Quantitative Determination of Natural Glycolipids from Oil Seed by Automated High‐Performance Thin‐Layer Chromatography (HPTLC)

The role of glycolipids in vegetable oil refining and production of bio‐based fuels has not been disclosed so far. Such investigations required a reliable and reproducible quantitative determination of these compounds. Fundamental data were therefore established on the quantitative determination of glycolipids in vegetable oil gums by means of high‐performance thin‐layer chromatography (HPTLC). Concentrating on five abundant natural glycolipid classes found in these oils, identification of a suitable separation method for the employed glycolipid mixture and those parameters relevant for successful detection were considered in detail. The special importance of sample volume when employing quantitative HPTLC was discussed. Acetone/chloroform/water 6:3:0.4 (v/v/v) was identified as a convenient mobile phase for the investigated issue. A derivatization reagent comprising methanol, copper(II)sulfate pentahydrate, sulfuric acid 98 %, and phosphoric acid 85 % was identified. Subsequent heating at 135 °C for 10 min finished the derivatization and enabled detection at λ = 370 nm. Calibration curves ranging from 1500 to 31.25 ng/mL, regarding both peak area and peak height, were determined. The good correlation of parameters enabled the application of the method to real oil gum samples from sunflower and soybean oil. This revealed that digalactosyldiglycerides in combination with either sterylglucosides or acylated sterylglucosides represented the major glycolipid classes in these oils.