Heparin isolation

Pejler, G., Danielsson, A., Bjork, I., and Lindahl, U. (1987). Structure and antithrombinbinding properties of heparin isolated from the dams Anomalocardia bmsiliana and Tivela mactroides. ]. Biol. Chem. 262,11413-11421. 

Lipmann and Fischer later confirmed the presence of nitrogen and sulfur in heparin isolated by the procedure of Charles and Scott. 

Heparan-sulphate was isolated for the first time as a byproduct of commercial heparin isolation in 1948 and described as a low sulphated heparin by Jorpes and Gardell. During its early stage some researchers thought heparan-sulphate to represent a new class of glycosaminoglycans. Others supposed it to be an intermediate of the heparin-biosynthesis. ... 

Dicoumarol [66-76-2] (5) was isolated from spoiled sweet clover hay. It is prepared synthetically by reaction of 4-hydroxycoumarin with formaldehyde (85). It is used in anticoagulant therapy often associated with heparin. 

Heparin was immobilized on the WPG-P A and the sorbent was used for isolation of the fibroblast growth factor from bovine hypophysis. Mitogenic activity of the factor purified on the heparin-WPG-PA and Heparin-Sepharose, as estimated with mice fibroblasts of line N1H-3T3, was virtually the same [130]. 

Protamine sulfate is a mixture of basic polypeptides isolated from salmon sperm that is used to neutralize heparin in vitro or in vivo. 

Some of the polysaccharides constituting intermediate stages in the biosynthesis of heparin have been physically isolated.23 Their structure is strikingly similar to those of heparan sulfates,24 thus re-opening the often-raised question as to whether heparan sulfate indeed represents products of incomplete biosynthesis of heparin. A detailed account of the structure and biological properties of heparan sulfate is beyond the scope of this article. However, in view of similarities of structural fea-... 

Most of the common methods of isolation of heparin (described in sufficient detail in monographs128-30) are based on a procedure, developed by Charles and Scott,31 involving autolysis of the tissue (originally beef liver and beef lung), extraction with alkali, coagulation of proteins by heating, and precipitation of a heparin - protein complex by acidification. Heparin is recovered from the complex by reprecipitation with ethanol, or acetone, or both. Fats are removed by extraction with ethanol, and proteins by treatment with trypsin. Modifications of this proce-... 

Examination of early -n.m.r. spectra of heparin and of chemically modified heparins121 prompted a reinvestigation of N,0-desulfated, carboxyl-reduced heparin, leading to the isolation of substantial amounts of L-iditol pentaacetate.121,122 In addition, improved conditions for the acid hydrolysis of heparin and carboxyl-reduced heparin gave increased recoveries of L-iduronic acid and 1,6-anhydro-L-idose, respectively.123 These findings confirmed the L-enantiomeric designation of the iduronic acid, and established that it is the main uronic acid in heparin. 

Both positional linkages (uronic acid to hexosamine and hexosamine to uronic acid) were established as being (1 — 4) by structural analysis of the previously mentioned (see Section IV), crystalline disaccharides containing D-glucuronic acid, isolated from an acid hydrolyzate of carboxyl-reduced heparin.128-129 Further evidence was obtained from the structure of the D-glucuronic acid-containing counterpart of disaccharides 6 and 8, obtained as minor products from pig-mucosal heparin following nitrous acid deamination,1110 136-138 and acid hydrolysis followed by N-acetylation,130 respectively. 

In vitro platelet activation is dependent on the anticoagulant that is used for blood collection. In one study it was demonstrated that PF4 levels in platelet-poor plasma isolated after incubation without any stimuli for 1 hour at 37°C were as follows conventional heparin, 1180 ng/ml hirudin, 469 ng/ml citrate, 440 ng/ml and EDTA, 217 ng/ml (110). EDTA appears to suppress platelet degranulation. PF4 levels obtained with a low-molecular-weight heparin preparation called Frag-min were, however, comparable to those obtained with hirudin (110). 

Howell88 considered it to be a derivative of D-glucuronic acid, but this view was not confirmed by Charles and Scott,89 who isolated a crystalline barium heparinate from beef lung and detected the presence in it of an amino sugar. 

Kotlikoff Coming back to the context of RyRs coexisting with InsP3 receptors, how do you conceive of this system working in isolation from the RyR Ca2+ sensor In some ways it is easier to do the experiment to exclude the InsP3 receptors because you can use something like heparin and completely block it, and have some confidence. The alternative experiment to block all RyRs is a little dirtier. What are your thoughts on how this system can exist in waves, and what is the wave transmission speed in the arteries ... 

Heparinized blood samples may be stored at 4°C for up to 48 h without affecting the SCE response (Lambert et al., 1982). If the test agent is known to react with serum or red blood cells, the mononuclear lymphocytes may be isolated by use of a Ficoll/Hypaque gradient (Boyum, 1968). 

Linhardt RJ, Ampofo SA, Fareed J, Hoppensteadt D, Mulliken JB, Folkman J (1992) Isolation and characterization of human heparin. Biochemistry 31 12441-12445. 

Gl. Ganesan, D., and Bradford, R. H., Isolation of apolipoprotein-free lipoprotein lipase from human post-heparin plasma. Biochem. Biophys. Res. Commun. 43, 544-549 (1971). 

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