Immobilized heparins anticoagulant activity

Methods for preparing heparin-containing polymeric materials by means of ionic and covalent immobilization of heparin on various polymers are surveyed. The data on the biological activity of heparin are discussed as well as the probable mechanisms of thromboresistance enhancement endowed to polymeric materials by this anticoagulant. Some approaches toward an increased efficiency of anticoagulant properties of immobilized heparin are analyzed, and the position of heparin-containing polymers among other biomedical polymers is discussed. 

Table 11 demonstrates, as in the previous case, the increased amount of immobilized heparin when the spacer was used the anticoagulant activity of immobilized heparin being around 10% of the initial heparin activity. The reason for the observed loss of initial activity is probably the inaccessibility of the immobilized heparin for high-molecular proteins (thrombin with a molecular mass of 34000 and antithrombin III with a molecular mass of 65 000) as the permeability of the grafted gels, whose water content was only 55 %, [Pg.113]

In all of the reported cases, anticoagulant activity of immobilized heparin, as was already stressed above, was less than the activity of heparin in solution, which was attributed to the low accessibility of heparin molecules for physiological substrates. Increasing the porosity of the polymer, i.e., increasing the accessibility of the immobilized heparin led to an increase in its anticoagulant activity 100). 

To elucidate heparin anticoagulant effects caused by covalent coupling via specific functional groups, those heparin groups involved in immobilization reactions are first derivatized, and the effect of derivatization on anticoagulant activity is determined. 

This chapter focuses on the effects of anticoagulant activity caused by specific functional group derivatization of heparin, and on preliminary immobilization spacer group evaluations. The functional groups selected for immobilization are hydroxyl and carboxylic heparin groups. 

Heparin with high anticoagulant activity has been recovered from crude commercial preparations by affinity chromatography on columns of immobilized human a-thrombin. ... 

Functions.—Heparin fractionated by gel filtration appeared to bind to two sites on antithrombin III (association constants 0.6 x 10 and 0.2 x 10 moll" ), whereas heparin prepared by affinity chromatography on matrix-bound antithrombin III appeared to bind to only one site (association constant 2.3 x 10 moll ). These results suggest that one of the binding sites on antithrombin III does not bind the most active heparin components, but accommodates heparin-like molecules which, although similar in size to the active heparin components, have little or no anticoagulant activity. Heparins with high or low affinities for antithrombin III exhibited no differences in their abilities to bind lipoprotein lipase. Studies of the interaction between the lipoprotein lipase from cow s milk and Sepharose-immobilized heparin have shown that heparin is poly-disperse. Whereas heparin facilitated complex formation between a-thrombin and antithrombin III, it had little effect on the interaction between p-thrombin and antithrombin III. ... 

Heparin fractions of different molecular weights and anticoagulant activities have been prepared by affinity chromatography on immobilized protamine. A heparin fraction (mol. wt. 7.9 x lO ) exhibited binding to anti-thrombin III with a stoicheiometry of 2 1, whereas two higher molecular weight fractions exhibited 1 1 binding. Similar results have been reported for two fractions from porcine... 

Dextran, a branched glycan polymer, can be chemically sulfated to prepare dextran sulfate,21 having low anticoagulant activity.2 Dextran sulfate has been used as a heparin replacement in anticoagulation and has recently been immobilized on plastic tubes to prepare non-thrombogenic... 

Fig. 9 demonstrates the proportionality between the amount of immobilized trypsin and the lytic activity of the trypsin-containing material. The higher lytic efficiency of the binary trypsin-heparin systems seems unexpected (cf. curves 2 and 4, and 5 and 6 of Fig. 9), however. Thus, immobilized trypsin and heparin of the described polymeric materials mutually affected each other. This is manifested by the enhanced anticoagulant action of heparin promoted by trypsin and by greater lytic efficiency of trypsin provoked by heparin. 

Anticoagulant properties are due to the formation of a complex between heparin and antithrombin (ATIII) heparin increases ATIII activity, inhibiting thrombin, which is responsible for the formation of the clot [8]. Although this complex is already characterized by a weak affinity, the exact mechanism of association between heparin and antithrombin is not exactly known. A multistep protocol of immobilization of heparin on silica beads permitted high-performance chromatographic phases to be obtained. Thus, it has been possible to evidence a slightly stronger affinity of heparin for antithrombin than for thrombin. 

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