Depolymerization of heparin by complexed ferrous ions.

Treatment of porcine heparin with the ferrous-EDTA complex and ascorbic acid for 24 h at 37 degrees C results in the degradation of most of the glycosaminoglycan to smaller fragments. About 65% of the products comprise oligosaccharides composed of less than 30 sugar units. The extent of depolymerization is decreased significantly if ascorbate or EDTA is not included in the reaction mixture. Gel filtration of the reaction products yielded fractions with narrow chain length ranges. The sulfate content of the fractions and their electrophoretic mobilities on cellulose acetate indicate that the components have equivalent charge densities. Depolymerization products with 20 or more sugar units retain significant anticoagulant potencies as measured by their effect in accelerating the neutralization of factor Xa by antithrombin.

The flagellar membrane of Ochromonas danica. Lipid composition.

The lipids of the whole flagella and the flagella membrane of the phytoflagellate Ochromonas danica were isolated and compared with those of the whole cell. The polar lipids were separated by two-dimensional thin layer chromatography. One-dimensional thin layer chromatography was used for the separation of the nonpolar lipids. In all respects the lipids of the whole flagella were identical with those of the flagellar membrane. These methods established the presence in flagellar membrane of the polychlorosulfolipids of O. danica as more than 90 molar per cent of the total polar lipids. These sulfolipids had been previously characterized as 1,14-docosanediol-1, 14-disulfate and 1,15-tetracosanediol-1,15-disulfate, containing zero to six chloro groups substituting for hydrogen on the chain. Seven unknown polar lipids were found. Both phosphorus analysis on each lipid and the molybdenum spray reagent for phospholipids on the chromatogram showed that there is no phospholipid present in O. danica flagellar membrane. Positive reactions to the diphenylamine spray reagent suggest that up to four of the unknown polar lipids are glycolipids. Of these, three reacted positively with ninhydrin. All of the unknown lipids reacted with the acidified 2,4-dinitrophenylhydrazine spray reagent suggesting the presence of aldehyde, ketone, glycoside, or plasmalogen. One unknown substance appeared near the origin of thin layer chromatograms. It showed a positive reaction with Dragendorff reagent, suggesting the presence of a quaternary amine group. This substance is presumed to be nonlipid, since it is not synthesized from [1-14C]acetate under the growth conditions used, as revealed by autoradiograms of thin layer chromatograms. It contained 35% hexose or hexosamine. It is devoid of phosphorus (0.7%) and is less than 4% protein (or phenolic groups or peptide), as judged by the Lowry assay using bovine serum albumin as a standard. Analysis of the nonpolar lipids of the flagellar membrane showed that free fatty acids constitute about 12 molar per cent of the total lipids. These fatty acids could be true components of the membrane or artifacts of the extraction procedure although every precaution was taken to prevent artifactual production of free fatty acids. The sterols constitute nearly 10 molar per cent of total lipids. Sterol esters were absent from the membrane. There are two additional major unknown nonpolar lipids present. The implications of such a high proportion of chlorosulfolipids as a polar lipid component in the membrane are important because of the unique structures of these lipids, which have ionic groups at or near both ends of the aliphatic chain.