1. ANTICOAGULANT

2. DEFINITION
A substance that prevents coagulation or clotting of blood but doesn’t dissolve an already formed clot.
Uses
Storage of blood for blood transfusion or hematological testing
Therapeutic

3. Anticoagulant Purpose of using anticoagulants:
When Blood is collected, it clots after sometime. The anticoagulants are the chemical agents which prevent the clotting of Blood when mixed with Blood in proper proportion.
Purpose of using anticoagulants:
For study of various constituents of Blood components.
Study of coagulation(clotting of Blood).
Preservation of Blood in Blood Bank.
Properties of anticoagulant
It must be soluble in Blood.
It must be keep the blood in fluid condition,
It must not be bring haemolysis of Blood cells.
It must not be change the size of RBC.
It must minimize destruction of leukocytes.
It must minimize aggregation of platelets.

4. Commonly used anticoagulants
EDTA(Ethylene Diamine Tetra Acetic acid)
Double oxalate
Sodium citrate
Sodium Fluoride
Heparin
ACD(Acid Citrate Dextrose)
CPD(Citrate Phosphate Dextrose)
CPDA(Citrate Phosphate Dextrose Adenine)

5. FOR ROUTINE HAEMATOLOGY ACD
EDTA
FOR ROUTINE HAEMATOLOGY
BUFFERED SODIUM CITRATE
FOR COAGULATION STUDIES
POTTASIUM OXALTE OR SODIUM FLOURIDE
GLUCOSE DETERMINATION
NO ADDITIVE
COLLECTION OF SERUM
ACD
PRESERVE RBC FOR BLOOD BANKING &
HLA Typing
HEPARIN
INHIBIT THROMBIN
ACTIVATION

6. Non-calcium chealeter
Anticoagulant
Calcium chealeter
Oxalates
Ammonium oxalate
Potassium oxalate
Double Oxalate
E.D.T.A.
Tri-Sodium citrate
Non-calcium chealeter
Heparin
Warfarin

7. Classification of anticoagulant
1.Calcium chelator
Bind with Calcium
Oxalates
It is following forms:-
-Ammonium Oxalate
-Potassium Oxalate
- Double Oxalate
EDTA (Ethylenediaminetetraacetiacid)
Tri-sodium citrate etc.
2. Non-calcium chelator
Do not bind with Calcium
Heparin
Warfarin etc.
Most of the anticoagulants used in laboratory act by binding with calcium and prevent clotting of blood since calcium ion is essential for many of the steps in coagulation mechanism.

8. CLASSIFICATION USED IN VIVO Parenteral anticoagulants
These are drugs used to reduce the coagulability of blood classified into:
USED IN VIVO
Parenteral anticoagulants
a) Heparins (ITI)
High Molecular Weight Heparin
Unfractionated Heparin (UFH)
Low molecular weight heparin (Enoxaparin, Dalteparin, Tinzaparin, Reviparin, Danaparoid)
b) Heparinoids (DTI)-Heparan sulfate, Hirudin, Lepirudin, Bivalirudin, Argatroban
1. Indirect Thrombin Inhibitors: HMWH, LMWH
2. Direct Thrombin Inhibitors: Hirudin, Lepirudin, Bivalirudin, Argatroban

9. Oral anticoagulants (1)Coumarin derivatives :Bishydroxycoumarin (dicumarol), Warfarin sod, Acenocoumarol (Nicoumalone), Ethylbiscoumacetate (2)Indandione derivative: Phenindione. USED IN VITRO (a)Heparin (b)Sod. citrate :used in blood banks to store the blood (c)Sod. oxalate : (d)Sod. editate :
used as an anticoagulant in laboratory

10. Ammonium oxalate:- Potassium oxalate:- Oxalates
This is used at a concentration of 2 mg for 1 ml of Blood. This anticoagulant causes swelling of the RBC therefore, it is not recommended for use with blood for PCV,ESR and cell morphology.
Potassium oxalate:-
It is used at a concentration of 2 mg for 1 ml of Blood. This anticoagulant is most often used for chemical analysis. It causes shrinkage of RBC therefore it is also is not recommended for the study of PCV,ESR and other cell morphology.
Double oxalate:-
Ammonium oxalate and Potassium oxalate are combined together to balance the swelling effect of Ammonium oxalate and the shrinking effect of Potassium oxalate on the red cells. It is the mixture of 3 parts of Ammonium oxalate and 2 parts of Potassium oxalate which is prepared as follows:-
Ammonium oxalate 2.4 gram
Potassium oxalate 1.6 gram
Distilled water ml

11. E.D.T.A.(Ethylenediamine tetra-acetic acid)
It is the most commonly used anticoagulant in Haematology lab because E.D.T.A. is the most powerful calcium chelating (binding) agent we have. It gives the best preservation to the cell morphology therefore E.D.T.A. is the preferred anticoagulant for all cell count and blood smear preparation.
E.D.T.A. is used in two different forms:-
Di-sodium E.D.T.A. salt (Versene)
Di-potassium E.D.T.A. salt (Sequestrene)

12. Tri-sodium citrate It is used as a liquid form. It binds with calcium.
The concentration of Tri- sodium citrate is used as 3.8%.
For PT (Prothrombin Time) a kind of coagulation test, 3.8% Tri-Sodium citrate is used at the ratio of 1:9 i.e. 1 part of anticoagulant and 9 part of Blood.( 200 µl of 3.8% Tri-Sodium citrate and 1.8 ml of Blood is used.)
Fro ESR (Erythrocyte Sedimentation Rate) by Westerngreen method
1 part of 3.8% Tri-Sodium citrate is mixed with 4 part of Blood that is the ratio of 1:4.( 400 µl of 3.8% Tri-Sodium citrate and 1.6 ml of Blood is mixed).
It do not preserve the cell morphology.

13. Heparin
It is a natural anticoagulant and is normally present in the blood in small amount and highly acidic. This is the best anticoagulant for open heart surgery and it causes minimum haemolysis.
It is very expensive.
It produce black back ground in the smear so it is not use for smear preparation.

14. Intrinsic Pathway Extrinsic Pathway Vit. K dependent Factors
Blood Vessel Injury
Tissue Injury
Tissue Factor
XII
XIIa
HMW-Kininogen
Thromboplastin XI
XIa
XIIa, Xia, Kallikrein
ca+ IX
IXa
VIIa
VII
ca+
ca+ X Xa X
XIII
ca+
ca+
Thrombin
Prothrombin
Factors affected
By Heparin
XIIIa
Fibrinogen
Fribrin insoluble
Vit. K dependent Factors
Affected by Oral Anticoagulants
Stabilised Fibrin threads

15. Mechanism + Heparin No heparin Active clotting factors
Slow AT III
Inactive clotting factors
Heparin
Active clotting factors
Fast AT III +
Inactive clotting factors

19. ORAL ANTICOAGULANTS

20. Vitamin K-Dependent Clotting Factors
VII
The four Vitamin K dependent clotting factors are synthesized in the liver. IX X II
Synthesis of Functional Coagulation Factors

21. Warfarin Mechanism of Action
Vitamin K
VII
Antagonism of
Vitamin K
Warfarin acts as an anticoagulant by blocking the ability of Vitamin K to carboxylate the Vitamin K dependent clotting factors, thereby reducing their coagulant activity. IX X II
No Synthesis of Functional Coagulation Factors
Warfarin

22. Mechanism of Action
Coumarins block the Gamma Carboxylation of glutamic acid residues of Clotting factors II,VII, IX, X , endogenous anti-coagulants C & S.
This is coupled with oxidative deactivation of Vit K
Coumarins and Indanediones (-) enzyme Vit K epoxide reductase that converts Vit K epoxide to its active hydroquinone (reduced form)
Thus they prevent the activation of Vit K and hence along with it carboxylations of clotting factor residues

23. Why Carboxylation is necessary?
Necessary for ability of clotting factors to bind Ca+ and to get bound to phospholipid surfaces which is necessary for coagulation
Factor VII affected first, then IX, X, and finally Factor II (depends upon half lives of circulating factors)

26. Anticoagulant used in Blood Bank
ACD (Acid Citrate Dextrose)
CPD ( Citrate Phosphate Dextrose )
CPDA (Citrate Phosphate Dextrose Adenine)