On the Relationship Between Molecular Mass and Anticoagulant Activity in a Low Molecular Weight Heparin (Enoxaparin)

A low molecular weight heparin (enoxaparin, mean molecular weight approximately 4,400) was separated by gel chromatography into eight different fractions with a narrow distribution around the following mean molecular weights: 1,800, 2,400, 2,900, 4,200, 6,200, 8,600, 9,800 and 11,000. We compared the influence of enoxaparin on the generation of thrombin in plasma to that of the eight fractions. We determined: a) the % of material with high affinity to antithrombin III (HAM) and the % of HAM above the critical chain length necessary to allow for thrombin inhibition (ACLM), b) the specific catalytic activity on the decay of endogenous thrombin, and c) the inhibition of over-all thrombin formation in the extrinsic and the intrinsic pathway. From b and c we calculated the inhibition of prothrombin conversion in these pathways. We found that a) there is a gradual decrease of the HAM fraction with decreasing molecular weight; b) the specific catalytic activity for the inactivation of thrombin does not vary significantly between the fractions when expressed in terms of ACLM; c) the potency to inhibit prothrombin conversion does not vary significantly between the fractions when expressed in terms of HAM.

[1]  J Jesty,et al.  The kinetics of inhibition of alpha-thrombin in human plasma. , 1986, The Journal of biological chemistry.

[2]  P. Owren,et al.  The control of dicumarol therapy and the quantitative determination of prothrombin and proconvertin. , 1951, Scandinavian journal of clinical and laboratory investigation.

[3]  E. A. Johnson,et al.  Molecular weight dependency of the heparin potentiated inhibition of thrombin and activated factor X. Effect of heparin neutralization in plasma. , 1979, Thrombosis research.

[4]  Å. Danielsson,et al.  Role of ternary complexes, in which heparin binds both antithrombin and proteinase, in the acceleration of the reactions between antithrombin and thrombin or factor Xa. , 1986, The Journal of biological chemistry.

[5]  J. Pieters,et al.  The limited importance of factor Xa inhibition to the anticoagulant property of heparin in thromboplastin-activated plasma. , 1988, Blood.

[6]  M. Griffith,et al.  Calcium inhibits the heparin-catalyzed antithrombin III/thrombin reaction by decreasing the apparent binding affinity of heparin for thrombin. , 1983, Archives of biochemistry and biophysics.

[7]  M. Samama,et al.  The inhibition of the generation of thrombin and the antithrombotic effect of a pentasaccharide with sole anti-factor Xa activity. , 1988, Thrombosis research.

[8]  T. Barrowcliffe,et al.  The effect of Ca2+, phospholipid and factor V on the anti-(factor Xa) activity of heparin and its high-affinity oligosaccharides. , 1987, Biochemical Journal.

[9]  V. Fuster,et al.  Role of Thrombin in Arterial Thrombosis: Implications for Therapy , 1991, Thrombosis and Haemostasis.

[10]  H. Hemker,et al.  The Mode of Action of CY 216 and CY 222 in Plasma , 2022 .

[11]  U. Lindahl,et al.  On the molecular-weight-dependence of the anticoagulant activity of heparin. , 1979, The Biochemical journal.

[12]  L. Andersson,et al.  Purification of antithrombin III by affinity chromatography , 1974 .

[13]  T. Barrowcliffe,et al.  The Effect of Calcium Chloride on Anti-Xa Activity of Heparin and Its Molecular Weight Fractions , 1989, Thrombosis and Haemostasis.

[14]  M. Samama,et al.  The inhibition of intrinsic prothrombinase and its generation by heparin and four derivatives in prothrombin poor plasma. , 1990, Thrombosis research.

[15]  H. Hemker,et al.  The Mode of Action of Heparin in Plasma , 1988, Thrombosis and Haemostasis.

[16]  E. Thaler,et al.  A Simple Two‐step Isolation Procedure for Human and Bovine Antithrombin II/III (Heparin Cofactor): a Comparison of Two Methods , 1975, British journal of haematology.

[17]  P. Schoen,et al.  The effect of sulfation on the anticoagulant and antithrombin III-binding properties of a heparin fraction with low affinity for antithrombin III. , 1990, Thrombosis research.

[18]  H. Hemker,et al.  Factor IXa Inhibition Contributes to the Heparin Effect , 1991, Thrombosis and Haemostasis.

[19]  J. Hirsh,et al.  The inhibition of thrombin-dependent positive-feedback reactions is critical to the expression of the anticoagulant effect of heparin. , 1987, The Biochemical journal.

[20]  R. Smith,et al.  Titration of activated bovine Factor X. , 1973, The Journal of biological chemistry.

[21]  H. Hemker,et al.  A Computer Assisted Method to Obtain the Prothrombin Activation Velocity in Whole Plasma Independent of Thrombin Decay Processes , 1986, Thrombosis and Haemostasis.

[22]  J. Jesty.  Analysis of the generation and inhibition of activated coagulation factor X in pure systems and in human plasma. , 1986, The Journal of biological chemistry.

[23]  H. Hemker,et al.  In situ-generated thrombin is the only enzyme that effectively activates factor VIII and factor V in thromboplastin-activated plasma. , 1989, Blood.

[24]  Å. Danielsson,et al.  The binding of low-affinity and high-affinity heparin to antithrombin. Competition for the same binding site on the protein. , 1978, European journal of biochemistry.

[25]  H C Hemker,et al.  The role of phospholipids and factor Va in the prothrombinase complex. , 1980, The Journal of biological chemistry.

[26]  M. Scully,et al.  The relative molecular mass dependence of the anti-factor Xa properties of heparin. , 1986, The Biochemical journal.

[27]  P. Hemker,et al.  Formation of Prothrombin Converting Activity , 1967, Nature.

[28]  D. Lane,et al.  Anticoagulant activities of heparin oligosaccharides and their neutralization by platelet factor 4. , 1984, The Biochemical journal.

[29]  L. M. Smith,et al.  An Approach to Assigning In Vitro Potency to Unfractionated and Low Molecular Weight Heparins Based on the Inhibition of Prothrombin Activation and Catalysis of Thrombin Inhibition , 1988, Thrombosis and Haemostasis.