The Use of High Pressure Liquid Chromatography for the Identification and Preparation of Pigments Concerned in Photosynthesis

The chlorophylls and carotenoids present in preparations from chloroplasts of marine algae can be extracted and separated by high pressure liquid chromatography (H.P.L.C.). The reverse-phase columns; Partisil 10 ODS (Whatman), μ Bondapak C18 and μ Bondapak CN (Waters Associates) gave good separation of the different pigments. Addition of the ion-pairing agent tetrabutylamnonium phosphate to the methanol/water solvents gave improved separations with complex mixtures and identification was facilitated by examination of the column eluant at 440nm where a ll the pigments absorbed and at 650nm where only chlorophylls absorbed. The method allowed the isolation of individual pigments for further study.