Lipids separation

Specific Chemical Detection Methods Used for Various Lipids Separated by TLC... 

Silica gel G contains a binder, calcium sulphate, to help it to adhere to the plate, but silica gel H does not and is preferable for some lipid separations, particularly polar mixtures. Some commercially prepared plates contain an alternative organic binder that does not interfere in the same way as calcium sulphate but can present some difficulty with location methods, particularly charring. The degree of hydration of the adsorbent and the particle size will effect the separation and because these cannot be guaranteed, authentic standards should always be run at the same time as the samples. 

Concentration and Identification of Flavor Volatiles in Heated Beef Fat fay SC-CO2 Extraction. Um et al. (30) studied the flavor intensities of lipids separated in different fractions of SC-CO2 extracts at two pressures from heated beef tallow. The tallow was heated at 100°C for 2 hr and extracted at 207 bar/50 C and 345 bar/50°C. Six 1 kg fractions of CO2 were used to extract 100 g of tallow at each pressure and separated at 34.5 bar/40°C. 

Ackman, R.G., 1991. Application of gas-liquid chromatography to lipid separation and analysis. In Analysis of Fats, Oils and Lipoproteins (E.G. Perkins, ed.) pp. 270-300. American Oil Chemists Society, Champaign, 111. 

The analysis of organohalogen substances (OHS) in biological samples includes extraction of the OHS and the lipids from the matrix, removal of the lipids, separation of various classes of OHS and finally, detection and quantification. As pointed out above, PCNs have physical and chemical properties similar to the PCBs and the methods used to analyze PCBs and PCNs are therefore rather similar. 

In argentation thin-layer chromatography, a major technique for lipid separations, application is made of the ability of silver to form complexes with lipids. Silver nitrate is incorporated in either the stationary or mobile phase, and separations are based on the type (such as cis-trans) and extent of unsaturation in the lipids. This type of chromatography has been reviewed by Morris. ... 

Capillary and packed (GC) columns are of value in the analysis of complex mixtures of lipids. The best capillary column length will depend on the complexity of the material injected, however, 30-m columns are often employed. In packed columns, many types of stationary phases are available for lipid separation, and these include silicone and alkylated or cyanogenated derivatives, polyesters, polyglycol, and carboranes. It is also common to derivatize the fatty-acid side chains to the corresponding methyl esters by reaction in BF3/ methanol prior to chromatographic analysis to achieve more distinct and uniform separations. 

Ackman, R.G. Application of Thin-Layer Chromatography to Lipid Separation. In Analysis of Fats, Oils and Lipoproteins Perkins, G.E., Ed. Amer. Oil Chem. Soc. Champaign, IL, 1991 60-82. 

Touchstone, J.C., Thin-layer chromatographic procedures for lipid separation, J Chromatogr B, 671 (1995) 169-195. 

The permeability barrier formed by lipids separating one aqueous environment from another is the defining characteristic of a biological membrane. This barrier enables... 

Clearly, the polyester stationary phases advocated earlier (79) in the field of lipid separations possess only a limited thermal stability. Cyanopropyl silicones [356,357] now provide selective and more stable stationary phases for the separation of FA esters. However, the column selectivity alone falls short of some resolution requirement, while capillary GC seems to be the most profitable route to pursue. The state-of-the-art separation of FA methyl esters is reflected in Fig. 3.21 [358], where many of the minor structural differences in the chromatographed FA molecules yield readily distinguishable peaks. Acquisition of suitable standards as well as extensive correlations of FA molecular parameters with retention characteristics [359] are now clearly needed. 

Great advances have been made in the direct analysis of lipid extracts (i.e., those requiring minimal preparative fractionation) by the use of dedicated analytical instrumentation such as TLC-FID and HPLC. Mass spectrometry enables the combination of aspects of lipid separation by molecular weight and structural detail from techniques of ionization-induced fragmentation. Combined with chromatographic procedures, this represents the most powerful but expensive tool available for lipid analysis. 

Antonopoulou, S., C.A. Demopoulos, and N.K. Andrikopoulos. 1996. Lipid separation from Urtica dioica Existence of plateletactivating factor. /. Agric. Food Chem. 44 3052-3056. 

The increasing use of triple-quadrupole and ion-trap mass spectrometers has significantly enhanced the ability to perform structural identification of multiple species in a single analysis. An increasing number of reports have made use of direct injection of lipid mixtures with no prior chromatographic lipid separation. 

The typical profile of polar and neutral lipids separation of the fish species on preparative thin layer chromatography is shown in Figure 4. 

In the lamellar phases, bilayers of lipid molecules alternate with aqueous layers to form a one-dimensional periodic stack. Many of the two-dimensional phases also consist of bilayer arrangements of lipids separated by water layers. In these phases, the bilayers exhibit periodic wave-like undulations with periods of 15-40 nm (150- 00 A) (Tardieu et aL, 1973). The hexagonal phases are made up of long cylinders of lipids packed in a two-dimensional hexagonal lattice with interstitial water or, alternatively, of long cylinders of water surrounded by interstitial lipids. 

The field of fatty acid separation was revolutionized by the development of GC, which allowed nearly complete separation of the common fatty acids. Later workers developed methods for GC separation of steroids, and of the steryl and glyceryl esters by high-temperature GC. The introduction of the flame ionization detector (FID) transformed GC from a method of qualitative separation into a method of quantification, which has remained unsurpassed to the present day. In parallel, TLC was developed as both a preparative and analytical technique for the resolution of the nonvolatile lipids. Because of the nearly universal applicability to all lipid analyses and the use of inexpensive apparatus, TLC made lipid separations accessible to all laboratories and it has remained unsurpassed as a method of sample preparation for modern analytical methods of lipid analyses. 

Polar lipids separated by TLC are commonly quantified by densitometry or the lipids may be scraped off and quantified by colorimetry or gas chromatography after derivatization. The latroscan TLC-flame ionization detector system is well suited to the quantification of polar lipids. In this procedure, TLC is performed on quartz rods coated with silica. The rods are then passed through a flame ionization detector to quantify each lipid band in turn. 

Touchstone (1995), Fried and Sherma (1996) and Shukla (1995). Several lipid separations have been standardized, which makes it easier to achieve effective separations even by a person not very familiar to this field however, other separations such as for alkaloids, flavones, medicine and other natural components are still being developed, and TLC of these compounds requires the operator to have more expertise and a greater understanding of the basic concept of TLC separation and analysis. Sherma (1994, 1996) has reviewed published articles on TLC every alternate year in the journal Analytical Chemistry. 

There are several stationary phases that can be used for separation of various compounds including lipids, natural products, biological compounds, drugs, etc. Examples are Celite (Supelco Inc., PA), cellulose powder, ion-exchange cellulose, starch, polyamides and Sephadex (Supelco Inc., PA), but the most popular ones for lipid separations include silica gel, alumina and kieselguhr. These adsorbents can also be modified by impregnation with other substances so as to achieve the desired separations. Based on their characteristics these phases can be classified as normal or reversed. 

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