FORM 2
THE PATENTS ACT, 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
Provisional Specification [See Sections 10 and rule 13]


Title: In-vitro method for testing bioequivalence of iron-sucrose formulation
Applicant: (a) Astron Research Limited
(b) Nationality: Indian
(c) 10th Floor, Premier House Bodakdev, Opp. Gurudwara Sarkhej - Gandhinagar Highway Ahmedabad 380054

The following specification describes the invention:


FIELD OF THE INVENTION
The present invention relates to a process for rapid assessment of bioequivalence of iron in iron-sucrose complex, based upon the conversion of Fe3+ to Fe2+ by breaking the iron sucrose complex without the addition of reducing agent.
BACKGROUND OF THE INVENTION
Iron carbohydrate complexes, administered either orally or parentrally, are used for the treatment of anemia due to iron deficiency. Iron sucrose injection is widely used in treatment of the iron deficiency and iron deficiency anemia and patients undergoing chronic Hemodialysis receiving supplemental erythropoietin therapy.
Iron sucrose injection replenishes body iron stores in patients with iron deficiency. Iron is a mineral that the body needs to produce red blood cells. When the body does not get enough iron, it cannot produce the number of normal red blood cells needed to keep person in good health. This condition is called iron deficiency (iron shortage) or iron deficiency anemia. Iron is sometimes lost with slow or small amounts of bleeding in the body that person would not be aware of and which can only be detected by your doctor. A doctor can determine if iron supplement is necessary.
Some conditions may increase your need for iron. These include bleeding problems, burns, hemodialysis, intestinal diseases, stomach problems, stomach removal, use of medicines to increase red blood cell count, etc.
Iron supplements are available in the following dosage forms:
Oral: Ferrous Fumarate, Ferrous Gluconate, Ferrous Sulfate, Iron-Polysaccharide
Parenteral: Iron Dextran, Iron Sorbitol, Iron Sucrose, Sodium Ferric Gluconate Complex


Now a days iron sucrose complex is used orally or parentrally for the treatment of iron deficiency anemia in patients. When iron-sucrose complex is given orally it will not be absorbed 100% from the GI tract. Hence, the absorbed iron-sucrose complex given orally is not adequate to stock up or maintain iron stores necessary for hematopoiesis during erythropoietin therapy.
Parenteral iron therapy in combination with erythropoietin has been shown to be successful and economic method to treat iron deficiency anemia in patients with chronic kidney disease (CKD).
To have high availability in the conditions like chronic hemodialysis, iron sucrose is given through intravenous route. Iron sucrose is taken up by cells of the reticuloendothelial system, which release ionic iron that binds to transferrin, which, in turn, transfers it to the bone marrow for erythropoiesis or to ferritin and the iron storage pool in the marrow, spleen and liver.
Thus in the human body, the metabolism of iron involves a series of reactions wherein the valence of the iron changes from Fe3+ to Fe2+ and vice versa.
US6911342 claims in vitro method to control and monitor the batch-to-batch bioequivalence of iron-sucrose complexes, by measuring the colloidal ferric hydroxide's rate of reduction from trivalent iron to divalent iron. In the method, iron sucrose complex is treated with a reducing agent and T75 for reduction kinetics of the complex is measured, wherein the T75 of less than 20 minutes indicates an effective bioequivalence of iron in the complex.
METABOLISM OF IRON SUCROSE
Iron sucrose is dissociated into iron and sucrose by the reticuloendothelial system and iron is transferred form the blood to a bone marrow. Ferritin, the iron storage protein binds and sequesters iron into a nontoxic iron that is easily available. The iron binds to
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plasma transferrin which carries iron through the extracellular fluid for supply to the tissues. The transferrin receptors presented in membrane binds transferrin iron complex which is then internalized in vesicles. Further, iron is released within the cell and transfemn-receptor complex returns to the cell membrane. Transferrin without iron is then released to the plasma. The intracellular iron becomes hemoglobin on circulating red blood cells.
When the amount of available iron exceeds ferritin's iron storage mechanism, an aggregated ferritin called hemosiderin is formed, which is a normal constituent of the monocyte-macrophage system. Hemosiderin is composed of molecules of ferritin, which have lost part of their protein shell and become aggregated. Hemosiderin accounts for about one third of normal iron stores and accumulates as insoluble granules in the cells of the reticuloendothelial system.
Upon administration to a patient, an iron sucrose complex is removed from the blood stream as a particle by the macrophages of the reticuloendothelial system and metabolized to replenish the body's iron stores of hemosiderin, ferritin and transferrin. The rate of removal from the blood stream is dependent on both the colloidal ferric hydroxide's particle size and composition.
Iron sucrose complex is composed of colloidal ferric hydroxide particles as core in complex with sucrose.
It is stated in US 6911342 that the colloidal ferric hydroxide complexes are dark red to brown solutions with a strong adsorption band at 450 nm. As the reduction to ferrous hydroxide occurs, the color is discharged, resulting in a decrease in absorbency. This decay (or dissociation) can be easily monitored in a temperature controlled (37+ldegree. C.) system.
In US 6911342, T75 time for the reduction of the iron-carbohydrate complex is used to determine the relative bioequivalence by reducing the complex with an appropriate
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reducing agent. A preferred bioequivalence standard for an iron-sucrose formulation is met if T75 reduction time is not more than 20 minutes and its reduction reaction plot of "Log (% Trivalent Iron Concentration)" versus "Time" is linear with a correlation coefficient absolute value of not less than 0.98.
The present invention is to achieve the same bioequivalence standard without addition of reducing agent.
OBJECTS OF THE INVENTION
First object of the present invention is to provide an in vitro bioequivalence method for iron sucrose without addition of reducing agent.
Another object of the present invention is to determine the kinetics for the conversion of Fe+3 to Fe+2 and comparing these kinetics to the reduction kinetics of a standard composition of known bioequivalence.
SUMMARY OF THE INVENTION
The present invention is a method of bioequivalence assessment of iron in iron-sucrose complexes, particularly iron-sucrose formulations for routine Quality control (QC) testing.
According to the present invention, the method of bioequivalence assessment for iron in iron-sucrose complex does not involve addition of a reducing agent.
The present invention also includes method to identify batches of iron-sucrose complexes having substantially the same bioequivalence. The method includes preparing iron-sucrose complexes, determining the conversion kinetics of each batch of iron-sucrose complex and identifying batches of iron-sucrose complex that meet the reduction kinetics of a standard composition of known bioequivalence.