Argentation TLC

Strocchi and Holman (1971), with the aid of argentation TLC and GLC-mass spectrometry, identified the fatty acids in Tables 4.7 and 4.8. We have presented all of their data because the identifications were obtained by unequivocal methods, many previously tentative identifications were confirmed, and the results were quantitative. Strocchi and Holman did not identify the positional isomers of the unsaturates but found two or three peaks for most of the carbon numbers. Iverson (1983) determined the quantities of minor and trace fatty acids, verifying the findings of other investigators. 

Smith et al. (1978) have described a procedure for the GLC determination of cis and trans isomers of unsaturated fatty acids in butter after fractionation of the saturated, monoenoic, dienoic, and polyenoic fatty acid methyl esters by argentation TLC. Total trans acids were much higher, as measured by infrared spectrophotometry than by GLC, probably because some of the acids could have two or more of the trans bonds designated as isolated by infrared spectrophotometry. Enzymatic evaluation of methylene-interrupted cis, cis double bonds by lipoxidase resulted in lower values than those obtained by GLC. The authors mention that the lipoxidase method is difficult, requiring considerable skill, and suggest that their method is suitable for the determination of the principal fatty acids in complex food lipids such as bovine milk fat. 

Reaction of shea triterpene alcohols with acetic anhydride and sulphuric acid to produce coloured products (Fitelson s reaction) was the basis of a sensitive early test for the presence of shea butter in cocoa butter (Fincke, 1975). Analysis of the triterpene fraction of a commercial cocoa butter by TLC fractionation followed by GC (Fincke, 1976), or argentation TLC followed by GC (Gegiou and Staphylakis, 1985), have been shown to have potential for detecting CBEs in chocolate based on the difference in levels of P-amyrin, butyrospermol and... 

The quantitative determination of individual isomers of tram-18 1 fatty acids in milk fat is not straightforward. It involves a multi-stage analytical procedure (i.e., transesterification of milk fat, argentation TLC of the fatty acid esters to separate the civ-isomers and tram-isomers, followed by capillary GC). This method gives an almost complete separation of the 13 individual tram-18 1 isomers, from A4 to A16 (Precht and Molkentin, 1996). 

Monocarboxylic esters were further separated by argentation TLC [silica GF254 impregnated with 5% (w/v) silver nitrate] into saturated esters (Rf = 0.51), monounsaturated esters (R = 0.42), diunsaturated esters (Rf = 0.29), and poly unsaturated esters (Rf < 0.29) by development in n-heptane-diethyl ether-methanol (90 10 1, v/v/v) (19). 

Quantitative determination of AMEs was achieved by high-resolution GLC analyses on a glass, support-coated, open, tubular (SCOT) SE-30 column (57 m x 0.5 mm i.d., S.G.E., Australia) in a Perkin-Elmer 910 gas chromatograph equipped with a flame ionization detector. Individual compounds were identified on the basis of GLC retention times, coinjection with authentic standards (Analabs), and their performance on argentation TLC. 

Petroleum ether (30-60 deg C) Counter current chromatogFaphy in hexane/nitromethane Preparative TLC on Silica gel argentation TLC IR,GC-MS. GC (27,47)... 

Porto et al. methylated a Clusia floral resin extract, chromatographed the derived compounds over silica gel, rechromatographed using preparative argentation TLC (5% silver nitrate), and thereby isolated seven polyisoprenylated benzophenones [68], Lokvam et al. [74] and de Oliveira et al. [59] also methylated a crude extract before isolating chamones I, II (119, 116), and nemorosone 11 (89) from Clusia species. 

It is also possible to modify the adsorption properties of the silica gel by incorporating substances such as bases or buffers enabling coatings with accurately defined pH to be prepared. In a like manner silver nitrate can be added this admixture changes the adsorptive properties to permit increased discrimination and separation of unsaturated compounds, especially alkenes. The technique is commonly known as Argentation TLC. 

Unsaturated fatty acids are usually separated by argentation-TLC (Section 4.4) prior to analysis by GLC. This separates acids on the basis of their total number of ethylenic bonds, and in addition, distinguishes trans from cis isomers (Emken and Dutton, 1974) (see Section 4.4). 

TLC is commonly used for the separation of different classes of wax components or for analysis of monomers from cutin and suberin depolymerization. A typical separation is shown in Fig. 6.12. By such methods, it is possible to separate hydrocarbons, wax esters, primary alcohols, secondary alcohols and /8-diketones from plant waxes (von Wettstein-Knowles, 1979). Products of hydrogenolysis from cutin can be separated by TLC into alkan-l-ols, alkane-a,ft>-diols, Cis triols, Ci6 triols and Cis tetrols (Kolattukudy, 1980). Unsaturated components can be resolved by argentation-TLC (Tulloch, 1976) and threo or erythro diastereoisomers separated by boric acid/silica gel TLC (Eglinton and Hunneman, 1968). Straight-chain compounds can be preferentially removed from branched compounds as their urea complexes (Kolattukudy, 1980). 

Silver ion TLC has also been used for separation of a variety of substituted unsaturated fatty acids such as epoxy, hydroxy and halohydroxy fatty acids, and these have been reviewed by Morris and Nichols (1972). Wax esters and steryl esters exhibit similar polarities and do not separate on column chromatography or normal silica TLC. Kiosseoglou and Boskou (1990) have separated the wax esters from steryl esters by using argentation TLC. They have used silica gel plates impregnated with 10% silver nitrate, with developing solvent hexane/chloroform (7 3 vol./vol.). 

Argentation TLC (on silver nitrate-impregnated silica gel plates) and hexane-ethyl acetate-diisopropyl ether (2 2 1, v/v/v) as mobile phase were used to separate vitamins Ki, K2, K3, ubiquinone-6, ubiquinone-9, and ubiquinone-10 (Rp values 0.71, 0.63, 0.57, 0.42, 0.21, and 0.28, respectively). ° ... 

Table 4 / p values of selected lipophilic vitamins, provitamins, and related compounds separated by argentation TLC. 

Most of the prenyUipids, such as chlorophylls, carotenoids, and prenylquinones, as well as tocopherols and vitamin Ki, which occur in plant hpid extracts, can be separated by TLC using sUica gel plates or special mixtures of silica gel with other adsorbents. But the compounds with one double bond per isoprene and others with a partially or fiiUy unsaturated isoprenoid chain can be separated efficiently by argentation TLC. The Rp values of selected hpophUic vitamins, their provitamins, and related compounds separated by argentation TLC using different mobile phases are listed in Table... 

The first publications on the separation of lipids on silvernitrate-impregnated adsorbent columns [207] and layers [6, 136, 208] appeared in 1962/63. Argentation-TLC quickly gained favour and has become one of the indispensable aids in the chemistry of lipophilic natural products. 

Adsorption TLC can be combined with argentation-TLC or reversed phase partition TLC on a single layer, using two-dimensional technique. The two last named procedures can be combined also. 

It has been possible to isolate several hitherto unknown fatty acids (mostly as their methyl esters) with the help of argentation-TLC. The relevant work has been summed up in a recent publication [143]. 

Reference may be made here to a procedure for separating mixtures of mono-olefinic fatty acids and for subsequently determining the structure of the components [11]. This method utilises two-dimensional TLC on silica gel, impregnated with paraffin and silver nitrate a similar procedure is described in [225]. A scheme for determination of the position and configuration of the double bonds in polyolefinic acids [170] depends on partial hydrogenation with hydrazine, separation of the reduction products through argentation-TLC and determination of... 

Table 71. Solvents for fractionation of pure compound classes using reversed phase partition TLG and argentation-TLC [123]...

Compound class Adsorp- tion TLC hRf Solvents for argentation-TLC (5% AgNOa in silica gel G) Solvents for partition TLC in reversed phase (paraffin oil on kieselguhr G)... 

Using argentation TLC, oleodistearin has been isolated from lard and stearodiolein from palm oil, malabar tallow and cocoa butter and it has been shown that these asymmetrical triglycerides are optically active [139]. 

Classes of triglycerides which cannot be separated by argentation-TLC are fractionated by reversed phase partition TLC [81, 89, 90, 124] (see Fig. 149, p. 143) or by counter current distribution [124]. 

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