduced its synthesis in the liver as well. These pesticides seemed to restrict the conversion of esterified cholesterol into free cholesterol without affecting the biosynthesis of cholesterol as such. In both sexes, esterification of free fatty acids to acyl glycerides and their mobilization from liver to gonads seemed to be restricted as a result of pesticides action.

Animals↗

Biochemistry of the sarcocyst of Sarcocystis fusiformis of buffalo Bubalus bubalis.

Sarcocysts of Sarcocystis fusiformis from oesophageal muscles of naturally-infected Indian water buffalo (Bubalus bubalis) were analysed for total lipids, phospholipids, cholesterol, fatty acids and glycerides and total protein. Protein and phospholipids constituted the major portion of the sarcocyst. Acetylcholinesterase and glutamate-oxalo-acetate transaminase activities when assayed were higher than glutamate-pyruvate transaminase in sarcocysts.

Acetylcholinesterase↗

Analysis of mono-, di- and triglycerides in pharmaceutical excipients by capillary supercritical fluid chromatography.

Mono-, di- and triglycerides are important components of oils, fats and other natural products. Since in general fatty acids are mixtures and glycerol can be differently substituted, finger-prints of the composition are suitable for better characterization. Since capillary supercritical fluid chromatography (SFC) employing carbon dioxide as mobile phase is compatible with flame ionization detection, it is possible to analyse many solutes at trace levels. Supercritical carbon dioxide offers higher solute diffusivity compared with the inert carrier gas conventionally used in gas chromatography and has a lower viscosity than the liquid solvents used in HPLC. Thus, glycerides of fatty acids can be separated and eluted at a lower temperature and with shorter analysis time in SFC. In this study the analysis of mono-, di- and triglyceride mixtures in several pharmaceutical excipients is reported using capillary SFC. Quantitative analysis is possible on the basis of a response factor established for each analyte. The accuracy of the method and its advantages are demonstrated.

Caprylates↗

Direct quantification of protein partitioning in oil-in-water emulsion by front-face fluorescence: avoiding the need for centrifugation.

The quantification of proteins adsorbed at the oil-in-water interface is often difficult since it requires separation of fat globules from the aqueous phase that may damage the fat globule size and/or modify the interfacial composition. Front-face fluorescence spectroscopy was used to characterize the protein partitioning between the aqueous and oil phases of emulsions without separating these two phases. Different emulsions based on skim milk powder (SMP), two mono- and di-glyceride (MDG) mixtures (saturated and partially unsaturated), and three fats (hydrogenated and refined coconut oils and refined palm oil) were studied. The impact of an ageing period (24 h at 4 degrees C) was also investigated to typify the first step of ice cream processing. The emulsions were characterized for protein partitioning, immediately following emulsification and after ageing, using the Bradford spectrophotometric method, applied to the aqueous phase recovered after emulsion centrifugation. In parallel, the emulsions were characterized by their tryptophan emission fluorescence spectra. The area of the peaks at 333 nm, of the fourth-derivative fluorescence spectra corresponding to the amount of proteins present in the aqueous phase of emulsions, was well correlated with the Bradford measurements (r2=0.91). This amount was also calculated from the fluorescence calibration curve obtained with SMP in solution. In conclusion, front-face fluorescence spectroscopy appeared to be a powerful and simple technique allowing the quantification of different populations of protein in an emulsified system, i.e., in the aqueous phase and loaded at the fat globule interface.

Centrifugation↗

Enhancing mechanism of Labrasol on intestinal membrane permeability of the hydrophilic drug gentamicin sulfate.

The aim of this study was to clarify the mechanism by which caprylocaproyl macrogol-8 glyceride (Labrasol) enhances the intestinal absorption of gentamicin sulfate (GM), a drug that has poor permeability but relatively high solubility. We studied the following characteristics: (i) the phase behavior of Labrasol in aqueous solution, (ii) the affinity of GM to Labrasol micelles, and (iii) the interaction between Labrasol and membrane lipids. We measured the critical micelle concentration of Labrasol in aqueous solution to be approximately 0.01%. The average diameters of Labrasol micelles in 2% and 25% solutions were approximately 10 nm and 20 nm, respectively, indicating that micelles increase in size with increasing Labrasol concentration. Although GM dissolved in 5% Labrasol solution was dialyzable, GM dissolved in either 25% or 50% Labrasol solutions was not, suggesting that GM exists in the hydrophilic region of the Labrasol micelle or in the high affinity region of the micelle surface where GM is retained. In membrane permeability experiments and electrophysiological studies conducted with rat ileum, only 25% Labrasol solution enhanced GM permeability, but did not remarkably affect membrane resistance. Furthermore, Labrasol increased membrane lipid fluidity as determined by fluorescence anisotropy in porcine intestinal brush border membrane liposomes. These results suggest that high concentrations of Labrasol solution enhance membrane permeability of GM via a transcellular rather than a paracellular route. We propose the following mechanism: Labrasol micelles grow when the concentration of Labrasol exceeds 20%, at which point GM shows high affinity for the hydrophilic region of the micelles. Since Labrasol micelles increase membrane lipid fluidity, the membrane permeability of GM is concomitantly enhanced.

Absorption↗

Preparation and characterization of a self-emulsifying pellet formulation.

The purpose of the current study is to investigate the feasibility of producing solid self-emulsifying pellets using the extrusion/spheronization technique. Pellets were made from a mixture of C18 partial glycerides, Solutol HS15 and microcrystalline cellulose. Pellets with good physical properties (size, shape, friability) and self-emulsifying properties were produced. The pellets were, in contrast to pellets lacking Solutol, able to transfer a lipophilic dye and a spin probe into the aqueous media. The release kinetics and the microenvironment of the pellets during the release process were assessed using electron spin resonance (ESR) spectroscopy. The ESR results showed that the hydrophobic spin probe was localized mainly in the lipid environment all over the release time. Furthermore, the formulation was capable of accelerating the release of the drug diazepam and achieving a diazepam concentration above its saturation solubility. In conclusion, spherical pellets with low friability and self-emulsifying properties can be produced by the standard extrusion/spheronization technique. The pellets are capable of transfering lipophilic compounds into the aqueous phase and have a high potential to increase the bioavailability of lipophilic drugs.

Azo Compounds↗

The preparation of pellets containing a surfactant or a mixture of mono- and di-gylcerides by extrusion/spheronization.

The ability to incorporate either of the two components of a self-emulsifying system (polysorbate 80 (PSG80) and a mixture of mono- and di-glycerides (MDG)) separately into pellets prepared by extrusion/spheronization and the properties of any resulting pellets has been investigated. The results have established that it is possible to prepare satisfactory pellets thus adding to the range of ingredients that can be included in pellet formulations. For PS80, it was found possible to prepare pellets containing at least 92% of the surfactant/water mixture, but with a mixture of (MDG) and water, however, it was not possible to prepare pellets, which contained more than 46% of MDG. By analysis of variance significant relationships were established between the ratio of lactose to MCC and the ratio of the PS80 or MDG to water and the properties of the pellets. There were both similarities and differences of the two input factors, which provided relationships for the two materials. The quantity of liquid required, the fluid content of the pellets, the tensile strength and porosity of the pellets provided relationships for both systems, whereas the extrusion force and the median pellet size gave relationships for the PS80 systems, but they did not for the MDG systems. The opposite was true for interquartile range (IQR), the yield in the modal fraction and the shape factor. It was possible to identify global relationships for these systems and those reported previously, when the two components were combined as a self-emulsifying system, by the application of perceptual mapping. It was found that, there were relationships between the size range, expressed as the IQR and the fluid level required to make pellets; the quantity of the non-aqueous component of the fluid and the pellet shape factor; the extrusion force and the tensile strength of the pellets and the yield in the modal fraction and the ratio of the non-aqueous to aqueous content of the fluid used to prepare the pellets. The ability to use the materials independently offers further alternatives for the formulation of water insoluble drugs into pellet formulations.

Biopharmaceutics↗