The complex lipids from a strain of Sarcina lutea were isolated and separated into fractions on diethylaminoethyl cellulose acetate and silicic acid columns. These fractions were monitored in several thin-layer chromatography systems. The various lipid types were characterized by their behavior in thin-layer systems and by an analysis of their hydrolysis products. The fatty acid composition of the column fractions was determined by gas-liquid chromatography. A number of components (13) were separated by thin-layer chromatography and characterized. The major components were polyglycerol phosphatide (17.0%), lipoamino acids (15.1%), phosphatidyl glycerol (13.8%), and an incompletely characterized substance (15.0%). Minor constituents included phosphatidyl inositol (5.5%), phosphatidic acid (4.2%), phosphatidyl serine (2.0%), and phosphatidyl choline (1.0%). No phosphatidyl ethanolamine was observed. Inorganic reagents, solvents, and Hyflo Super-Cel were obtained from Fisher Scientific Co., Silver Spring, Md., and were the highest grades available. Chloroform, hexane, and propionic acid were redistilled before use. The methanol, acetone, benzene, and diisobutyl ketone were gaschromatographically pure. Synthetic phosphatidyl ethanolamine, phosphatidyl serine, and phosphatidyl choline (all dipalmitoyl) were Agrade from Calbiochem. Lecithin and cephalin standards were isolated from soy phosphatides by silicic acid column chromatography, according to Fleischer et al. (1962), and examined for homogeneity by thin-layer chromatography (TLC) according to Horrocks (1963). All other lipid standards used were the highest grades available from Mann Research Laboratories, New York, N.Y., or Applied Science Laboratories, Inc., State College, Pa. Rhodamine 6G and ninhydrin were obtained from Eastman Organic Chemicals, Rochester, N.Y. TLC apparatus and materials were obtained from Brinkmann Instruments, Inc., Great Neck, N.Y. Culture conditions. S. lutea ATCC 533 was cultured at 25 C for 24 hr in tryptic soy broth (Difco) under forced aeration. The prepared medium was found to contain less than 0.009% lipid. Cells were harvested, as previously described (Huston and Albro, 1964), immediately after the growth; all operations were performed below 5 C. Lipid extraction. Extraction of lipids and isolation of the "complex lipid" fraction was accomplished as previously described (Huston and Albro, 1964). Nonlipid contaminants that were not removed by the Folch washing procedure were separated on Sephadex (Wells and Dittmer, 1963) and on DEAE cellulose columns (Rouser et al., 1961). These contaminants amounted to 8 to 14% 768
The complex lipids of a strain of Saroina lutea were isolated and separated into fractions on DFAE cellulose acetate and silicic acid columns.These fractions were monitored in several thin-layer cbromatography aystems.The various lipid types were characterized by their behavior in thin-layer systems and by an analysis of their hydrolysis products.The fatty acid composition of the column fractions was determined by gas-liquid chromeatography.Thirteen components were separated by thin-layer chromatography and characterised. Ethanolamine is the most commonly reported nitrogen-containing component; chloline is found in some species. Recent reports have also described the isolation of lipoamino acid complexes in the complex lipids of bacteria. -6We have reported 7 that Sarcina lutea contains a complex mixture of highly polar lipids that comprise approximately 23% of the total extractable lipid. The determination of the composition of this polar fraction is reported in this manuscript. II. MATERIALSThe preciseness of the experimental procedures and .h;! small quantities of lipids under investigation required extreme purity and reliability in the reagents employed. For these reasons the following materials were obtained from the sources shown: I) Whatman diethylaminoethyl (DEAE) celluloseScientifica, Clifton, New Jersey.2) Silicic acid, analytical:reagent grade -Mallinckrodt Chemical Works, New York, New York.3) Inorganic reggents, solvents, and Hyflo Super Cel -Fisher Scientific Company, Silver Spring, Maryland.4) Synthetic phosphatidyl ethanolamine, phosphatidyl serine, and phosphatidyl choline (all dipalmitoyl) A-grpde -Calbiochem,
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