Densitometry on Thin-Layer Chromatographic Plates by Fourier Transform Near-Infrared Spectrometry

Abstract

Direct measurement by Fourier transform near-infrared (FT-NIR) spectrometry on thin-layer chromatographic plates is described. Zones separated on a high-performance thin-layer chromatographic plate fixed on the sample stage of a microscope linked with an FT-NIR spectrometer were scanned with a narrow beam of light (0.4-mm square). The NIR chromatogram and spectra of the separated zones were then obtained by the diffuse transmission method. Three kinds of phospholipids that have essentially no absorption bands in the ultraviolet and visible regions but that do have native absorption bands in the NIR region were used to evaluate the usefulness of this method. The limit of detection for phosphatidylcholine was under 1 μg, and the correlation coefficient of the calibration curve obtained from derivative spectra was 0.998 for amounts from 1.25 to 10 μg.