Method for isolation of non-esterified fatty acids and several other classes of plasma lipids by column chromatography on silica gel

Ingalls, Stephen T.; Kriaris, Maria S.; Yang, Xu; DeWulf, David W.; Tserng, Kou‐Yi; Hoppel, Charles L.

doi:10.1016/0378-4347(93)80441-6

Cited by 50 publications

(32 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…7 shows that there was an extensive remodeling of the triglyceride-bound fatty acids in both groups. Namely, the relative amount of saturated fatty acids decreased (e.g., diet I: 16 …”

Section: Resultsmentioning

confidence: 99%

Influence of diet on the modeling of adipose tissue triglycerides during growth

Brunengraber¹,

McCabe²,

Kasumov³

et al. 2003

American Journal of Physiology-Endocrinology and Metabolism

108

View full text Add to dashboard Cite

We have studied the accretion of lipids in growing mice. We measured the rates of synthesis and degradation of triglycerides in epididymal fat pads of mice maintained for 44 days on a low-fat, high-carbohydrate diet (I) or a high-fat, lowcarbohydrate diet (II).2 H2O was added to the drinking water for 14 days. Rates of incorporation/washout of 2 H to/from C1 of triglyceride-glycerol showed that triglyceride synthesis was greater than triglyceride degradation (net triglyceride balance was ϳ2.5 times greater in II than in I). The data also show that the contribution of de novo lipogenesis to triglyceride-bound palmitate was ϳ3 times greater in I than in II. This was consistent with a greater relative intake of carbohydrate in I vs. II. The rates of incorporation and washout of newly synthesized ( 2 H-labeled) palmitate into and from triglycerides were also measured. Those data suggested a remodeling of triglyceridebound fatty acids. On measuring the profile of triglyceridebound fatty acids, we observed a decrease in the relative abundance of triglyceride-bound palmitate and stearate and an increase in triglyceride-bound oleate and linoleate. This was observed in I and II. In summary, diet substantially affects the deposition and modeling of triglycerides in adipose tissue during growth. 2 H2O can be used to examine the mechanisms responsible for the accumulation of triglycerides, e.g., factors that affect 1) triglyceride synthesis and degradation and 2) the source of fatty acids that are used in esterification. deuterium; stable isotopes; lipid metabolism; pediatric obesity; insulin resistance THERE IS AN ALARMING INCREASE in the number of reported cases of pediatric obesity (26, 32). Like obese adults, obese children present with metabolic abnormalities (e.g., impaired glucose tolerance, dyslipidemia) that are associated with an increased risk of developing a chronic disease (e.g., diabetes, cardiovascular disease) (8,9,21,24,32). A special concern regarding obese children is that many will remain obese as adults, which further increases the likelihood of developing a chronic disease. Presumably, the development of effective treatments for pediatric obesity will benefit from knowledge of how specific biochemical reactions drive lipid accumulation during growth.We hypothesized that, during growth, the nature of one's caloric intake will affect the rate of, and the mechanism of, fat accumulation. For example, as the ratio of dietary fat to carbohydrate increases, the rate of triglyceride accretion will increase. Also, the relative amount of carbohydrate in the diet will affect the relative contribution of de novo lipogenesis to the pool of triglyceride-bound fatty acids. We developed the use of 2 H 2 O to measure the rates of the reactions involved in triglyceride accumulation (e.g., triglyceride synthesis and triglyceride degradation and the contribution of de novo lipogenesis to the pool of triglyceride-bound fatty acids). This approach also allows us to determine the rate of remodeling of fatty acids that are bo...

show abstract

“…7 shows that there was an extensive remodeling of the triglyceride-bound fatty acids in both groups. Namely, the relative amount of saturated fatty acids decreased (e.g., diet I: 16 …”

Section: Resultsmentioning

confidence: 99%

Influence of diet on the modeling of adipose tissue triglycerides during growth

Brunengraber¹,

McCabe²,

Kasumov³

et al. 2003

American Journal of Physiology-Endocrinology and Metabolism

108

View full text Add to dashboard Cite

show abstract

“…Samples were then subjected to silica gel chromatography and the lipid classes were serially eluted in six fractions using solvents of increasing polarity ( 19 ): Fraction 1 contained cholesterol esters and was eluted with isooctane:ethylacetate (95:1), fraction 2 contained triglycerides and was eluted with isooctane:ethylacetate (20:1), fraction 3 contained cholesterol and diglycerides and was eluted with isooctane:ethylacetate (75:25), fraction 4 contained free fatty acids and was eluted with isooctane:ethylacetate:acetic acid (75:25:2), fraction 5 contained monoglycerides and was eluted with isooctane:ethylacetate:acetic acid (75:25:2), and fraction 6 contained the combined phospholipids and was eluted with methanol. This fi nal fraction, containing cardiolipin, was evaporated to dryness while the other factions were stored for other experiments.…”

Section: Biological Samplesmentioning

confidence: 99%

Separation and characterization of cardiolipin molecular species by reverse-phase ion pair high-performance liquid chromatography-mass spectrometry

Minkler¹,

Hoppel

2010

Journal of Lipid Research

Self Cite

View full text Add to dashboard Cite

This article is available online at http://www.jlr.orgCardiolipin [bis-(1,2-diacyl-sn -glycero-3-phospho)-1' -3 ′ -snglycerol] is a unique phospholipid present in mitochondria ( 1 ) and is a component of bacterial membranes ( 2 ). In eukaryotic cells, it is present almost exclusively in the inner mitochondrial membrane ( 3 ), where it is required for normal mitochondrial function. In bacteria, it has been identifi ed in the cytoplasmic membrane ( 4 ). Structurally, the diversity of cardiolipin molecular species is found in both the identity and position of its four fatty acyl moieties; the characterization of these differences and the relative amounts of these different species has been the focus of much investigation ( 5-9 ).We demonstrated the feasibility of characterizing cardiolipin molecular species by reverse-phase HPLC-MS/MS using a quadrupole ion trap mass spectrometer and recursive mass spectrometry (MS, MS/MS, and MS 3 ) ( 10-12 ). Following analysis of many samples, we found two areas in this procedure that we wished to improve: 1 ) the chromatographic conditions yielded variable retention time results with the resolution among the cardiolipin molecular species sometimes not suffi cient to distinguish the molecular species, and 2 ) the key fatty acid MS/MS mass fragment for cardiolipin (e.g., 279 m/z in tetra-linoleoyl cardiolipin) was outside of the range of the ion-trap mass spectrometer because of the instrument's low mass cut-off. This fragment was observable in an MS 3 spectrum, but it was less prominent than desirable.Abstract An improved high-performance liquid chromatography-mass spectrometry method for the separation and characterization of cardiolipin molecular species is presented. Reverse-phase ion pair chromatography with acidifi ed triethylamine resulted in increased chromatographic retention and resolution when compared with chromatography without acidifi ed triethylamine. Using a hybrid triple quadrupole linear ion trap mass spectrometer to generate MS/MS spectra revealed three regions within each spectrum that could be used to deduce the structure of the cardiolipin molecular species: the diacylglycerol phosphate region, the monoacylglycerol phosphate region, and the fatty acid region. Cardiolipin standards of known composition were analyzed and exhibited expected chromatographic and mass spectral results. Two minor components in commercial bovine heart cardiolipin, (with the same molecular weight but different chromatographic retention times), were shown to differ by fatty acid composition: (C18:2) 2 (C18:1) 2 versus (C18:2) 3 (C18:0) 1 . These compounds were then analyzed by HPLC-MS 3 to examine specifi c diacylglycerol phosphate generated fatty acid fragmentation. Also, two commercial sources of bovine heart cardiolipin were shown to have minor differences in cardiolipin species content. Cardiolipin isolated from rat liver, mouse heart, and dog heart mitochondria were then characterized and the relative distributions of the major cardiolipin species were determined. -Minkler, P. E....

show abstract

“…The lower organic phase was removed to a clean tube and dried under N 2 . Glyceryl lipids were separated following the method described by Ingalls et al [28]. A silica SPE cartridge (DSC-Si, 100 mg) was washed with 4 mL isooctane/ethyl acetate (80:1).…”

Section: Extraction and Isolation Of Neutral Lipidsmentioning

confidence: 99%

Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry

Murphy

James

McAnoy

et al. 2007

Analytical Biochemistry

142

122

View full text Add to dashboard Cite

Triacylglycerols are neutral lipids present in all mammalian cells as energy reserves and diacylglycerols as intermediates in phospholipid biosynthesis and as signaling molecules. The molecular species of triacylglycerols and diacylglycerols present in mammalian cells are quite complex and previous investigations revealed multiple isobaric species having molecular weights at virtually every even mass between 600-900 daltons, making it difficult to assess changes of individual molecular species after cell activation. A method has been developed using tandem mass spectrometry and neutral loss scanning to quantitatively analyze changes in those glyceryl ester molecular species containing identical fatty acyl groups. This was carried out by neutral loss scanning of 18 common fatty acyl groups where the neutral loss corresponded to the free carboxylic acid plus NH 3 . Deuterium labeled internal standards were used to normalize the signal for each nominal [M +NH 4 ] + ion undergoing this neutral loss reaction. This method was applied in studies of triacylglycerols in RAW 264.7 cells treated with the toll-like receptor 4 ligand Kdo 2 -lipid A. A 50:1-TAG containing 18:1 was found to increase significantly over a 24 hr time course after Kdo 2 -lipid A exposure whereas an isobaric 50:1-TAG containing 16:1 did not change relative to controls.

show abstract

Method for isolation of non-esterified fatty acids and several other classes of plasma lipids by column chromatography on silica gel

Cited by 50 publications

References 37 publications

Influence of diet on the modeling of adipose tissue triglycerides during growth

Influence of diet on the modeling of adipose tissue triglycerides during growth

Separation and characterization of cardiolipin molecular species by reverse-phase ion pair high-performance liquid chromatography-mass spectrometry

Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry

Contact Info

Product

Resources

About