[CITATION][C] Quantitative analysis of phospholipids by thin-layer chromatography and phosphorus analysis of spots

G Rouser, AN Siakotos, S Fleischer - Lipids, 1966 - Springer
G Rouser, AN Siakotos, S Fleischer
Lipids, 1966Springer
p ROCEDURES FOR ANALYSIS of phospholipid composition by thin-layer chromatography
(TLC) and phospho~ nts analysis have been reported from a number of laboratories. These
procedures usually depend upon one-dimensional TLC and elution of spots before analysis.
The method reported here has the advantage of improved separations by two-dimensionM
TLC, direct aspiration of spots by suction, and phosphorus analysis without prior elution. Our
procedure depends upon two-dimensional TLC with the solvent pairs 1) …
p ROCEDURES FOR ANALYSIS of phospholipid composition by thin-layer chromatography (TLC) and phospho~ nts analysis have been reported from a number of laboratories. These procedures usually depend upon one-dimensional TLC and elution of spots before analysis. The method reported here has the advantage of improved separations by two-dimensionM TLC, direct aspiration of spots by suction, and phosphorus analysis without prior elution. Our procedure depends upon two-dimensional TLC with the solvent pairs 1) chloroform/methanol/water 65/25/4 a~ ld n-butanol/acetic acid/water 60/20/20; and 2) chloroform/methanol/2S% aqueous ammonia 65/35/5 followed by chloroform/acetone/methanol/acetic acid/water 5/271/1/0.5. The adsorbent composed of silica gel plain/magnesium silicate 9/1 (1) after spreading with a conventional Desaga spreader (0.25 nnn layer) is heat activated for 20 rain at 120C, cooled for 30 rain, spotted, and ehromatograms developed in chambers lined with solvent-saturated paper (2). Spots are detected by spraying with a 0.6% solution of potassium dichromate in 55%(by wt) sulfuric acid followed by heating for 30 rain at 180C in a forced draft oven or by exposure to iodine vapors. After development, spots are circled and lettered for identification and several blank areas corresponding in size to the sample spots are marked off. A typical ehromatogram of each series is photographed (Polaroid camera) and the spots recovered by aspiration.
Aspiration of the spots directly into 30 ml Xjeldahl digestion flasks is accomplished by fitting a rubber stopper with two plastic tubes removed from plastic wash bottles. One tube with a pointed end serves as the intake and the other tube for attachment to a water pump for suction. Adsorbent is prevented from passing out of the digestion flask during aspiration by adding 0.9 ml of 72% perehlorie acid (used subsequently for digestion) to the flask to act as a liquid trap by moistening the lower bulb portion of the flask and by insertion of a 1 cm square of" Kimwipe" or similar light weight paper into the end of the suction tube to serve as a filter. After aspiration, the plastic tubes are tapped to remove any dry powder and the paper filter pushed with a wire plunger into the flask. Digestion of the flask contents is carried out on an electrically heated Kjeldahl rack with water-pump suction to remove any escaping
Springer