Nanogram detection of phospholipids on thin-layer chromatograms.

An extremely sensitive method for the quantitative determination of phospholipids on silica gel-precoated thin-layer chromatography plates is reported. The procedure is based on the successive application of two lipid spray reagents. Firstly, phospholipid chromatograms are sprayed with the lipophilic fluorochrome 1,6-diphenyl-1,3,5-hexatriene, followed by application of the phosphorus-specific molybdenum blue (Dittmer-Lester) reagent. Compared to the single use of the respective dyes, the consecutive utilization of both spray reagents resulted in an enormous enhancement in sensitivity of at least one order of magnitude. The procedure was proved with eight phospholipids of animal origin, and visual detection limits of down to 10 ng were achieved. The described technique provides a sensitive and specific means for the analysis of phospholipids on the nanogram scale by augmenting the molybdenum blue staining with the fluorochrome 1,6-diphenyl-1,3,5-hexatriene operating as an enhancer.