Normal-phase HPLC Separation of Possible Biosynthetic Intermediates of Pheophytin a and Chlorophyll a′

Abstract

Normal-phase HPLC conditions have been developed for separating the C17³ isoprenoid isomers, which are expected to be formed as biosynthetic intermediates of chlorophyll (Chl) a, Chl a′ (C13²-epimer of Chl a), pheophytin (Pheo) a and protochlorophyll (PChl). The application of these conditions to pigment composition analysis of greening etiolated barley leaves allowed us to detect, for the first time, the C17³ isomers of Chl a′, a possible constituent of the primary electron donor of photosystem (PS) I, P700, and those of Pheo a, the primary electron acceptor of PS II, in the very early stage of greening. The C17³ isomer distribution patterns were approximately the same between Chl a and Chl a′, but significantly different between Pheo a and Chl a′, probably reflecting the similarity and difference, respectively, in the biosynthetic pathways of these pigment pairs.