Patent Application
20250127952
The present invention relates to the field of polymeric matrix for haemostatic application. In particular, the Invention provides an efficient polymeric haemostatic device/bandage with high swelling rate, enhanced mechanical property and high blood clotting ability.
The following background discussion includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.
Most bleeding casualties arise within the first 30 minutes of the injury leading to high patient mortality which can be reduced by effective and especially with immediate action. Besides controlling bleeding quickly, haemostatic products that are ready-to-use, easy to apply for on-site responders in trauma situations, antiseptic and has high shelf life are generally accepted for forward care in the battle zone. Although, the advancement in material science has led to the development of various promising dressing, only a few made it to clinical translation.
In battlefields and trauma cases, uncontrolled haemorrhages and its associated severe complications remain one of the major factors in both defense and civilian deaths. From many decades, the mortality rate has increased due to continuous loss of blood which can be prevented by an ideal haemostatic dressing that forms a stable clot as well as stops excessive bleeding.
In the past, compression with gauze was a widespread practice for stopping blood loss in most injuries. Development of some new generation materials had already overcome some of the prevailing limitations, but lack of rapid coagulation and insufficient provision limited its more extensive application. These are also expensive, unable to curtail microbial infection and may lead to allergic reactions. The currently available haemostatic dressings (materials that prevent blood loss) are also not efficient enough to stop excessive bleeding.
A wide variety of materials are used at an individual level for haemostasis but fail in clinical use as they lack to fulfill all requirements. As elaborated above, many strategies for developing the haemostatic material have been devised using specific technologies like electrostatic spinning, emulsion polymerization, layer-by-layer self-impregnation, etc. The use of such methods has solved the problem of developing the composite materials to some extent; however, issues like flexibility, blood absorption rate, coagulation time, cost, incorporation of haemostatic agent, etc. are worth to mention which still persist. Many of such materials individually/composites have proven less effective, inadequate, expensive, associated with severe complications in in-vivo studies.
CN104208741A discloses adhesive bandage based on chitosan, oligochitosan, and enhancing polymer, wherein the chitosan and oligochitosan accounts for 85 to 95% of the total weight of the adhesive bandage, and the enhancing polymer is one or more of sodium alginate, glucan, hyaluronic acid polymer, and collagen.
U.S. Pat. No. 9,950,091B2 discloses composition based on a hydrogel matrix that includes at least one polymer cross linked, via ionic or covalent bonding, with both hyaluronic acid and alginic acid. The polymer is chitosan, poly L-Lysine, or a combination thereof.
However, the technologies disclosed in US '0091 and '741 are mainly based on chitin which is not efficient in stopping the heavy bleeding and there is still a need of efficient and better mechanisms and related product on the same.
U.S. Pat. No. 4,822,349 describes a non-bandage material used to treat bleeding. The material is sold by Z-Medica as “Quick-Clot” and is a granular form of zeolite, an aluminum silicate mineral. While use of this material may be preferable to bleeding to death, the attendant burning of tissue at and near the wound (and possible burn injury of medical personnel who are administering the material) is clearly a severe disadvantage. This side effect also reduces the ability of the material to be used for internal hemorrhage.
No perfect solution currently exists for the treatment of excessive or severe bleeding.
Many haemostatic products have been synthesized and are commercially available, presenting an approach of art as well as science to choose dressings over one another. However, the primary FDA approved haemostatic dressings like dry fibrin sealant dressing (DFSD), those based on collagen, chitosan, semcitite and zeolite (HemCon, QuikClot, WoundStat, etc) have significantly promoted rapid and effective hemostasis, but majority of them fail to fulfill the needed ideal requirements. The major drawbacks of these dressings are expensive, availability, non-antiseptic, and lack of rapid coagulation and may lead to allergic reactions. So, there is an immediate need to develop a dressing material which can have a high haemostatic ability and should protect the wound from secondary bacterial infections. It has also been observed that various haemostatic agents impregnated in the developed material or gauze leach-out into blood vessels, leading to severe vascular clots that may eventually become deadly.
Hence, there is a need of an efficient, cost effective and flexible bandage with high blood absorbing and antimicrobial capacity. There is also needed an efficient method of production of the flexible bandage with advantageous properties of quick blood clotting, high blood absorbing and antimicrobial capacity.
Primary object of the present invention is to overcome the limitation of prior art.
Another object of the present invention is to provide an efficient polymeric bandage for haemostatic application.
Another object of the present invention is to provide an efficient polymeric haemostatic device/bandage with high swelling rate, enhanced mechanical property, enhanced absorption rate and high blood clotting ability.
Another object of the present invention is to provide an efficient polymeric haemostatic device/bandage based on naturally derived polymers comprising chitosan and dextran.
Another object of the present invention is to provide an efficient polymeric haemostatic device/bandage based on cryogelled polymers.
Another object of the present invention is to provide a cryogel composition with high absorption rate and rapid blood clotting ability.
Another object of the present invention is to provide a cost effective bandage with prolonged shelf life in lyophilized form.
Another object of the present invention is to provide a method, for preparing the cryogel composition, which combines biocompatible and haemocompatible polymer-based materials with other topical haemostatic agents.
Another object of the present invention is to provide a method based on cryogelation for obtaining the polymeric bandage for haemostatic application.
In an aspect, there is provided a cryogel composition for a hemostatic bandage comprising:
In another aspect, there is provided a method of surface modification of dextran which comprises oxidation of hydroxyl groups at adjacent carbons of glucose units by sodium periodate (NaIO4) leading to the formation of two aldehydic groups per glucose unit due to the cleavage of bond between the adjacent carbons containing hydroxyl groups and the formation of multifunctional dextran with high reactive groups for coupling with molecules having amines.
In another aspect of the Invention, there is provided a process of preparing the composition as described above, comprising the steps of:
In another aspect, there is provided a therapeutic bandage comprising a base layer embedded with a polymeric matrix of cryogel composition as described above, wherein said cryogel composition comprises:
To further clarify advantages and features of the present invention, a more particular description of the invention will be rendered by reference to specific embodiments thereof, which is illustrated in the appended drawings. It is appreciated that these drawings depict only typical embodiments of the invention and are therefore not to be considered limiting of its scope. The invention will be described and explained with additional specificity and detail with the accompanying drawings in which:
For the purpose of promoting an understanding of the principles of the invention, reference will now be made to the embodiment illustrated in the drawings and specific language will be used to describe the same. It will nevertheless be understood that no limitation of the scope of the invention is thereby intended, such alterations and further modifications in the illustrated system, and such further applications of the principles of the invention as illustrated therein being contemplated as would normally occur to one skilled in the art to which the invention relates.
It will be understood by those skilled in the art that the foregoing general description and the following detailed description are exemplary and explanatory of the invention and are not intended to be restrictive thereof. Throughout the patent specification, a convention employed is that in the appended drawings, like numerals denote like components.
The present invention provides a cryogel composition and highly efficient polymeric bandage based on the same for haemostatic application especially during excessive bleeding in case of battlefields and civilian trauma. The invention is efficient in stopping excessive blood loss and will be useful to common public and defense people. The bandage will act as a first-hand emergency bandage in critical conditions (war, trauma), where seconds matter to save a precious life.
The Invention provides an efficient polymeric haemostatic device/bandage with high swelling rate, enhanced mechanical property and high blood clotting ability. This polymeric bandage works synergistically to maintain haemostasis by forming a stable clot, besides accelerating the clot formation. The bandage has high blood absorption rate besides efficient in rapid clotting. The haemostatic device/Bandage is based on cryogelation technology and a new composition to stop excessive bleeding, specifically critical and deadly bleeding in trauma care and battlefields. The cryogelation will help to provide a well interconnected macroporous network within the bandage, necessary for rapid fluid uptake and high swelling rate. The use of cryogelation technology further ensures proper integration of multiple haemostatic agents into the bandage for efficient and rapid clotting of blood. In addition, the technology used for fabrication of this invention substantially stopped the leaching of haemostatic agents from the material during practical application thereby avoiding any chances of risks due to systemic thrombosis and emboli formation. The in-vitro and in-vivo evaluation of the developed haemostatic bandage showed best results with rapid blood coagulation and good antiseptic properties.
The cryogel composite showed high absorption rate and rapid blood clotting, an important property of an ideal haemostatic dressing. The bandage is flexible, ready to use that can be cut in various sizes for distinct wounds and has high shelf life.
In an embodiment, the composite haemostatic bandage prepared from polymeric materials i.e. chitosan and oxidized dextran incorporated with other topical agents (kaolin and calcium chloride) using cryogelation technology. The chitosan based bandage not only controls bleeding quickly but also have a high blood uptake capacity. This helps to eliminate the need for frequent changing of the dressing. The bandage has optimum swelling and mechanical properties, which is enhanced further by incorporation of cotton mesh. Further, the bandage is antiseptic due to the inherent antimicrobial property of the incorporated polymer and has a high shelf life. The developed haemostatic matrices are able to stop bleeding quickly without having any issues of biocompatibility.
In an embodiment, the bandage comprises a cotton mesh incorporated within the polymeric cryogel, that enhances its mechanical stability and flexibility along with controlling the swelling of the bandage. Besides, being able to stop bleeding quickly, the chitosan-based composite is non-cytotoxic and inexpensive as compared to other available products.
The composite haemostatic bandage is prepared from chitosan and tailored the properties of dextran, incorporated with kaolin and calcium chloride as the potent haemostatic agents. The Invention uses specific cryogelation technologies, to combine the biocompatible and haemocompatible polymer-based materials with other topical haemostatic agents, which can directly activate the coagulation cascade to improve the haemostatic potential. The method of fabrication has led to an interconnected porous structure with optimum pore size, high fluid absorption rate, and very rapid blood clot formation.
These advantageous properties of the invention directly depends on certain major fabrication steps, mainly the appropriate concentration of polymers and haemostatic agents besides using a desired crosslinking agent. The porous interconnectivity as well as the mechanical property is highly dependent on the concentration of ingredients and temperature of fabrication. To have a desired bandage for ideal haemostatic applications, time of incubation during cryogelation also plays a critical role. The claimed invention has customized these parameters to achieve the best result.
In an embodiment, the bandage is based on polymers (chitosan and oxidized dextran) and haemostatic agents (comprising kaolin and calcium chloride) in the cryogel bandage. Kaolin, a natural clay consists of a network of aluminosilicate units, giving a net negative surface charge that plays a significant role mainly in activating the factor XI and XII thereby formation of fibrin. Calcium ions play a critical role in blood clotting, they are important in the activation of many clotting factors of multistep coagulation cascade as well as are involved in the formation of fibrin clot by triggering the production of thrombin.
In an embodiment, the Invention provides a polymeric bandage based on non-limiting biocompatible materials comprising chitosan (C) and dextran (D). The biocompatible materials are cryogelled at the sub-zero temperature using cryogelation technology.
In an embodiment, the cryogel composition for a hemostatic bandage comprises:
In an embodiment, the composition comprises approx. Ig chitosan, 2.5 g of Kaolin, 0.5 g of CaCl2, approx. 3 g of oxidised dextran.
In an embodiment, the dextran is oxidized through the surface modification.
In an embodiment, the surface modification of dextran comprises oxidation of hydroxyl groups at adjacent carbons of glucose units by sodium periodate (NaIO4) leading to the formation of two aldehydic groups per glucose unit due to the cleavage of bond between the adjacent carbons containing hydroxyl groups and the formation of multifunctional dextran with high reactive groups for coupling with molecules having amines.
In an embodiment, the dextran is partially oxidized in the presence of an aqueous solution of sodium periodate by the steps of:
In an embodiment, the dextran is partially oxidized in the presence of an aqueous solution of sodium periodate by the steps of:
In an embodiment, there is provided a process of preparing the composition as described above. The process comprises the steps of:
In an embodiment, the therapeutic bandage comprising a base layer embedded with a polymeric matrix of cryogel composition as described above. The cryogel composition comprises:
In an embodiment, the therapeutic bandage comprises approx. 1 g chitosan, 2.5 g of Kaolin, 0.5 g of CaCl2, approx. 3 g of oxidised dextran.
In an embodiment, the base layer is made of material comprising cotton or material with alike properties.
In an embodiment, the bandage is prepared by following steps:
In an embodiment, the composite was fabricated by dissolving chitosan (1 g) in 60 ml of acetic acid (1%) for 4-6 h. Next, 2.5 g of Kaolin and 0.5 g of CaCl2 were mixed in 15 ml of mQ-water. After proper mixing, the solution was added to the chitosan solution and was kept on rocker to allow complete homogenization. In addition, 3 g of oxidised dextran was dissolved in 25 ml of mQ-water at 70° C. with continuous stirring for 30 min. The two solutions were then pre-cooled separately at 4° C. for half an hour. After cooling of the solutions for desired time, the dextran solution was added to the other solution (containing chitosan, Kaolin and CaCl2) and were mixed appropriately followed by pouring of the solution in the desired mould having thin cotton membrane. The mould was then allowed for cryogelation by keeping it in a cryostat at −12° C. for 14 h. The synthesized cryogel columns/sheets were then given freeze thaw cycles to remove all the unreacted polymers and increase their strength followed by lyophilization for their proper storage.
In an embodiment, the synthesis happens at sub-zero temperature comprising −12° C. for 14 h followed by freeze-thawing cycle. The optimized temperature of −12° C. for 14 h allows to have a desired crosslinking within the polymers, forming mechanically flexible and stable haemostatic bandage. The developed haemostatic bandage at sub-zero temperature resulted to the formation of macroporous structure with interconnected porous architecture that is essential for rapid and efficient fluid uptake without getting disintegrated or collapse during practical application. The freeze thaw cycle aids in achieving proper pore structure and in mechanical stability of the haemostatic bandage.
In order to make the bandage robust, proficient and cost effective, kaolin (K) and calcium chloride (Ca) as haemostatic agents were incorporated in the developed bandage that provides a synergistic effect in controlling excess bleeding. These components of fabricated matrix cause occlusion by protein precipitation or coagulation (cell entrapment), while others act on later steps of the clotting pathway thus initiating a biological response to stop bleeding. The polymer based cryogel bandage (CDKCa) is formed in various moulds to get appropriate shape for specific applications at distinct sites of injuries.
The physical properties of the developed haemostatic bandage were evaluated to examine its suitability for quick control of bleeding. The CDKCa haemostatic bandage showed very high absorption and swelling rate both in water and simulated body fluid. The property is very significant for an ideal haemostatic dressing and is attributed to the interconnectivity of pores in the developed cryogel sheet. Furthermore, CDKCa bandage showed rapid blood coagulation (in vitro) in the 30 s as compared to other chitosan based compositions as well as commercially available QuikClot. The CDKCa will come in contact with blood and thus was tested for haemolysis. The results showed the material is heamocompatible. Mechanical strength is a very important factor in a haemostatic dressing, and in the developed polymeric composite, it can be easily modulated by varying the concentration of the polymers or the temperature of synthesis. The fabricated CDKCa composite system and the bandage based on it exhibits a rapid and strong effect as a haemostatic dressing, which is highly efficient over the commercially available products.
The surface modification of dextran comprises the oxidation of hydroxyl groups at adjacent carbons of glucose units by sodium periodate (NaIO4). This reaction leads to the formation of two aldehydic groups per glucose unit due to the cleavage of bond between the adjacent carbons containing hydroxyl groups. The NaIO4 based modification of dextran leads to the formation of multifunctional dextran with high reactive aldehydic groups for coupling with molecules having amines. In this invention the functionalized dextran forms Schiff base by reacting with amine groups present in the chitosan.
In an embodiment, the product is developed as a single unit in the form of cryogel patch/bandage using cryogelation process. The cryogelation technique makes the fabrication process easy and cost effective. The size and format of this can be easily modified depending on the desired application. The developed bandage has interconnected porous architecture leading to rapid fluid uptake as well as high swelling ability. The invention led to rapid blood clotting during excessive bleeding. The fabrication method of cryogelation does not any leaching of material ingredients from the bandage thereby preventing the risk of forming thrombosis or systemic emboli.
The Invention is further described with the help of non-limiting examples:
The haemostatic bandage comprises naturally derived polymers chitosan and dextran. The dextran is oxidized through the surface modification thereby allowing it to be used as a crosslinker as well as haemostatic agent due to its negative charge. The invention employs cryogelation technology in the fabrication of haemostatic bandage from hydrophilic polymers and eases while incorporating the haemostatic agents Kaolin and Ca2+ (from calcium chloride). The use of cryogelation technology not only helped to overcome the unwanted release of haemostatic agents at the injury site but also maintained the uniformity of fine porous structure, mimicking to the naturally occurring fibrin mesh. The porous network of the developed haemostatic device leads to fast swelling rate and entrapment of platelets and red blood cells thereby effectively reducing the coagulation time. In addition, the present invention provides a unique composition of the components in the form of developed haemostatic bandage with highly efficient and improved haemostasis. In particular, the composition of the inventive haemostatic bandage has been further embedded with the cotton membrane at the time of synthesis to present a flexible and mechanically robust bandage in order to assure its application at any unique geometrical shape and size.
Furthermore, this composition has led to the haemostatic bandage which is user friendly, less expensive, long shelf life, no exothermic reaction when applied. The cumulative and synergetic effect of these polymeric components i.e. chitosan and oxidized dextran with incorporated haemostatic agents i.e. Kaolin and Ca2+ in the developed material has shown the ability to rapidly form a stable fibrin clot. The clotting occurs due to surface charge interaction i.e. positive charge of chitosan with platelets and Ca2+ ions, a crucial role in activating clotting factors of the cascade, thrombin formation and finally role in the conversion of fibrinogen into fibrin. The haemostatic property is also efficiently improved by the negatively charged surface of oxidized dextran and kaolin through the stimulation of intrinsic coagulation cascade via autoactivation of clotting factor XII, which has a direct role in fibrin generation.
The fabricated dressings for haemostatic application have been evaluated for various physio-chemical properties as well as in both in-vitro and in-vivo studies. The experimental analysis was carried out and studied using specific samples.
Different types of polymer (chitosan)-based cryogel sheets that can be used as ideal haemostatic dressings were formed. These cryogel materials have been fabricated using different natural polymers and inorganic compounds. The materials used exhibit distinct advantages in stability, accessibility and cost efficiency with no biological toxicity or risk of disease transmission. Chitosan has been used as a haemostatic material due to its cationic and antimicrobial nature, besides being biocompatible in nature. Dextran is a hydrophilic polysaccharide and can easily be configured by altering its surface functionality. It has been used in the haemostatic application for both civilian and combat trauma. Oxidation of dextran will help both in crosslinking as well as in blood coagulation. So, initially, dextran was subjected to partial oxidation in the presence of an aqueous solution of sodium periodate to generate aldehyde functionalities. The generated aldehyde groups help in the crosslinking of chitosan in the fabricated bandages. The amount of aldehyde generated was calculated around 30%. The oxidized dextran was then used to fabricate the chitosan based cryogel sheets in different formats with and without haemostatic agents (Kaolin/Calcium chloride). Kaolin and Calcium chloride as a haemostatic agent has shown the efficacy in the stabilization of severe arterial hemorrhage and diffuse long-term bleeding from lethal liver injuries along with other materials. Kaolin also called as Chinese clay consists of aluminosilicate network that leads to the activation of blood coagulation cascade by stimulating the intrinsic pathway. Also, the negative charge on Kaolin surface triggers binding of factor XII, its autoactivation along with factor XI and their direct role in fibrin formation.
The approach towards the development of an ideal haemostatic dressing is very critical, and currently, cryogelation technology is one of the promising approaches for material fabrication. The cryogel sheets were developed using various polymers such as chitosan, gelatin, polyurethane, etc. with or without haemostatic agents during the optimization process. The various combinations of cryogel sheets developed, characterized and evaluated for blood clotting study were: (1) Chitosan (C)-O-Dex (D)-Kaolin (K)-CaCl2) (Ca) composite (2) Gelatin Cryogel only (3) Chitosan-Gelatin (4) Chitosan only (5) Chitosan-O-Dex-Kaolin (6) Chitosan-Polyurethane (not formed). All the cryogel bandages were synthesized at sub-zero temperature (−12° C. for 14 hours) followed by freeze-thawing cycle, resulting in melting of the frozen solvent crystals, leaving behind a well interconnected porous network as confirmed by scanning electron microscopy as in
The bandage was made mechanically robust by embedding with cotton fibers to enhance its flexibility and mechanical property as in
Next, as a clotting indicative, the absorbance of haemoglobin (540 nm) was evaluated to test the haemostatic efficiency of developed composite bandage. Higher absorbance of haemoglobin indicates poor clotting while as low absorbance indicates better clotting. The developed bandage was examined for hemolytic analysis, carried out with separated human erythrocytes. Blood was drawn from volunteered human subjects by registered staff at the Department of Medicine and Transfusion, Govt. Medical College, Kanpur with blood samples were tested. Subjects were healthy adults ranging in age from 25 y to 30 y. Sodium citrate (3.8%) was used as an anticoagulant. The erythrocytes were separated as per the standard protocol, briefly; the samples were centrifuged at 1500 rpm for 10 min followed by the washing of pellet 3 times using 1× phosphate buffer saline (PBS, pH 7.2). The samples were incubated in 10 ml of blood: PBS (1:9) for 1 h at 37° C. and then measured at 540 nm post centrifugation. As shown in
As a functional assay, the developed materials were evaluated for blood coagulation test initially on goat blood (I and II) and later on human blood samples (III).
The influence of the developed CDKCa on blood clotting and to further observe the mechanism of heamostasis were evaluated by the effect of material interface on the adhesion of the platelets and red blood cells (RBC'S). The blood cells adhered on the material surface as observed by SEM images (
The animal studies were carried out on Sprague-Dawley (SD) rats (250-300 g) which were directed in accordance with the “Guide for the Care and Use of Laboratory Animals.” All the in-vivo experimental protocols were approved by the Indian Animal Ethical Committee of Indian Institute of Technology Kanpur—IAEC IITK (Protocol No. ITK/IAEC/2017/1061).
The animals anesthetized using 2.5% isoflurane along with 21% of oxygen support were shaved and sanitized by 70% ethanol at desired sites, and then the rats were fixed on Styrofoam board, tilted at 45 degrees approx. The Axiostat and QuikClot, commercially available FDA approved products, were used as a standard haemostatic dressing for evaluating our developed CDKCa bandage for haemostatic application.
The rats were divided into 5 groups (No treatment group: control; Treated groups: chitosan only, QuikClot, CDKCa, and Axiostat) with 6 animals per group. Briefly, the 5 mm deep liver puncture model was created by the 20-G needle on the left lateral liver lobe exposed by abdominal incision. A pre-weighed filter paper was kept under the liver lobe, and blood flow from the injury was allowed freely for 5 s before applying the dressings. The evaluation of haemostatic effect by the treatment groups was evaluated by bleeding time and determining the difference in the weight of filter paper used within the respective groups after a liver puncture (
The model was developed to mimic the field challenges of bleeding during trauma and battlefield. Also, initially bleeding from injured artery predominates but later venous bleeding becomes more appropriate due to mean pressure drop in the artery. The study of this femur model involves 6 treatment groups with 6 animals in each group. The 6 groups include control (no treatment group), gauze only, axiostat, QuikClot, chitosan only and CDKCa. All the animals were anesthetized and sterilized before any surgical procedure as mentioned in the above section. The femoral vein and artery and were exposed after making an incision in the left leg, and these exposed vessels were cut using a scalpel. A pre-weighed filter paper was immediately put under the injured leg to absorb the blood from the cut vessels prior to application of the bandage at the site. The total amount of blood loss accumulated on the filter paper and time to stop bleeding was recorded for efficient evaluation of haemostasis by the treating groups. The results of CDKCa group displayed rapid blood coagulation and a significant reduction in clotting time in comparison to other treated groups. In CDKCa group, the clotting was achieved quickly within 92 s without any further flow of blood to the filter paper (
| Number | Date | Country | Kind |
|---|---|---|---|
| 202111039585 | Sep 2021 | IN | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/IB2021/059807 | 10/27/2021 | WO |
