Fractionation methods thin-layer

Standard methods of analysis of total sterol content of oils involve saponification of the oil, followed by extraction and isolation of total sterols from the unsaponihable fraction by thin layer chromatography (TLC) (AOCS, 1998). Quantification of individual sterols involves silylation of the sterol fraction and analysis by gas chromatography (GC). Sterols and steryl esters in oils and fats can be analysed by LC-GC after silylation or acylation of the free sterols (Artho et al., 1993). An alternative approach to the analysis of intact steryl esters involves separation of sterols and steryl esters by solid phase... 

Other separating techniques may be used to separate total hydrocarbons into different classes. Thus, the normal paraffins are selectively removed by 5 A molecular sieve (Mortimer and Luke, 1967) or by urea adduction, although it is less specific than the former method. Unsaturated hydrocarbons are separated from the saturated fraction by thin layer or column chromatography on silicic acid/AgNOj. 

While no analytical methods were located that are specific for detecting Stoddard solvent in sediment, as with water and soil, methods that detect other hydrocarbon mixtures may be applicable. For example, quantification of fuel oil hydrocarbons from sediments is a relatively involved process. Following extraction, the saturated and olefinic hydrocarbon fraction is separated from the aromatic hydrocarbon fraction using thin-layer chromatography or column chromatography. Fractions are subsequently analyzed by GLC (Gearing et al. 1980). 

Tissue Fatty Acid Composition. Tissue fatty acids were extracted using the method of Folch et al. (13). Aliquots of total tissue lipid extracts were separated into different phospholipid fractions by thin-layer chromatography using chlo-roform/methanol/water/triethylamine (4 5 1 4, by vol) as the developing system. The fatty acids in the phosphatidyl-... 

The total phosphoms content of the sample is determined by method AOCS Ja 5-55. Analysis of phosphoUpid in lecithin concentrates (AOCS Ja 7-86) is performed by fractionation with two-dimensional thin-layer chromatography (tic) followed by acid digestion and reaction with molybdate to measure total phosphorous for each fraction at 310 nm. It is a semiquantitative method for PC, PE, PI, PA, LPC, and LPE. Method AOCS Ja 7b-91 is for the direct deterrnination of single phosphoHpids PE, PA, PI, PC in lecithin by high performance Hquid chromatography (hplc). The method is appHcable to oil-containing lecithins, deoiled lecithins, lecithin fractions, but not appHcable to lyso-PC and lyso-PE. 

Spectrophotometric deterrnination at 550 nm is relatively insensitive and is useful for the deterrnination of vitamin B 2 in high potency products such as premixes. Thin-layer chromatography and open-column chromatography have been appHed to both the direct assay of cobalamins and to the fractionation and removal of interfering substances from sample extracts prior to microbiological or radioassay. Atomic absorption spectrophotometry of cobalt has been proposed for the deterrnination of vitamin B 2 in dry feeds. Chemical methods based on the estimation of cyanide or the presence of 5,6-dimethylben2irnida2ole in the vitamin B 2 molecule have not been widely used. 

SERS has also been applied as a sensitive, molecule-specific detection method in chromatography, e.g. thin layer, liquid, and gas chromatography. SERS-active colloids were deposited on the thin layer plates or mixed continuously with the liquid mobile phases. After adsorption of the analytes, characteristic spectra of the fractions were obtained and enabled unambiguous identification of very small amounts of substance. 

Nonionic surfactants, including EO-PO block copolymers, may be readily separated from anionic surfactants by a simple batch ion exchange method [21] analytical separation of EO-PO copolymers from other nonionic surfactants is possible by thin-layer chromatography (TLC) [22,23] and paper chromatography [24], and EO-PO copolymers may themselves be separated into narrow molecular weight fractions on a preparative scale by gel permeation chromatography (GPC) [25]. 

Because of the instability of many of the compounds involved, it is necessary to determine the chemical recoveries in all cases. This requires the use of macro quantities (10 mg up to several hundred mg) of carriers and target compounds. This, in turn, makes it impractical to use the various thin-layer methods, such as paper and thin-layer chromatography and paper electrophoresis, although such methods have proved useful in identifying products and in checking the purity of fractions. The separation methods now most commonly used are column chromatography and sublimation. 

The conventional approach to solvent extraction is the batch method. Early work with this method was hampered by the low concentration of the compounds present and the relative insensitivity of the methods of characterization. Thus lipids and hydrocarbons have been separated from seawater by extraction with petroleum ether and ethyl acetate. The fractionation techniques include column and thin-layer chromatography with final characterisation by thin-layer chromatography, infrared, and ultra-violet spectroscopy and gas chromatography. Of these techniques, only gas chromatography is really useful at levels of organic matter present in seawater. With techniques available today such as glass capillary gas chromatography and mass spectrometry, much more information could be extracted from such samples [20]. 

Multi-dimensional Chromatography. Multi-dimensional chromatography is the term used to describe a variety of methods where fractions from one chromatographic system are each transferred to another for further separation. Combinations of SEC with thin-layer chrcmatography have been shown to enable separation of copolymers by composition in a "cross-fractionation". OC utilizes a combination of two SECs in a cross-fractionation approach. 

In earlier times, thin-layer chromatography (TLC), polyamide chromatography, and paper electrophoresis were the major separation techniques for phenolics. Of these methods, TLC is still the workhorse of flavonoid analysis. It is used as a rapid, simple, and versatile method for following polyphenolics in plant extracts and in fractionation work. However, the majority of published work now refers to qualitative and quantitative applications of high-performance liquid chromatography (HPLC) for analysis. Llavonoids can be separated. 

The low-molecular-weight water-soluble fraction of LCP flour was found by thin layer chromatographic methods to contain several flavonoid components. To establish the role of flavonoids in the production of yellow color in biscuits, these components were extracted from LCP and glandless cottonseed flours with 85Z aqueous isopropyl alcohol (which is a better solvent for flavonoids than water). Before removal of the flavonoids, the flours had been treated with petroleum ether to extract residual lipids that could interfere with flavonoid isolation. Extraction of the residual lipids did not significantly alter the color of biscuits prepared with the extracted flours (Figure 7). 

Many methods exist to separate triglycerides into fractions on the basis of their degree of unsaturation. These include fractional crystallization from solvents, and separation by column and thin layer chromatography (TLC). The classes of triglycerides may then be studied and after methylation the fatty acid content determined by GLC. Yet even this is not the whole story since the position of fatty acids on the triglyceride molecule may uniquely affect the physical and biological properties of the lipid. 

The long story of the methods for the separation of the individual rare earths may broadly be divided into two main parts a) classical methods b) modern methods. Old-fashioned classical techniques like fractional crystallization, fractional precipitation and fractional thermal decomposition were not only used by the early workers in the past, but still remain as very important methods for economical production of rare earths on commercial scales. Modem methods like solvent (liquid-liquid) extraction, ion exchange or chromatographic (paper, thin layer and gas) techniques have both advantages and limitations. 

Figure 1 shows the scheme for the preparation of purified lipid A from endotoxin. S. typhimurium G30/C21 was extracted by the method of Galanos t aK (24) and submitted to one of two different conditions of hydrolysis (a) 0.1 N HC1 [in methanol-water (1 1, v/v)], 100 °C, 45 min, to yield the crude monophosphoryl lipid A (nontoxic), and (b) 0.02 M sodium acetate, pH 4.5, 100 °C for 30 min (two cycles) to yield the crude diphosphoryl lipid A (toxic). The 0.1 N HC1 hydrolysis product was fractionated on a Sephadex LH-20 column (23). Each of these fractions was then separated by preparative thin layer chromatography (TLC) on silica gel H (500 ym), with the solvent system chloroform-methanol-waterconcentrated ammonium hydroxide (50 25 4 2, v/v) as previously described (23) to yield TLC fractions 1-7 and 1-9 respectively. 

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