Barium acid heparinate

L-arabinose and D-xylose in hemi-cellulose of sugar Cane, V, 279 Banana starch. See under Starch. Barium acid heparinate, III, 146 Barium 2-desoxy-D-gIuconate, III, 144 Barium D-gluconate, III, 144 Barium hypobromite, III, 163 Barium salts, in preparation of aldonic acids with NaCN, I, 23 Bark, tree, pentosan content, V, 271 Barley, alpha amylase of malted, V, 255-265... 

Wolfrom ML, Weisblat DI, Karabinos JV, McNeely WH, McLean J. Chemical studies on crystalline barium acid heparinate. J Am Chem Soc 1943 65 2077-2085. 

Astrup and Jensen converted the crystalline barium acid salt to the sodium salt of heparin, which they subjected to elemental analysis. Results in good agreement with those of Charles and Scott were claimed w hen allowance was made for the hydration. Reinert and Winterstein also prepared heparin by the benzidine-salt procedure and obtained 25-50 mg. 

In 1943, Wolfrom and coworkers published the results of a thorough investigation of the elemental composition of the barium acid salt of heparin, and special attention was given to the preparation of anhydrous material. The results are quoted in Table II, column 4. Summation of the analytical results apparently accounted for 88.6% of the material present. However, there was by no means unaninious agreement that the barium acid salt of heparin was the most pure and active material prepared. In particular, Jorpes, ° who had been very critical of the evidence for the crystallinity of the barium acid salt, continued to purify his heparin prepa-... 

At this point, it is pertinent to review the information that may be gained from analytical data concerning the structure of heparin. Since 1935, many analytical data on heparin have been accumulated, but, as noted earlier, much of the work was performed on inhomogeneous or incompletely dried samples. The most thorough quantitative examination of the composition of heparin was made by Wolfrom and coworkers" on the barium acid salt. 

Meyer and Wolfrom and their coworkers detected the uronic acid in the barium acid salt of heparin both qualitatively and quantitatively, and considered it to be the source of the acidity in the acid salt. Amino sugar was also estimated. These results, which cannot be said to be as precise as the elemental data, indicate a ratio of 1 1 for amino sugar and hexuronic acid. 

Wolfrom and coworkers have summated their analytical data on the barium acid salt of heparin, and have expressed the data in terms of a tetrasaccharide unit which comprises two molecular proportions each of 2-amino-2-deoxy-D-glucose residue and D-glucuronic acid residue, and 5 (rather than 6) ester sulfate groups, in an N S ratio of 2 5. The barium sulfur ratio was 1 2, which indicated that the sulfur is essentially present as ester sulfate, and hence the acidity must be due to the carboxyl group, in accordance with the shape of the titration curve. " ... 

Little work has been published concerning the isolation of oligosaccharides following graded, acidic hydrolysis of heparin. Wolfrom and co-workers have studied the partial, acid hydrolysis of a carefully purified heparin. (Purification was effected through the benzidine salt and the neutral barium salt was fractionally precipitated from water. The less-soluble fraction, after conversion to the barium acid salt, had a N S ratio of 2 5 and 5 sulfate groups per tetrasaccharide period, consumed 1 mole of periodate per tetrasaccharide period, and was shown to be homogeneous by electrophoresis and by countercurrent distribution. The molecular... 

Application of the procedure to heparin gave a degraded product wffiich was sulfur-free. In work described in a later paper, this material was de-0-acetylated and then was found to consume 2 moles of periodate per tetra-saccharide unit, without the formation of formic acid or formaldehyde. The original heparin (see p. 355) consumed 1 mole of periodate, and contained 5 sulfate groups, per tetrasaccharide unit. From these data, together with those reported for heparosinsulfuric acid (see p. 356), Wolfrom and coworkers have postulated VI as a probable structure for the barium acid... 

The reagent forms water-insoluble cetyltrimethylammonium salts with acidic polysaccharides which are useful for isolation and purification of such substances. - It is an effective precipitant for sulfated carbohydrates, even of sugar sulfates of low molecular weight. For acidic, nonsulfated polysaccharides molecular weight is important, since the salts of simple uronic acids are water soluble. Wolfrom- purified commercial heparin through the cetyltrimethylammonium salt to a product readily convertible into the crystalline barium acid salt. 

The Isolation of L-Iduronic Acid from the Crystalline Barium Acid Salt of Heparin, M. L. Wolfrom, S. Honda, and P. Y. Wang, Carbohyd. Res., 10, 259-265 (1969). 

Wolfrom ML, Honda S, Wang PY. Isolation of L-iduronic acid from the crystalline barium acid salt of heparin. Carbohydr Res 1969 10 259-265. 

Glycosaminoglycanes (hyaluronic acid, heparin, chondroitin sulfate, etc.) Cellulose acetate membranes 0.2 mol 1" Barium acetate, pH 5.0 or pH 8.0, 0.05moll" lithium chloride, 0.01 mol 1" HCI (pH 2.0). Two dimensional 1st direction 0.1 moir pyridine, 0.47 mol r formic acid, pH 9.0 2nd direction 0.1 moir barium acetate, pH 8.0 Aldan blue... 

Heparin in the acid state, and its amorphous sodium and potassium salts are very soluble in water, whereas its crystalline barium acid salt and its amorphous lead salt have a low solubility. In addition to the barium salt, other salts of heparin have a crystalUne appearance, for example those formed with piperidine, -pentylamine, and isopentylamine. Values of the optical rotation of the barium salt have been reported from -1-45 to -t-yo . 

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. 

It is of interest to note that the crystalline barium heparinate, which is an acid salt, tends to lose its anticoagulant powers very readily and that the authors consider that sulfur content gives no indication of heparin activity.93 Some polysaccharide sulfuric esters from marine algae possess anticoagulant activity. 

Subsequently, Howell improved the separation of heparin from protein by introducing the use of Lloyd s reagent (aluminum silicate) in acetic acid. The final stage then involved the precipitation of the heparin with excess barium hydroxide. Howell claimed that his heparin preparation con-... 

Some difficulty was experienced in establishing a strict correlation between liberation of amino group and sulfate release. For example, Wolfrom and McNeely, and Jorpes, Bostrom and Mutt, did not obtain correspondence between these when heparin was treated with hot, dilute acid. It was suggested that retention of the barium sulfate in the colloidal state (peptization) by the heparin might be responsible for inaccurate results in sulfate analysis. If, however, heparin was first treated with hot, dilute alkali, then, on subsequent acid treatment, correspondence between release... 

Available forms Solid elemental sulfur, sulfate in weak hydrochloric acid, barium sulfide in barium hydroxide solution, elemental sulfur in benzene solution, in tagged compounds such as carbon disulfide, chlorosulfonic acid, thiourea, sulfanilamide, thiamine, heparin, insulin, Sucaryl, etc. 

The crystalline barium salt of beef heparin proved to have a potency of 100 units/mg. One-third by weight of this salt is base and water and variation in percent of base and of water can be responsible for differences in other values for heparin. Hence it is better to express the values in equivalent anhydrous free acid. When I did this for... 

Chapter 9. A substrate used in the assay of L-iduronic acid 2-sulphate sulphatase was obtained from sodium heparinate by methanolysis followed by nitrosation. Disaccharide monosulphates presumed to contain D-(methyl a-L-idopyranosiduronate-2-sulphate)-(l 4)-2,5-anhydro-D-mannose were isolated by gel filtration. Brucine L-guluronate and o-mannuronate, on the other hand, were obtained by treatment with brucine of the barium salts of the acids formed on hydrolysis of alginic add. ... 

Heparin ( a-heparin ), a blood anticoagulant, is found in liver, lung, thymus, spleen, and blood. It can be extracted from autolyzed beef liver by sodium hydroxide. After tryptic digestion of the proteins, heparin is precipitated with acidic ethanol (57), Heparin is usually crystallized as the barium salt. Purification through the free acid and reformation of the barium salt causes a decrease in the activity, as heparin is labile toward acidic... 

The most convenient starting material for obtaining large amounts of dermatan sulfate is the residual liquor of the technical preparation of heparin. Treatment with alcohol gives a precipitate, which is fractionated through the barium salt with acetic acid, then through the zinc salt with methanol. From i kg of crude precipitate, 20 g of dermatan sulfate was thus obtained. Further purification has also been carried out through the cobalt salt. 

---