Diacylglycerol Biosynthesis in Everted Sacs of Rat Intestinal Mucosa

1975 ◽  
Vol 53 (11) ◽  
pp. 1170-1183 ◽  
Author(s):  
W. C. Breckenridge ◽  
A. Kuksis
Keyword(s):  
Fatty Acids ◽  
Phosphatidic Acid ◽  
Thin Layer ◽  
Free Fatty Acids ◽  
Thin Layer Chromatography ◽  
Intestinal Mucosa ◽  
Gas Liquid Chromatography ◽  
Acid Pathway ◽  
Layer Chromatography ◽  
Everted Sacs

The molecular specificity in the biosynthesis of diacylglycerols by rat intestinal mucosa was examined by means of radioactive markers, thin-layer chromatography with silver nitrate and gas-liquid chromatography with radioactivity monitoring. Bile salt micelles of alternately labeled monoacylglycerols and free fatty acids were incubated with everted sacs of intestinal mucosa for various periods of time and the diacylglycerols were isolated by solvent extraction and thin-layer chromatography. Stereospecific analyses of the X-1,2-diacylglycerols labeled from 2-monoacylgiycerols showed that the sn-1,2-isomers (45–55%) were slightly in excess of the sn-2,3-isomers (34–45%) with the X-1,3-diacylglycerols accounting for the rest of the radioactivity (5–10%). This suggests that racemic diacylglycerols may be intermediates in the resynthesis of dietary fat in rat intestinal mucosa. Detailed analyses of the molecular species of the sn-1,2-diacylglycerols labeled from free fatty acids revealed that 10–45% of the total did not contain the acid present in the 2-monoacylglycerol supplied, and therefore had originated from the phosphatidic acid pathway. These findings are at variance with those obtained in isolated microsomes, which have suggested an inhibition of the phosphatidic acid pathway by monoacylglycerols as well as have given evidence of an exclusive synthesis of sn-1,2-diacylglycerols from 2-monoacylglycerols.

Triacylglycerol Biosynthesis in Everted Sacs of Rat Intestinal Mucosa

1975 ◽  
Vol 53 (11) ◽  
pp. 1184-1195 ◽  
Author(s):  
W. C. Breckenridge ◽  
A. Kuksis
Keyword(s):  
Fatty Acids ◽  
Thin Layer ◽  
Free Fatty Acids ◽  
Thin Layer Chromatography ◽  
Intestinal Mucosa ◽  
Molecular Species ◽  
Triacylglycerol Biosynthesis ◽  
Acid Pathway ◽  
Layer Chromatography ◽  
Everted Sacs

The molecular specificity of the biosynthesis of triacylglycerols by rat intestinal mucosa was examined by means of radioactive and mass tracers, and thin-layer chromatography with silver nitrate and gas–liquid chromatography with radioactivity monitoring. Bile salt micelles of alternately labeled monoacylglycerols and free fatty acids were incubated with everted sacs of intestinal mucosa for various periods of time and the triacylglycerols isolated by solvent extraction and thin-layer chromatography. Analyses of the molecular species of the triacylglycerols labeled from monoacylglycerols showed that the 2-monoacylglycerol pathway was responsible for the biosynthesis of a maximum of 90% and the X-1-monoacylglycerol pathway for about 10% of the total radioactive triacylglycerols. Detailed analyses of the molecular species of triacylglycerols labeled from free fatty acids showed that the phosphatide acid pathway contributed a minimum of 20–30% of the total labeled triacylglycerol formed. There was a preferential utilization in triacylglycerol biosynthesis of the more unsaturated diacylglycerols arising from the monoacylglycerol pathway and of the more saturated diacylglycerols originating from the phosphatidic acid pathway. The above experiments do not allow a demonstration of the utilization of the sn-2,3-diacylglycerols in triacylglycerol biosynthesis but are not inconsistent with it.

Stereospecific analysis of menhaden oil triacylglycerols and resolution of complex polyunsaturated diacylglycerols by gas–liquid chromatography on polar capillary columns

1990 ◽  
Vol 68 (1) ◽  
pp. 336-344 ◽  
Author(s):  
J. J. Myher ◽  
A. Kuksis ◽  
L.-Y. Yang
Keyword(s):  
Fatty Acids ◽  
Liquid Chromatography ◽  
Thin Layer ◽  
Phospholipase C ◽  
Thin Layer Chromatography ◽  
Molecular Species ◽  
Gas Liquid Chromatography ◽  
Menhaden Oil ◽  
Grignard Degradation ◽  
Layer Chromatography

The sn-1,2-, sn-2,3-, and X-1,3-diacylglycerols derived by Grignard degradation of purified menhaden oil triacylglycerols were isolated by conventional thin-layer chromatography with boric acid complexing. The sn-1,2(2,3)-diacylglycerols were resolved into sn-1,2- and sn-2,3-diacylglycerols by stepwise digestion with phospholipase C of the corresponding phosphatidylcholines and the positional distribution of the fatty acids were determined. Diacylglycerols were converted into trimethylsilyl ethers and resolved on the basis of molecular weight and degree of unsaturation by gas–liquid chromatography using a polar capillary column and isothermal or programmed temperatures. The order of chromatographic elution was established for more than 70 major and minor species by reference to primary and secondary diacylglycerol standards and by calculation of relative retention times. The identified molecular species ranged in carbon number from 28 to 44 and in double bond number from 0 to 12 being made up of C14–C22 fatty acids with 0 to 6 double bonds each and representing the n – 9, n – 6, n – 4, n – 3, and n – 1 series. The gas–liquid chromatographic determinations yielded proportions of all major species that were consistent with those calculated from the knowledge of the stereospecific distribution of the fatty acids in the original triacylglycerol molecules.Key words: Grignard degradation, rac-phosphatidylcholines, phospholipase C, enantiomeric diacylglycerols, thin-layer chromatography, molecular species of diacylglycerols, composition of fatty acids.

Biosynthesis of triacylglycerols by rat intestinal mucosa in vivo

1976 ◽  
Vol 54 (2) ◽  
pp. 145-152 ◽  
Author(s):  
W. C. Breckenridge ◽  
S. K. F. Yeung ◽  
A. Kuksis
Keyword(s):  
Fatty Acids ◽  
Phosphatidic Acid ◽  
Free Fatty Acids ◽  
Intestinal Mucosa ◽  
Stereospecific Analysis ◽  
Weight Fraction ◽  
Exact Estimate ◽  
Gas Liquid Chromatography ◽  
Butter Oil

The structure of mucosal triacylglycerols was studied in rat intestinal mucosa in vivo during the absorption of a low molecular weight fraction of butter oil and of the corresponding free fatty acids of medium and long chain length. The mucosal lipids were isolated by solvent extraction and the acylglycerol structures were determined by combined AgNO3 – thin-layer chromatography and gas–liquid chromatography techniques and stereospecific analysis. Evidence was obtained for a rapid biosynthesis of triacylglycerols from diacylglycerols arising from the operation of both the monoacylglycerol and the phosphatidic acid biosynthetic pathways. Both sn-1,2- and sn-2,3-diacylglycerols appeared to be converted to triacylglycerols at significant rates, but a preferential utilization of sn-1,2-diacylglycerols could not be excluded. Endogenous dilution varied from a minimum of 5% during triacylglycerol biosynthesis from monoacylglycerols to 15% during their synthesis from free fatty acids, and was characterized by a preferential placement of the endogenous acids in the sn-3 and 2 positions of the triacylglycerol molecules. Exogenous myristic acid was preferentially associated with the sn-3 position, and stearic acid became preferentially bound to the sn-1 position. The complexity of the triacylglycerol end products prevented an exact estimate of the contribution of the phosphatidic acid pathway, but the acylglycerol structures were compatible with a minimum of 20% of total triacylglycerol yield at all times.

Application of Thin-layer Chromatography to the Quantitation of Plasma Neutral Lipids and Free Fatty Acids

1967 ◽  
Vol 13 (9) ◽  
pp. 773-787 ◽  
Author(s):  
Robert T Louis-Ferdinand ◽  
Donald G Therriault ◽  
William F Blatt ◽  
Milton Mager ◽  
Edward J Metheson
Keyword(s):  
Fatty Acids ◽  
Sulfuric Acid ◽  
Thin Layer ◽  
Free Fatty Acids ◽  
Thin Layer Chromatography ◽  
Silica Gel ◽  
Neutral Lipids ◽  
Layer Chromatography

Abstract Neutral lipids and free fatty acids were extracted from plasma and separated on thin-layer chromatoplates coated with silica gel G. The plates were charred by heating after spraying with dichromate-sulfuric acid, and then evaluated densitometrically. Plasma analyses obtained by this procedure were compared with parallel determinations performed by established chemical technics.

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