Electrospun janus wound dressing and associated methods

Abstract

A Janus wound pad which can be used as a wound dressing or wound closure is disclosed. The Janus wound pad comprises: a hydrophobic layer comprising electrospun strands of a hydrophobic material; and a hydrophilic layer comprising electrospun strands of a hydrophilic material. The Janus wound pad has an optical transmittance of at least 10%. The hydrophobic and hydrophilic layers may allow water at the wound side to be removed, while preventing water from external sources entering the wound. The optical transmittance may allow the wound to be properly aligned when the wound pad is being applied, and the wound to be monitored during the healing process.

Description

Electrospun Janus Wound Dressing and Associated MethodsTECHNICAL FIELD

[0001] The invention relates to wound closures and dressings and, in particular, to electrospun Janus closures and dressings with a hydrophobic side and a hydrophilic side.BACKGROUND

[0002] Skin provides a physical and chemical barrier between the outside environment and the inside tissues of the body. It is one of the most common medical problems that open wounds, such as cuts, lacerations, abrasions, high-temperature scalds, and diabetic ulcers, are difficult to heal, especially after being infected.

[0003] Wound closures and wound dressings are important components for optimizing healing by aligning the wound, protecting the wound, preventing contamination, and promoting absorption of wound exudate while permitting air flow to prevent wound hypoxia and bacterial infection.

[0004] Numerous types of wound closures are available in the market such as sutures, glue, sterile strips, and staples; however, they all have their own limitations.

[0005] For example, sutures are widely available for use in emergency medicine but require specialized equipment and skill sets and frequent follow-up management for removal utilizing additional health care resources. Further, the standard approach for closure using sutures can be traumatic for children and has limited applicability with the fragility of skin for elderly populations.

[0006] Numerous types of wound dressings are available in the market; however, they all have their own limitations.

[0007] For example, gauze wound dressings are inexpensive but can damage the wound site upon removal.

[0008] Semi-permeable films permit gas exchange, but they have a low absorption capacity. In contrast, foam dressings possess a larger absorption capacity; however, their long residence time leads to dryness of the wound area. Hydrogel dressings can maintain a moist microenvironment surrounding the wound, but they can have low mechanical stability and increase the risk of bacterial infections.

[0009] It remains a challenge to optimally close a wound and prepare effective wound dressings with good combinations of desired properties for wound healing. For instance, a leading commercial product, Tegaderm™ by 3M™, is a waterproof wound dressing, but the mechanical strength is only -0.25 MPa.SUMMARY

[0010] In accordance with the present disclosure, there is provided a Janus wound pad comprising: a hydrophobic layer comprising electrospun strands of a hydrophobic material; and a hydrophilic layer comprising electrospun strands of a hydrophilic material, wherein the Janus wound pad has an average transmittance (e.g., the mean average of the transmittance values for the different wavelengths) of at least 10% for wavelengths between 350-800nm (e.g., from 350 to 800 nm).

[0011] The Janus wound pad may have a two-dimensional mesh pattern comprising areas of relatively lower density of electrospun strands bounded by elongate portions of relatively higher density of electrospun strands. Each area of relatively lower density may be surrounded on all sides by elongate portions. The elongate portions may form a boundary around each area of relatively lower density. The boundary may extend around at least 75% of the perimeter of each area of relatively lower density.

[0012] The elongate portions may be formed from electrospun strands which are axially aligned with the elongate portions. E.g., each elongate portion may have an elongate axis extending along the length of the elongate portion. The strands within the elongate axis may be aligned with this axis. The strands in the areas of relatively lower density may be randomly oriented.

[0013] The spatial frequency (or mean average spacing) of the elongate portions along two orthogonal axes may be between 1-5mm. The spatial frequency may be calculated as the length of a straight line along an axis divided by the number of elongate portions which intersect the straight line. That is, it corresponds to the mean average spacing of the elongate portions along that axis.

[0014] The elongate portions may form a mesh pattern. The elongate portions may form a regular grid, where the spacings between the mesh strands are spaced between 1-5mm apart. Each area of relatively lower density may be surrounded on by four elongateportions. The four elongate portions may form a rectangular or square shape. The areas of relatively lower density may form a rectangular or square shape. Each elongate portion along each side of the area of relatively lower density may be longer than they are wide.

[0015] The Janus wound pad may have an ultimate tensile engineering strength of at least 10 MPa.

[0016] The Janus wound pad has a water vapor transition rate of between 1.0-5.0 kg / m2 / day.

[0017] The Janus wound pad may comprise chitosan.

[0018] The hydrophilic layer may comprise ethylene-vinyl alcohol (EVOH).

[0019] EVOH is a polymer known for its excellent barrier properties. These barrier films may provide a high level of protection against the entry of oxygen, moisture, and bacteria into the wound site. While maintaining a moist wound environment is generally desirable, in some cases, such as when managing heavily exuding wounds or wounds with infection concerns, a barrier film containing EVOH can be used to help keep the wound dry and isolated from external contaminants. EVOH-containing layer in a dressing may act as a barrier to reduce the risk of contamination and infection. It can create a sterile or semi- sterile environment around the wound.

[0020] The hydrophobic layer may comprise polycaprolactone (POL).

[0021] POL is a biodegradable polymer, which means it can break down naturally over time. This property can be advantageous in wound dressings, especially for wounds that require longer-term care. As PCL degrades, it may reduce the need for frequent dressing changes, minimizing disruption to the wound bed. PCL is a biocompatible, flexible and mouldable material. This property allows it to conform to the contours of the wound, providing a comfortable and secure fit. It can be used in dressings for wounds in challenging anatomical locations. Its biocompatibility allows it to integrate with the surrounding tissue as it degrades, potentially aiding in the healing process.

[0022] The Janus wound pad may comprise an adhesive. The adhesive may be on the hydrophilic side. The adhesive may allow the pad to be securely attached to skin. The adhesive may be weak enough to allow the pad to be removed from skin without damaging the skin.

[0023] The Janus wound pad may be flexible.

[0024] The hydrophobic layer may have a water droplet contact angle greater than 90°.

[0025] The hydrophilic layer may have a water droplet contact angle less than 90°.

[0026] The Janus wound pad has a lateral dimension of at least 3cm.

[0027] The hydrophobic material and the hydrophilic material are capable of hydrogen bonding with each other.

[0028] The electrospun strands in the hydrophilic and hydrophobic layers have a mean yarn diameter of between 5 and 50 pm. The mean yarn diameter may be determined by imaging the yarn and measuring diameters (or widths) of the yarn (across different yarns and at different positions along each yarn), and calculating a mean average.

[0029] According to a further aspect, there is provided a method of producing the Janus wound pad described herein, wherein the method comprises: providing an electrically conductive mesh layer; and electrospinning the hydrophobic and hydrophilic layers onto the mesh layer.

[0030] The mesh layer may be formed of metal.

[0031] The mesh layer may be formed of aluminium.

[0032] The hydrophobic layer may comprise one or more of the following: Hydrophobic- Polyurethane (Pll), Polymethyl Methacrylate (PMMA), Polystyrene (PS), Polycaprolactone (PCL), Polyvinylidene Fluoride (PVDF), and Polyethylene (PE).

[0033] The hydrophilic layer may comprise one or more of the following: Chitosan, Hyaluronic Acid (HA), Collagen, Polyvinyl Alcohol (PVA), Carboxymethyl Cellulose (CMC), Alginate, EVOH (Ethylene Vinyl Alcohol), and Natural polymer gum.

[0034] The mean average width of the areas of relatively lower density may be at least 50% of the spatial frequency of the elongate portions along two orthogonal axes. The mean average width of the elongate portions may be at most 40% of the spatial frequency of the elongate portions along two orthogonal axes. The mass area density of the elongate portions may be at least 120% of the mass density of the pad (e.g., as a whole). The mass area density of the elongate portions may be at most 400% of the mass density of the pad (e.g., as a whole). The mass area density of the areas of relatively lower density may be at most 80% of the mass density of the pad (e.g., as a whole). The mass area density of the areas of relatively lower density may be at least 20% of the mass density of the pad (e.g., as a whole).

[0035] The optical transparency or transmittance of the fabricated wound pad allows for the wound to be properly aligned when the closure is applied, and / or for the monitoring of wound healing progress without having to replace the dressing or disturb the healing process. The hydrophobic layer may provide a level of water resistance to the closure, such that it can prevent water (e.g., from rain or a shower) entering the wound via the closure.

[0036] The wound pad may comprise a polycaprolactone (hydrophobic) layer on the top surface of an ethylene-vinyl alcohol (hydrophilic) nanofiber layer. The hydrophilic layer may absorb the excess biofluid of the wound and prevents bacterial infections.

[0037] The wound pad may comprise chitosan. The wound pad may comprise 1 , 3, and 5% of chitosan. The hydrophilic layer may comprise chitosan. The hydrophobic layer may be free of chitosan. Chitosan may improve biocompatibility and accelerate the healing process via its antimicrobial activity and skin-repairing function.

[0038] The wound pad may have two or more characteristics of the following: high strength, Janus wettability, good WVRT, transparency, and antibacterial properties.

[0039] The Janus wound pad may be a wound dressing and / or a wound closure. A dressing is a sterile pad applied to a wound to promote healing and protect the wound from further harm. A dressing may be designed to be in direct contact with the wound. Many modern dressings are self-adhesive. A wound closure is configured to span across a wound and to hold elements of the wound in place (e.g., two edges of a cut) while the wound heals. E.g., a wound closure may be applied across a laceration in a manner which pulls the skin on either side of the wound together.

[0040] The thickness of the Janus wound pad may be between 50-400 pm (e.g., 75-150 pm or around 100 pm). The thickness may be measured using a micrometer. A nominal pressure may be applied to the Janus wound pad to ensure consistency in measurement (e.g., 0.1 kPa).

[0041] The mesh or fabric preferably has a filament or yarn diameter of between 5 and 50 pm (e.g., 15-20 pm). The nanofiber diameter may be measured from the images collected by using scanning electronic microscope. The images were taken over several locations on a Janus wound pad, to provide sufficient data for statistical analysis. The diameter may be a mean diameter of the strands visible in multiple SEM locations.

[0042] The area density of the wound pad may be between 0.001 and 0.005 g / cm2. The volume density of the wound pad may be 0.1-0.5 g / cm3, based on a volume calculated by multiplying the length, width and thickness of the wound pad.

[0043] The hydrophobic and / or hydrophilic layers may be formed from polymers. A polymer may be a semi-crystalline polymer comprising a mixture of both crystalline and amorphous structures. A polymer material may be transparent across the visible spectrum (e.g., 380 to 700 nm).

[0044] The water vapor transmission rate (WVTR) is a measure of the passage of water vapor through a substance. It is a measure of the permeability for vapor barriers. ASTM F1249-20 is a standard test method for measuring the water vapor transmission rate through flexible barrier materials used in packaging or industrial applications.

[0045] The mass / volume percent (M / V%) of a solution may be defined as the ratio of the mass of solute that is present in a solution in grams, relative to the volume of the solution in milliliters, as a whole multiplied by 100.

[0046] Transmittance may be taken as the percentage of incident light power of a specific wavelength which passes via a sample (e.g., a Janus wound pad). Transmittance is a typically expressed as a percentage between 0% (all light is absorbed) and 100% (no light is absorbed). Average transmittance over a wavelength range may correspond to the mean value of the wavelength-dependent transmittance values within (and / or spanning) the wavelength range.

[0047] Chitosan is a sugar that comes from the outer skeleton of shellfish, including crab, lobster, and shrimp. It may be used as medicine and in drug manufacturing. Chitosan may help blood clot when applied to wounds.

[0048] The hydrophilic layer may comprise EVOH (Ethylene-vinyl alcohol). The hydrophobic layer may comprise PCL (Polycaprolactone).

[0049] Ultimate tensile strength is the maximum stress that a material can withstand while being stretched or pulled before breaking. Engineering stresses and strain are determined based on initial dimensions of the sample.

[0050] The ultimate tensile engineering strength, S, may be calculated based on the following formula:where F is the maximum tensile force applied during the stretching of a piece of closure (until fracture occurs), w is the width of the closure being stretched, d is the thickness of the closure (wd representing the cross-sectional area perpendicular to the force), and matis the porosity of the wound closure (e.g., the porosity being the ratio of void volume to the total volume of the wound closure). The ultimate tensile engineering strength may correspond to the peak or maximum value in the engineering stress-strain curve.BRIEF DESCRIPTION OF THE DRAWINGS

[0051] In the Detailed Description section below, one or more embodiments of the present technology are described in relation to the attached figures. These embodiments are intended to provide a better understanding of the invention, how the invention may be put into practice, and to demonstrate some of the advantages of the invention. The drawings are not necessarily to scale, emphasis instead being placed upon illustrating the principles of various embodiments of the invention. Similar reference numerals indicate similar components.Figure 1(A) is a schematic of the apparatus used to make the Janus wound pads.Figure 1(B) is a schematic of how collimated light from a laser passed through various pads, including embodiments of the present disclosure, and corresponding images of the light projected onto a screen.Figures 2(A)-(D) are images of a logo overlaid with the various pads.Figure 2(E) is a graph of the transmittance of the various pads as a function of wavelength.Figure 2(F) is a stress-strain graph of the various pads.Figures 2(G)-(I) are images of the various pads.Figure 3(A) is an IR spectroscopy graph of the transmittance as a function of wavenumber for each of the components making up a Janus wound pad in according to the present disclosure, and for a Janus wound pad itself.Figure 3(B) is a schematic representation of the two layers of an embodiment of a Janus wound pad at the molecular level.Figure 3(C) is a Differential Scanning Calorimetry (DSC) curve of the heat flow as a function of temperature for each of the components making up a Janus wound pad in according to the present disclosure, and fora mixture of the nanofibers itself. Figure 3(D) is a Thermo-Gravimetric Analysis (TGA) curve of the weight loss as a function of temperature for each of the components making up a Janus wound pad in according to the present disclosure, and for a mixture of the nanofibers itself.Figure 4 is a schematic representation of the measurements made on droplets resting on the hydrophilic and hydrophobic sides of two embodiments of Janus wound pads.Figures 5(A)-(B) are graphs of transmittance as a function of wavelength for various embodiments of Janus wound pads with chitosan.Figure 5(C) are photos of various embodiments of Janus wound pads overlaid on text with chitosan.Figures 5(D)-(E) are stress-strain graphs for various embodiments of Janus wound pads with chitosan.Figures 6(A)-(B) are graphs of transmittance as a function of wavelength for various embodiments of Janus wound pads with chitosan.Figures 6(C)-(D) are stress-strain graphs for various embodiments of Janus wound pads with chitosan.Figures 7(A)-(B) are graphs showing the radius of the zone of inhibition for various embodiments of Janus wound pads with chitosan.Figures 8(A)-(C) are graphs showing the blood clotting factors for various embodiments of Janus wound pads compared with a commercial gauze.DETAILED DESCRIPTIONIntroduction

[0052] Wound pads, such as dressings and closures, ideally should have a large number of different properties, such as transparency, the ability to control water flow, the ability to control air flow, mechanical strength, and antimicrobial properties.

[0053] For example, a combination of transparency and strong mechanical strength of the wound pads (dressings or closures) is highly desirable for wound care. The transparencyallows for visual inspection during the alignment of the wound. Appropriate closure of the wound and aligned edges can potentially promote the natural healing process and reduce scarring.

[0054] After the alignment of the edges, strong tensile strength of the wound pads is important to hold the closed wound in place. Throughout the healing process, the transparency permits visual inspection which enables examination of the healing process and management of the wound.

[0055] Water repellent dressings allow the patients to perform many daily activities without removing the wound pad, such as taking a brief shower or washing their hands. In the present technology, a Janus dressing is composed of a hydrophobic layer to render the outer side water repellent, and a water-absorbing hydrophilic inner layer to absorb wound exudates and control water loss from the wound. In addition, active ingredients that can provide antibacterial properties and promote wound healing may also be integrated into the dressings. Air flow can also be important to allow oxygen to pass through the wound pad and to the wound surface.

[0056] Aside from the chemical and mechanical properties of an electrospun nanofiber membrane, it is still a challenge to design transparent fibrous electrospun membranes due to significant surface light reflection and internal scattering. In the present disclosure, embodiments are described with multiple characteristics, including two or more of: high transparency (e.g., 82-90 %), water control, and high strength (11-13 MPa). The present disclosure also relates to a method of manufacture configured to make such a wound pad.

[0057] Various aspects of the invention will now be described with reference to the figures. For the purposes of illustration, components depicted in the figures are not necessarily drawn to scale. Instead, emphasis is placed on highlighting the various contributions of the components to the functionality of various aspects of the invention. A number of possible alternative features are introduced during the course of this description. It is to be understood that, according to the knowledge and judgment of persons skilled in the art, such alternative features may be substituted in various combinations to arrive at different embodiments of the present invention.Wound Pad Composition

[0058] In this disclosure, strong, and Janus electrospun membranes are described. These embodiments are fabricated using PCL (Polycaprolactone) and 27 EVOH (Ethylene-vinylalcohol) polymers. EVOH comes in a variety of compositions, usually designated by the mol% ethylene in the polymer backbone, ranging from about 27 to 40 mol%. The 27 in the term 27 EVOH refers to a 27mol% composition. For other embodiments, other concentrations of EVOH may be used, e.g., from about 27 to 40 mol%.

[0059] POL is an aliphatic linear polyester and well suited for preparing nanofibers using an electrospinning process. It is also highly biocompatible, slowly biodegradable, mechanically stable, bioresorbable, and a low-cost synthetic polymer, due to its semicrystalline and hydrophobic nature, it exhibits strong wound protection against environmental moisture and bacterial infections.

[0060] EVOH is a hydrophilic semi-crystalline polymer, derived from the hydrolysis of poly (ethylene-co-vinyl acetate). It is commonly used in various biomedical applications because of its good chemical resistance, good thermal stability, good physical properties, excellent biocompatibility, and low cost. Due to its hydrophilic nature, it absorbs wound exudate, controls the humidity, and prevents bacterial infections.

[0061] These materials are spun into nanofibrous membranes by using 1.58 mm and 2.5 mm square aluminium mesh collectors. In this square frame structure, which includes the outer square frame and middle space region, nanofibers are mainly accumulated in highly ordered on the surrounding square frames, and the less amount of fibers accumulation in the area between the frames or center space region.

[0062] An antibacterial material embedded in the polymer may be beneficial for the wound healing process. In this disclosure includes a description of wound pads fabricated with 1 %, 3%, and 5% chitosan-loaded antibacterial electrospun nanofiber along with EVOH nanofiber to accelerate the wound healing process.

[0063] The prepared transparent electrospun wound pads demonstrated good flexibility, robust mechanical properties, higher WVTR, and high transparency.

[0064] The stretchability of the pad can be assessed from the stress-strain curve. One common parameter to evaluate stretchability is the elongation at break, which is the percentage increase in length of the film at the point of failure (breakage). This value represents how much the film can stretch before it breaks and provides an indication of its stretchability.Manufacture

[0065] Figure 1(A) shows the fabrication apparatus for producing transparent Janus electrospun nanofiber wound pads.

[0066] Electrospinning uses an electric potential to draw charged threads from polymer solutions or melts. In this case, the electrospinning apparatus comprises a syringe 101 with an electrically conducting needle 102, and an electrically conducting collector cylinder 103.

[0067] In operation, a voltage is applied between the needle 102 and the receiving cylinder 103. This voltage charges liquid being dispensed by the syringe and generates a fine stream of polymer 104 which is attracted to the collector cylinder 103. This stream will form the strands in the resulting electrospun membrane or pad 107. Controlling concentration of the precursor, flow rate, temperature, and the voltage and distance between the needle and cylinder allows the thickness of the produced strands to be controlled.

[0068] In this case, there were two precursor solutions prepared for electrospinning:(i) 15% by mass PCL in DMF: CHCI3(30:70), and(ii) 22% by mass 27EVOH in DMSO.

[0069] Both the solutions were heated at 80-90 °C with 500 RPM stirring speed for 3 hours to produce the homogeneous solution.

[0070] Afterward, the prepared solution was drawn into a plastic syringe 101 and then installed on the electrospinning machine equipped with a high-voltage power supply 108.

[0071] Electrospinning is a scalable and versatile technique where fibers with micron to nanometer sizes are spun from feedstock solutions or melt by applying an electric field. Electrospinning can be used with different synthetic as well as natural origin polymers. The physicochemical properties of the wound pad can be adjusted by the composition of the polymer solution, flow rate, applied voltage, and temperature. However, electrospun membranes are typically non-transparent because of their high porosity and surface roughness. The refractive index difference between air and the nanofibers causes the loss of light at the interface between the two components.

[0072] For this disclosure, different substrates were used to collect the nanofibers. Using different substrates, or collectors, was found to affect the transparency of the membrane.These substrates were placed over the electrically conductive collector drum 103. The substrates included a continuous Al foil 105 (S1), and square electrically conductive meshes 106 (S2).

[0073] For the Al foil 105 (S1), a continuous sheet of aluminium foil was used as a collector overlaid on the metallic cylinder 103.

[0074] For the square meshes, an aluminium (Al) mesh (1.58 mm or 2.5 mm) was used as a collector overlaid on the metallic cylinder 103. It will be appreciated that other collectors may be used to produce other wound pads.

[0075] In each case, the distance between the needle to the collector and the speed of the collector was kept at 15 cm and 300 RPM.

[0076] As there are two layers, the fabrication was carried out in two stages. First, EVOH nanofibers were fabricated for 30 min at 0.5 ml / hrs and a voltage of 11 kV, and then, in a second stage, PCL nanofibers were fabricated on top of the EVOH nanofibers at 1 .5 ml / hrs flow rate using 11 kV voltage. PCL electrospinning time was 30, 60, and 120 min. In the results described below, the 60 min electrospinning time pads were used. Light transmittance of the resulting pad may be controlled by adjusting one or more of: the thickness of the polymer layers; the strand thicknesses; and the size of Al collector mesh. The mass ratio of the finished wound pad is 81% PCL and 19% EVOH.

[0077] For other embodiments, chitosan was added to the polymer. To develop the PCL / EVOH nanofibers with chitosan molecules, 1 %, 3%, and 5% (M / V%) by chitosan in 1 % by volume acetic acid solution were prepared and the EVOH nanofibers were fabricated along with the chitosan nanofibers at the same flow rate and times. Finally, the PCL nanofibers were fabricated on the top side of the nanofibers. In these embodiments, just the hydrophilic layer comprises the chitosan. That is, the EVOH and Chitosan solutions were prepared separately and added to separate dedicated syringes, and both types of fibers were fabricated simultaneously for same time. The composition of the materials used to make the pads, and of the pads themselves are provided in Table 1 below.

[0078] Table 1 : Composition of Materials used to make Pads.

[0079] The electrospinning dope solution was prepared by adding different concentrations (% W / V) of used polymers in their respective solvents (15% PCL in CHCl3:DMF 70:30, 22% 27 EVOH in DMSO, and 1% or 3% or 5% Chitosan in 1% acetic acid solution). To obtain a homogenous dope solution; the mixture was stirred for 24 hrs at 60 °C and then sonicated to avoid aggregation. The electrospinning process was conducted at room temperature and 25 - 30% relative humidity (RH) atmosphere. A specific volume of 3 ml for each solution was placed in the syringe pump and electrospun onto aluminum foil or screen mesh as the backing layer. The distance between needle-collector and speed of the collector was kept at 15 cm and 300 RPM. First, EVOH and Chitosan dope solution were spun separately and simultaneously with same electrospinning conditions (0.5ml / hr for 30 min., 11Kv). After the fabrication of EVOH and Chitosan nanofibers, PCL spinning on top of it with conditions of 1.5 ml / hr for 60 min and using voltage 12 kV respectively.Experimental Testing

[0080] The transparency of prepared electrospun nanofibers was measured using a UV- Visible spectrophotometer (GENESYS 15™, Thermo Scientific™), at 25 °C in the visible light wavelength range of 200 - 800nm. The spectrometer was configured to illuminate an area of 25 mm x 50 mm (i.e., spanning several mesh spacings).

[0081] To understand the molecular interactions between PCL and EVOH nanofibers, Fourier transform infrared (FTIR, Bruker™) spectroscopy was performed.

[0082] The surface morphology and the arrangement of the PCL / EVOH electrospun nanofiber within the square framework were examined using a field-emission scanning electron microscope (FE-SEM, Mira 3™, Tescan™).

[0083] Differential scanning calorimetry and thermos-gravimetric analysis (TA-Q500, USA) were conducted to determine the thermal properties of solid PCL, solid EVOH, and PC / EVOH composite nanofiber membranes, including glass transition temperature (Tg)and decomposition temperatures (Tm). A heating rate of 10 °C / min was used with a nitrogen atmosphere around the sample.

[0084] The mechanical strength of all the electrospun nanofibers and adhesive efficiencies of the nanofibers was examined using an Instron Corporation™ Automated Materials Testing system at room temperature.

[0085] To determine the wettability of nanofibers, water contact angles were detected by using Contact Angle Meter dosing unit which was produced by Kruss Scientific™ (KRLISS GmbH 22453 Hamburg, Germany).

[0086] WVTR (water vapor transmission rate) is the steady- state rate at which water vapor permeates through a nanofiber at specified conditions of temperature and relative humidity. A DI water bottle was sealed with nanofiber and placed in the oven at 37 °C for 24 hrs and then calculated WVTR (i.e., g / m2 / day) with known nanofiber area or bottle opening area.

[0087] The antibacterial properties of the PCL / EVOH nanofibers were studied against E. coli (ATCC 25922) and S. aureus (ATCC 29213) bacterial strains and the blood clotting - hemolysis profile of the nanofibers was examined on human blood samples.Transparency and arrangements of nanofiber

[0088] In general, strong light scattering can result in opaque electrospun nanofiber membranes. In the recent literature, various approaches have been developed to produce (semi-) transparent electrospun membranes, including solution casting, hot pressing, thermal treatment, electroless deposition, sputtering, and among others. The present technology is based on controlling the structure and alignment of the electrospun strands to improve transparency.

[0089] Figure 1(B) includes a schematic illustration of light propagation patterns in three different media (i) air, (ii) a fibrous deposited on Al foil (S1), and (iii) a fibrous deposited on Al square mesh (S2). Figure 1(B) also shows an image of laser light hitting a screen after having passed through each of the three media. To generate these images, three media were perpendicularly illuminated using a 650 nm wavelength (red) laser and the light projected onto a screen.

[0090] In the case of air, there was hardly any change in the laser beam’s intensity as it passed through the air media and arrived at the screen.

[0091] However, the size of the center light spot was significantly dispersed as the laser traveled through randomly oriented fibrous membrane S1 to the screen, and the scattering light area was very large. The rough surface of the nanofibers and multiple solid-air interfaces in the fibrous structures significantly increased light scattering, and decreased laser beam intensity as well.

[0092] When the laser beam was applied to the square shape membrane, S2, there was very little loss of light intensity as the laser propagated through the square framework to the screen. The center light spot was still visible, and the area of scattering was small compared with the S1 membrane.

[0093] Figure 2(A) is a photo of the University of Alberta logo. Figure 2(B) is a photo of the same logo with an S1 overlaid over the logo. Figures 2(C)-(D) are photos of two S2 pads overlaid over the logo. Figure 2(C) corresponds to a mesh size of 1.58 mm, and figure 2(D) corresponds to a mesh size of 2.5mm. As the photos show, both S2 pads provide a considerably clearer view of the logo compared with the S1 logo. The larger mesh size increases clarity. It will be appreciated that this test indicates the utility of having a transparent pad to allow the wound to be properly aligned as the wound pad is being applied. A properly aligned would aids healing and reduces long-term scarring.

[0094] To determine the transparency of the fabricated membranes, a UV-visible spectrometer was used to measure the transmittance of the various pads. The structure of the substrate (size and shape) played a significant contribution to achieving maximum light transmittance. As can be shown in Figure 2(E), the polymeric nanofibers collected on substrates had light transmittances of <1% (S1 ; Al foil), 16% (S2;1.58 mm), and 26% (S2; 2.5 mm) respectively. The light transmittance analysis indicates that a fibrous membrane structure on the square mesh has superior light transmittance than the randomly arranged S1 pad. Unless otherwise specified, transmittance percentage values in this disclosure are the average transmittances over the wavelength range 350-800 nm.Pad Strength

[0095] For wound closure and healing applications, these transparent electrospun nanofibers must have strong mechanical properties. To better understand the robust mechanical strength, the tensile strength of S1 and S2 (1.58 mm and 2.5 mm) nanofiber membranes were measured. Figure 2(F) shows that the stress-strain curves were largely similar for all three pads, and all the pads had ultimate tensile engineering strengths ofabout ~ 12 to 13 MPa, respectively. This indicates that the S2 fibrous pads with high light transmittance did not affect the mechanical strength. This may be because they have similar levels of polymer deposition.Arrangement of Fibres

[0096] Figure 2(G-I) displays the FE-SEM (Field-Emission Scanning Electron Microscopy) images of fibrous membranes developed on the different substrates S1 and S2. It was found that fibrous membranes deposited over aluminum foil produced randomly orientated and disorganized 3D fibrous structures - see Figure 2(G).

[0097] Figure 2(H) includes three images of the 1.58 mm S2 pad, and Figure 2(l) includes three images of the 2.5 mm S2 pad. In each figure, the central image is a zoomed-out region showing the two-dimensional mesh pattern comprising areas of relatively lower density of electrospun strands bounded by elongate portions of relatively higher density of electrospun strands; the left image is a zoomed-in portion showing aligned strands in the elongate portions; and the right image is a zoomed in portion showing randomly aligned strands in a lower density area.

[0098] In these pads with the square framework structures (1.58 mm and 2.5 mm), the majority of the fibers were deposited at the frame region and relatively few fibers are deposited in between the square frames. The fibrous membrane transmitted more light with minimal loss of light at the center region Figure 2(H) and 2(l).

[0099] As shown in figures 2(H) and 2(l), in these embodiments, in the elongate portions, the electrospun strands are aligned with the direction of the elongate portions.

[0100] Because the elongate portions are aligned with the mesh during manufacture, the spatial frequency of the elongate portions along two orthogonal axes correspond to the spacing of the mesh. The spacing would typically be between 1-5 mm. It will be appreciated that other mesh sizes and shapes may be used.Molecular interaction between nanofibers

[0101] The chemical interactions between the constituents of nanofibers were studied by FTIR (Fourier Transform Infrared) spectroscopy. FTIR spectra of the PCL, EVOH, and POL / EVOH nanofiber are compared in Figure 3(A).

[0102] The FTIR characteristic peaks of PCL appeared at:• 2950 cm-1(asymmetric -CH2 stretching),• 2860 cm-1(symmetric -CH2 stretching),• 1728 cm-1(carbonyl stretching),• 1294 cm-1(C-0 and C-C stretching),• 1240 cm-1(asymmetric C-O-C stretching), and• 1170 cm-1(symmetric C-O-C stretching).

[0103] Characteristic peaks of EVOH appeared at:• 3350 cm-1(-OH stretching),• 2930 and 1440 cm-1(-CH2 stretching and bending, respectively), and• 2860 and 1330 cm-1(-CH2 stretching and bending, respectively).

[0104] The -C=O band of the PCL and -OH band of the EVOH are red-shifted during PCL / EVOH nanofiber fabrication (3300 cm-1and 1712 cm-1), indicating that intermolecular hydrogen bonding is responsible for the stable and strong Janus nanofiber formation Figure 3(B).DSC and TGA

[0105] The DSC and TGA curves for pure PCL, pure EVOH, and PCL / EVOH PCL nanofibers are shown in Figure 3(C-D). The melting behavior of pure PCL, pure EVOH, and PCL / EVOH PCL nanofibers were observed around 76.69 °C, 170.01 °C, and 70.92 - 172.44 °C Figure 3(C).

[0106] From the TGA curve Figure 3(D), it can be observed that pure PCL shows a degradation step in the range from 388 to 439 °C with a 96% weight degradation.

[0107] The EVOH degradation profiles followed a two-step process, with the first mass loss (76%, 350 - 410 °C) assigned to the degradation of vinyl alcohol units, and the second one, (ca. 24%, 410 - 483 °C) attributed to the degradation of ethylene units.

[0108] The degradation of a pathway of PCL / EVOH combined nanofiber followed a single step process, with 85% degradation at 340 - 410°C. The decreases in melting and degradation temperature of PCL / EVOH nanofiber can be attributed to the multiple hydrogen bonding interactions between PCL / EVOH.Janus water wettability of nanofibers

[0109] To verify the wettability of the composited PCL and EVOH layers of the Janus membrane, static water contact angles (WCA) are tested. A static water contact angle may be measured using an optical tensiometer by, for example, placing a droplet on thesolid surface and recording an image of the drop. The static contact angle is then defined by fitting Young-Laplace equation around the droplet, although other fitting methods such as circle and polynomial can also be used.

[0110] As shown in Figure 4(A), the water contacts of the top layer (PCL) and bottom layer (EVOH) of both nanofiber membrane was 122.6° (PCL - 1.58 mm size mesh), 116.6° (PCL - 2.5 mm size mesh) and 83.5° (EVOH - 1.58 mm size mesh), 76.8° (EVOH - 2.5 mm size mesh), respectively. These results are showing a great difference in wettability between the two layers in single nanofiber fabrication.

[0111] The asymmetric wettability of the two sides of the membrane facilitated the absorbing efficiency of water. The 30-35 pL water absorbing processes in the positive direction of the PCL / EVOH nanofiber membrane are shown in Figure 4(A-B).

[0112] For the top layer (PCL) of the Janus nanofiber membrane, the droplet spread on the surface of the hydrophobic layer due to gravity and becomes hemispherical. There was no obvious infiltration in contact angle, height, and length of the water droplet within 15 min.

[0113] In contrast, for the bottom layer (EVOH) of the membrane, the water quickly penetrated within 15 min and the contact angles and height reasonably reduced until it reached values of 0. The above results show a strong wetting force from the EVOH nanofiber membrane, which accelerated the process of directional water absorption for the enlarged wettability gradient.

[0114] This asymmetry in wettability means that the wound pad can absorb and release aqueous fluids generated by a wound, while preventing external water (e.g., from a shower or rain), form entering the wound from the outside.Chitosan nanofibers

[0115] In further embodiments, chitosan as an active ingredient was integrated into the membrane, giving additional wound healing and antibacterial properties, 1%, 3%, and 5% chitosan (percentage by mass) was incorporated during EVOH nanofiber fabrication at the same flow rate and time. Chitosan is a natural and readily available polymer, and it has been extensively applied in wound dressing because of its effective antibacterial properties, water absorption, biocompatibility, biodegradability, non-cytotoxicity, nonantigenicity, and biological membrane function.

[0116] As shown in figure 5(A), the transmittance with 1%, 3%, and 5% chitosan in 1.58 mm size mesh was observed to be 14%, 13%, and 12%, respectively. These results are visually shown in figure 5(C) which shows a series of photos of the three pads overlying paper with text printed on it.

[0117] As shown in figure 5(B) the transmittance with 1%, 3%, and 5% chitosan in the 2.5 mm size mesh was 24%, 21 %, and 18%. That is, additional chitosan is associated with a slightly lower transmittance.

[0118] Decreasing the transparency after the encapsulation of chitosan may be due to the higher density of the fibers. The mechanical strength of the nanofiber after incorporating chitosan molecules was found in both cases to range between ~11 to 13 MPa. There was no significant difference in mechanical strength with and without chitosan. These results are shown in stress-strain curves in Figure 5(D) and (E) for a 1.58 mm S2 pad and a 2.5 mm S2 pad respectively.

[0119] As shown in figure 5(C) that although the transparency has decreased, the logo is still largely visible through the chitosan infused wound pads. This shows that these pads would still allow the wound to be aligned and / or monitored through the healing process. In addition, the chitosan will also provide an antibacterial environment for the wound and aid healing. Therefore, it will be appreciated that the benefits of higher concentrations of chitosan can be weighed against the decreased transparency, and a compromise may be found between the two parameters to provide the advantages of antibacterial properties and reasonable transparency.Adhesive property of nanofibers

[0120] The adhesiveness in wound dressing material can firmly attach to external and internal wounds, and form a barrier to prevent bacterial invasion, accelerate healing, and avoid secondary damage caused by sutures. Many wound dressings cause the patient pain upon removal or replacement, and this makes selecting an adhesive that much more important, especially on sensitive skin.

[0121] An adhesive material (e.g., 2-octyl cyanoacrylate; 10pL in 6.25 A2 fiber) may be used to adhere the pad to skin tissue.

[0122] As shown in Figures 6(A)-(B), which are respectively graphs of optical transmittance of a 1.58 mm S2 pad and a 2.5 mm S2 pad, it was found that thetransparency of both the S2 pads with 1%, 3%, and 5% chitosan significantly increased with adhesive material and reached to ~ 82 - 90%.

[0123] At the same time, we studied the mechanical strength of both the nanofiber with 1 %, 3%, and 5% chitosan and representative stress-strain curves and the tensile properties for a 1.58 mm S2 pad in Figure 6(C) and for a 2.5 mm S2 pad in Figure 6(D).

[0124] The mechanical strength was observed between ~ 11-13 MPa with higher stretchability.

[0125] The increase in transmittance may be because of the adhesive material filling the empty pores to reduce the number and spatial frequency of air-nanofiber boundaries. This reduces scattering, and a higher transparency is achieved.Healing and weight loading efficacy of nanofibers

[0126] The healing efficiency of a 1.58 mm S2 pad and a 2.5 mm S2 pad was measured. In each case, the pad was torn apart. When both the nanofibers were joined as a whole entity after 4 hrs, the reconstructed nanofiber was lifted, it could sustain tensile force in a direction perpendicular to the original tear.

[0127] The rejoining of the tear was effected as follows: after cutting the nanofiber pad into two fragments (2.5cm width x 5cm length), the adhesive material was applied to one of the fragments (area covered by adhesive 2.5cm width x 2.5cm length). The second fragment of the sample was then brought in contact with the first one. The contact area between the sample and adhesive was chosen to be 2.5cm width x 2.5cm or 6.25cm2. After 2-3 hrs, both the pad fragments were completely rejoined. The repair was a result of polymerization via covalent bonds of cyanoacrylates and hydroxyl ions in the EVOH polymer.

[0128] It was further observed that the self-repaired nanofiber film could be stretched without further breaking. After stretching beyond its limit, the breakage takes place at the healed site, showcasing the excellent self-healing properties of the nanofiber. We measured the healing efficiency of both nanofiber by using ultimate tensile engineering strength measurements. The fracture stress for 1.58 mm and 2.5 mm mesh nanofiber was 12.04 MPa and 12.01 MPa, representing a recovery of ~90 % or more of the original ultimate tensile engineering strength.Real-world Testing

[0129] It was found that the closure maintains excellent transparency, skin adhesiveness, and stretchability of the nanofiber applied to the front of a knee, even upon 90° bending of the leg, and taking an extended (4 km) walk.

[0130] Moreover, the macroscopic tear strength of different materials was determined by introducing a 5 mm tear at the edge of a 2.5 cm pad on both a 1.58 mm and a 2.5 mm S2 pad (both with 5% chitosan). For both pads, with a 2.5 N (0.26 kg weight) load applied in a direction perpendicular to the tear, there was no change in the size of the cracks. At a higher load of 10.4N (1.06 kg) the tear enlarged. The tearing strength of the material would be 4N or larger.In vitro degradation and WVTR of nanofibers

[0131] We studied the in vitro degradation of the electrospun nanofibers in terms of morphological changes, and weight loss. It was found that there are no significant changes in weight loss or fibrous morphology and arrangement were observed after 14 days of incubation time at 37°C. This confirms the formation of stable square arrangements of the PCL and EVOH nanofibers.

[0132] Here the WVTR of the various samples is summarized in Table 2. With an increased concentration of the chitosan or with adhesive material the rate of the water transition decreases and the lower values of the WVTR that defines indicate the more compact structures of the nanofiber.

[0133] Table 2: Water vapor transition rateSize of the mesh Chitosan (%W / V) % WVTR (gm / / »2 / day)1.580 1620.29 ± 40.131.581 1592.35 ± 27.321.583 1526.00 ± 13.071.585 1459.66 ± 21.871.58 mm 5 with Adhesive A 1444.11 ± 16.322.5 mm 0 1833.70 ± 18.512.5 mm 1 1725.50 ± 11 .212.5 mm 3 1658.70 ± 54.322.5 mm 5 1579.08 ± 33.042.5 mm 5 with Adhesive A 1556.21 ± 11.12

[0134] The rate of water vapor transmission is an important parameter of the dressing and represents the dressing’s ability to retain moisture. It was found that the dressing witha WVTR of 2028.3 ± 237.8 g / m2 / day was able to maintain an optimal moisture content for the proliferation and regular function of epidermal cells and fibroblasts in a three- dimensional culture model.In vitro antibacterial study of nanofibers

[0135] The antibacterial performance is a characteristic that a dressing material must have to inhibit harmful microorganisms and prevent bacterial infection. The antibacterial activity of both the nanofiber with and without chitosan was evaluated against gram negative E.coli and gram positive S. aureus bacterial strains using the disc diffusion method. The results of antimicrobial activity shown in Figure 7(A-B), show that the positive control (without chitosan) did not exhibit an antibacterial response against the both the bacterial strains.

[0136] However, with increasing the concentration of chitosan from 1 % to 5%, both the nanofibers showed the slightly higher zone of inhibition against E. coli (23.9 mm, and 22.3 mm) indicating that inhibitory capacity was higher for gram negative bacteria than gram positive (S. aureus). Thus, the antibacterial activity of both the nanofibers are mainly due to the presence of chitosan molecules, the fabricated nanofibers could be used as the wound dressing material to kill the both the bacteria and prevent infection with improved the healing process.In vitro blood clotting and hemolysis profile of nanofibers

[0137] A dynamic whole blood clotting model was used to study the in vitro hemostatic property. In comparison to commercial medical gauze, there was more blood clots formed on the hydrogel film’s surface. The commercial gauze control group showed a clearly increased color in the water after rinsing with 10 ml of water, showing the blood was difficult to clot on the commercial gauze. Whereas the supernatant was transparent and clear, and the blood had completely coagulated on the hydrogel film structures. This demonstrated that blood may coagulate effectively on the hydrogel film structure, thus reducing the time for blood to coagulate. The value of absorbance for the hemoglobin solution released by the uncoagulated erythrocytes is used to calculate the blood-clotting index (BCI).

[0138] As shown in figure 8(A), the commercial gauze sample had the highest BCI value (82.2 ± 2.9%), which indicated the slowest blood-clotting speed. The BCI values for the 1.58 mm and 2.5 mm S2 pads were 27.4±3.1% and 32.1 ±2.1 % respectively, lower thanthe commercial gauze sample. These results suggest that the hydrogel film shows excellent in vitro hemostatic.

[0139] The blood clotting time of both the 1 .58 mm and 2.5 mm S2 pads with 5% chitosan and commercial gauze samples during hemostatic were measured. This is shown in figure 8(B). After a specific amount of time of adding blood to the test tubes, both samples could produce blood clots. The blood clotting time for the 1.58mm and 2.5mm size mesh nanofiber was 145 ± 12 sec and 168 ± 8 respectively, which was considerably quicker than the time required by commercial gauze to clot blood (241 ± 5 sec). Due to the porous structures of the nanofiber, which dramatically expands the surface area and makes chitosan contact with the blood cells. This caused platelets to clump together to form blood coagulation and increase entire blood viscosity.

[0140] Figure 8(C) shows the hemolysis ratio for the commercial gauze and the 1 .58 mm and 2.5 mm S2 pads. Hemolysis is the rupturing (lysis) of red blood cells. Each sample had a negligible hemolysis ratio (< 2%), indicating that it was not causing the hemolysis and showing good hemocompatibility.

[0141] Although the present invention has been described and illustrated with respect to preferred embodiments and preferred uses thereof, it is not to be so limited since modifications and changes can be made therein which are within the full, intended scope of the invention as understood by those skilled in the art.

Claims

CLAIMS1 . A Janus wound pad comprising: a hydrophobic layer comprising electrospun strands of a hydrophobic material; and a hydrophilic layer comprising electrospun strands of a hydrophilic material, wherein the Janus wound pad has an average transmittance of at least 10% for wavelengths between 350-800nm.

2. The Janus wound pad according to claim 1 , wherein the Janus wound pad has a two-dimensional mesh pattern comprising areas of relatively lower density of electrospun strands bounded by elongate portions of relatively higher density of electrospun strands.

3. The Janus wound pad according to claim 2, wherein in the elongate portions, the electrospun strands are axially aligned with the elongate portions.

4. The Janus wound pad according to any one of claims 2-3, wherein the spatial frequency of the elongate portions along two orthogonal axes is between 1-5mm.

5. The Janus wound pad according to any one of claims 2-4, wherein the elongate portions form a mesh pattern.

6. The Janus wound pad according to any one of claims 1-5, wherein the Janus wound pad has an ultimate tensile engineering strength of at least 10 MPa.

7. The Janus wound pad according to any one of claims 1-6, wherein the Janus wound pad has a water vapor transition rate of between 1 .0-5.0 kg / m2 / day.

8. The Janus wound pad according to any one of claims 1-7, wherein the Janus wound pad comprises chitosan.

9. The Janus wound pad according to any one of claims 1-8, wherein the hydrophilic layer comprises ethylene-vinyl alcohol (EVOH).

10. The Janus wound pad according to any one of claims 1-9, wherein the hydrophobic layer comprises polycaprolactone (PCL).

11. The Janus wound pad according to any one of claims 1-10, wherein the Janus wound pad comprises an adhesive.

12. The Janus wound pad according to any one of claims 1-11 , wherein the Janus wound pad is flexible.

13. The Janus wound pad according to any one of claims 1-12, wherein the hydrophobic layer has a water droplet contact angle greater than 90°.

14. The Janus wound pad according to any one of claims 1-13, wherein the hydrophilic layer has a water droplet contact angle less than 90°.

15. The Janus wound pad according to any one of claims 1-14, wherein the wound pad has a lateral dimension of at least 3cm.

16. The Janus wound pad according to any one of claims 1-15, wherein the hydrophobic material and the hydrophilic material are capable of hydrogen bonding with each other.

17. The Janus wound pad according to any one of claims 1-15, wherein the electrospun strands in the hydrophilic and hydrophobic layers have a mean yarn diameter of between 5 and 50 pm.

18. A method of producing the Janus wound pad of any of claims 1-16, wherein the method comprises: providing an electrically conductive mesh layer; and electrospinning the hydrophobic and hydrophilic layers onto the mesh layer.

19. The method according to claim 17, wherein the mesh layer is formed of metal.

20. The method according to any one of claims 17-18, wherein the mesh layer is formed of aluminium.

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