Composition and method for arresting blood flow and for forming a persistent microbial barrier

Abstract

A composition and method useful in promoting healing of a bleeding wound site. The composition preferably includes a substantially anhydrous acid form of a cation exchange resin, which when applied over blood, provides an antimicrobial against planktonic microorganisms and biofilms in the wound. The resin is also capable, when applied in sufficient quantities, of providing a continuing and persistent antimicrobial against planktonic microorganisms and biofilms through dehydration and ion exchange with cations present in the blood and other body fluids. When the resin has a concentration of at least 26 mg/ml, it provides a >3 log reduction in biological activity of MRSA, MRSE and Pseudomonas aeruginosa.

Claims

1. A method of promoting healing of a wound having human blood comprising the steps of: providing a seal-forming composition consisting of a substantially anhydrous acid form of a cation exchange resin that, when forming a seal with the wound, results in the seal having a pH less than 3; applying an amount of the seal-forming composition to the human blood, said amount sufficient to form a microbial barrier seal over the wound; and maintaining the seal-forming composition in place over the wound for a time sufficient to allow formation of a reservoir of the seal-forming composition to promote healing of the wound.

2. A method of promoting healing of a wound having human blood comprising the steps of: providing a seal-forming composition consisting of a substantially anhydrous acid form of a cation exchange resin that, when forming a seal with the wound, results in the seal having a pH less than 3; applying an amount of the seal-forming composition to the wound, said amount sufficient to form a microbial barrier seal over the wound; maintaining the seal-forming composition in place over the wound for a time sufficient to allow formation of a reservoir of the seal-forming composition to promote healing of the wound; and discarding excess seal-forming composition that is above the microbial barrier.

3. The method of promoting healing of a wound having human blood as set forth in claim 1 wherein the seal-forming composition is applied to the wound in an amount to provide a resin concentration of at least 26 mg of acidic cation exchange resin per ml of blood.

4. The method of promoting healing of a wound having human blood as set forth in claim 2, wherein the seal-forming composition is applied to the wound in an amount to provide a resin concentration of at least 26 mg of acidic cation exchange resin per ml of blood.

Description

DETAILED DESCRIPTION OF THE INVENTION

(1) The following is a brief explanation of how the acid form of a cation exchange resin provides hemostasis and antimicrobial action.

(2) 1. The dry resin absorbs four times its weight of water. In contact with blood, it absorbs the water in the blood quickly and concentrates the clotting factors to form a strong seal to stop bleeding. Likewise, a strong seal is formed in contact with serum or exudates.

(3) 2. The blood contains sodium, potassium, magnesium and calcium cations represented as M.sup.+ and M.sup.++, respectively. The cations exchange with the acid ion-exchange resin to yield protons.

(4) 3. M.sup.++R−H.fwdarw.R−M+H.sup.+

(5) 4. M.sup.+++R−H.fwdarw.R−M+2 H.sup.+

(6) a. The pH in the seal formed with blood falls to <3 with the liberation of protons.

(7) Microbes are 80 to 90% water and contain sodium and potassium cations. The resin is capable of absorbing four times its weight in moisture. When a microbe passes through the resin, the microbe loses critical moisture in a low pH environment and dies. The cations necessary for microbial viability are extracted from the microbe and exchanged by the resin for a proton. The combination is lethal.

(8) Microbes come in different shapes, but all have an exterior cell wall which provides structure to the organism. Without wishing to be bound, when the microbe is in contact with resin, there is an entropy driving force for cations to leave the microbe and enter the resin and be replaced with acidic protons. The driving force is analogous to an osmotic pressure difference, except the driving force is based on the cation concentration difference between the interior of the microbe and the interior of the resin. When this cation extraction occurs, the structure of the organism changes and its viability is compromised. Even after the resin has absorbed all the water it can, there is a continuing antimicrobial effect because the resin can still absorb potassium and sodium ions and destroy the microbe's reproduction capability. The mechanism of action is considerably different than described in the U.S. Pat. No. 6,187,347 patent and for weak acid antimicrobial agents such as sorbic acid and acetic acid.

Example 1

(9) The adhesiveness and strength of a seal formed by fresh human blood and a dry ion exchange resin—hydrogen form of sulfonated, 2% crosslinked polystyrene resin was evaluated. A 0.1 mL sample of fresh human blood was placed on a plastic boat with a one square inch circular template. The blood was spread evenly on the template. 400 mg of the dry resin was poured on top of the blood. After 90 seconds, the excess resin was discarded, after which a seal formed from coagulated blood and resin was observed. After scrapping with a 6 mm wide spatula to remove weakly adhered parts of the seal, 30 mg of the seal was retained, indicating that a strong and adhesive seal was formed.

Example 2

(10) The in vitro antibacterial activity of a dry ion exchange resin (hydrogen form of sulfonated, 2% crosslinked polystyrene resin) was evaluated. Twice washed liquid cultures were resuspended in sterile water to provide a concentration of approximately 10.sup.8 CFU/ml for each strain. The liquid cultures were added to 50 ml conical tubes containing the dry resin that had been pre-weighed to produce the desired concentration upon addition of 1 mL of bacterial suspension. The samples were vortexed and allowed to stand undisturbed for 30 minutes at room temperature. Each sample was serially diluted and quantitative culture on Luria-Bertani (LB agar) was performed. The hydrogen resin demonstrated significant bactericidal activity. No organisms were recovered by quantitative culture at exposure levels of 5 mg/mL. Dose responses were seen. The estimated dose required to effect 99% killing of three bacteria are presented in Table 1 below.

(11) TABLE-US-00001 TABLE 1 Mg/mL of hydrogen form of sulfonated 2% crosslinked polystyrene resin (hydrogen resin) required for 99% bacterial killing Organism Mg/mL of hydrogen resin Pseudomonas aeruginosa PA7 0.83 ± 0.04 Staphylococcus epidermidis MRSE 1.87 ± 0.08 (ATCC #700565) Staphylococcus aureus MRSA 4.22 ± 0.26 (ATCC #43300) All values reported as mean ±standard error. Thus, the hydrogen resin is a powerful antibacterial agent against at least two of the most antibiotic resistant bacteria commonly occurring in hospital settings, i.e., MRSA and MRSE.

Example 3

(12) This example demonstrates that the same hydrogen resin mentioned in Example 2 has persistence in vitro antibacterial activity. Hydrogen resin was added to 10 mL of Luria-Bertani (LB) media and allowed to hydrate for 30 minutes with mixing. Single bacterial colonies were added and placed in an incubator at 37° C. and 200 RPM for 8 hours. Bacterial proliferation was measured as the change in sample turbidity during incubation. The hydrogen resin inhibited bacterial growth at both 1 and 5 mg/mL as measured by liquid culture turbidity. The inhibition was significantly greater at 5 mg/mL compared to 1 mg/mL (see Table 2 below).

(13) TABLE-US-00002 TABLE 2 Persistency of in vitro antibacterial activity of hydrogen form of sulfonated 2% crosslinked polystyrene resin (reported as reduction in bacterial colonies) Organism 1 mg/mL 5 mg/mL Pseudomonas aeruginosa PA7 80% reduction 95% reduction Staphylococcus epidermidis MRSE 30% reduction 99% reduction (ATCC #700565) Staphylococcus aureus MRSA 15% reduction 99% reduction (ATCC #43300)

Example 4

(14) The ability of hydrogen form of sulfonated 2% crosslinked polystyrene resin (hydrogen resin) to eradicate biofilm was studied with the Calgary Biofilm Device (CBD) (reference—Laila Ali et al., “Investigating the suitability of the Calgary Biofilm Device for assessing the antimicrobial efficacy of new agents”, Bioresource Technology 97 (2006) 1887-1893). The CBD assay was developed by the University of Calgary as a simple assay to reliably culture 96 identical biofilms at a time. The CBD assay provides rapid testing of compounds for anti-biofilm activity. The hydrogen resin was evaluated for anti-biofilm activity against MRSA, MRSE and Pseudomonas aeruginosa using the CBD. The results are summarized in Table 3 below.

(15) TABLE-US-00003 TABLE 3 Biofilm eradication by the hydrogen form of sulfonated 2% crosslinked polystyrene resin (hydrogen resin) Hydrogen resin concentration in mg/mL Pseudomonas Microorganism MRSA MRSE aeruginosa MIC breakpoint ≤26 ≤26 ≤26 MBC breakpoint ≤26 ≤26 ≤26 MBEC breakpoint ≤26 ≤26 53 Minimum concentration that kills 26 26 26 biofilms with logR ≥ 3 Legend: MRSA = Methicillin-Resistant Staphylococcus aureus 399 MRSE = Methicillin-Resistant Staphylococcus epidermidia ATCC 35984 P aeruginosa = Pseudomonas aeruginosa ATCC 27853 MIC—minimum inhibitory concentration against planktonic microorganism MBC—minimum bactericidal concentration against planktonic microorganism MBEC—minimum biofilm eradication concentration Breakpoint—concentration at or below logR—log reduction

(16) The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) breakpoints against all three planktonic microorganisms are the same at 26 mg/mL. The minimum biofilm eradication concentration (MBEC) breakpoint is surprisingly the same at 26 mg/mL for MRSA and MRSE and about twice the concentration at 53 mg/mL for Pseudomonas aeruginosa.

(17) When biofilms are formed, they are difficult to remove as they show an increased resistance to biocides and antibiotics when compared to planktonic microorganisms. Studies have shown greater than a hundred to a thousand fold resistance to antibiotics of biofilms compared to the same bacteria in planktonic state. Therefore the identical or similar concentration breakpoints between MBEC and MIC/MBC for hydrogen resins are very significant.

(18) The results suggest that the hydrogen resin is able to rob the water from the biofilm, that ion exchange is proceeding to free up the proton, and that the proton is able to penetrate through the complex matrix of the biofilm to inactivate and destroy it. The concentration of hydrogen resin required to provide a .gtoreq.3 log reduction in activity of the three biofilms is equal at 26 mg/mL. At 26 mg/mL, hydrogen resin provided 3.2 log reduction of MRSA biofilm, 3.5 log reduction of MRSE biofilm and 4.5 log reduction of Pseudomonas aeruginosa biofilm.

Example 5

(19) Eight concentrations of dry hydrogen resin powder were studied in the Calgary Biofilm Device (CBD) as shown in Table 4 below. This was achieved by weighing the indicated weights into different wells of the CBD. Next, a precise amount (microliter, ul) of saline was pipetted into the corresponding wells of the CBD following the quantities specified in column 1 of Table 4. The resulting concentrations in mg/mL are shown in column 2.

(20) TABLE-US-00004 TABLE 4 The ability of hydrogen form of a sulfonated 2% crosslinked polystyrene resin to eradicate biofilm Pseudomonus mg/ul H.sup.+ mg/mL H.sup.+ MRSA MRSE aeruginosa resin/saline resin/saline Log Reduction 160/0  pure powder, no saline 3.2 3.48 5.98 75/106 745 3.2 3.48 5.63 50/138 384 3.2 3.48 5.63 38/153 255 3.2 3.48 5.98 25/169 154 3.2 2.98 5.98 15/181 88 2.85 3.48 5.98 10/189 58 3.2 3.48 5.98  5/194 26 3.2 3.48 4.48

(21) At the lower concentrations (26, 58, 88 and 154 mg/mL), the hydrogen resin has fully absorbed the saline and excess saline is available outside the swelled resin phase. The excess saline is greatest at 26 mg/mL and lowest at 154 mg/mL. At 255 mg/mL, the resin is fully absorbed with saline with no excess saline is available. At 384 mg/mL, the resin is only hydrated to 69% assuming the resin is uniformly hydrated. At 745 mg/mL, the resin is only 35% hydrated. The extreme case is where the well contains only 160 mg resin with no saline. The assay was run in triplicate. When challenged with MRSA biofilm (see column 3) or

(22) MRSE biofilm (column 4), the whole concentration range of hydrogen resin showed a log reduction close to 3 or higher. When challenged with Pseudomonas aeruginosa biofilm (column 5), the lowest concentration of 26 mg/mL gave a log reduction of 4.48 while the rest gave log reductions of 5.6 or higher.

(23) The results indicate that hydrogen resin eradicates biofilm very efficiently even at a very low concentration of 26 mg/mL. Furthermore, the data suggest that excess capacity or persistency for biofilm eradication is available at higher concentrations. In particular, at the concentrations where the hydrogen resin is not fully hydrated, there is excess capacity or persistency to extract water and cations from any new biofilm that may form after the first wave of biofilm has been destroyed. At the extreme, a completely dry hydrogen resin provides the greatest capacity for repeatedly killing biofilms, increasing its persistency.

(24) This finding can be applied to a bleeding or exuding wound from a vascular access procedure or percutaneous catheters and tubes where excess dry hydrogen resin is applied to stop bleeding or to form a strong seal around wounds. Once the powder dressing is applied, a secondary dressing is applied over the powder dressing to keep the resin above the seal dry as a reservoir for preventing biofilms from forming and for continually eradicating planktonic cultures and biofilms. The persistency for biofilm eradication is determined by the height and quantity of the reservoir of dry hydrogen resin available on the wound site.

ALTERNATE EMBODIMENTS

(25) In another embodiment, a powder containment device (PCD) is employed to build the height and to contain the dry hydrogen resin powder within the device. In another embodiment, the resin is applied over minor-to-severely bleeding wounds such as surgical wounds including post-operative, post-suturing, donor sites and dermatological wounds to stop bleeding as well as to provide a microbial barrier to prevent infection. In yet another embodiment of the invention, the resin is used for managing exuding wounds such as pressure ulcers, venous ulcers, diabetic ulcers and arterial ulcers taking advantage of its microbial barrier properties. In yet another embodiment, the optional inclusion of other cationic forms (such as silver and alkali metal cations and quarternary ammonium cations) of the ion exchange resin with the hydrogen form of the resin may further extend its antimicrobial properties.

(26) Other embodiments of this invention include the optional presence of other substances such as hydrophilic inorganic or organic polymers, clays, gums, natural polysaccharides, oxidized cellulose, regenerated cellulose, chitosan and the like that, when added to the protonic ion exchange resin, can impart adhesive and strengthening properties to the seal formed with blood.

(27) While a number of exemplary aspects and embodiments have been discussed above, those of skill in the art will recognize certain modifications, permeations and additions and subcombinations thereof. It is therefore intended that the following appended claims and claims hereinafter introduced are interpreted to include all such modifications, permeations, additions and subcombinations that are within their true spirit and scope.

(28) All patents, patent applications, provisional applications, and publications referred to or cited herein are incorporated by reference in their entirety, including all figures and tables, to the extent they are not inconsistent with the explicit teachings of this specification.

(29) It should be understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and the scope of the appended claims. In addition, any elements or limitations of any invention or embodiment thereof disclosed herein can be combined with any and/or all other elements or limitations (individually or in any combination) or any other invention or embodiment thereof disclosed herein, and all such combinations are contemplated with the scope of the invention without limitation thereto.