Analysis of lipid content and quality in Arabidopsis plastids

Chloroplasts of plants contain an intricate membrane system, the thylakoids, which harbor the complexes of the photosynthetic machinery. Chloroplasts are confined by two membranes, the inner and outer envelope. The major glycerolipids of chloroplasts are the glycolipids monogalactosyl diacylglycerol (MGD), digalactosyl diacylglycerol (DGD), and sulfoquinovosyl diacylglycerol (SQD). Furthermore, two phospholipids, phosphatidyl glycerol (PG) and phosphatidyl choline (PC), are found in chloroplast membranes. The photosystems and light-harvesting complexes in the thylakoids are rich in photosynthetic pigments (chlorophyll, carotenoids, and xanthophylls) and contain a unique set of prenylquinol lipids (tocochromanol/vitamin E, plastoquinol, and phylloquinol/vitamin K1). In this chapter, methods for the isolation and quantification of chloroplast and leaf glycerolipids and prenylquinol lipids are presented. Glycerolipids are separated by thin-layer chromatography prior to conversion of the fatty acids into methyl esters. Fatty acid methyl esters are subsequently quantified by gas chromatography. Prenylquinol lipids are separated by HPLC and quantified by UV absorption (plastoquinol) or fluorescence (tocochromanol, phylloquinol).