Nanofibre wound dressing containing rutin

Abstract

The invention relates to a nanofibre wound dressing containing rutin. The nanofibre wound dressing that is the subject of the invention comprises methacrylic acid - methyl methacrylate copolymer (1:1) and rutin hydrate active ingredient and is produced by electrospinning method. By means of the rutin active ingredient contained in the nanofibre wound dressing that is the subject of the invention, success is increased in the angiogenesis, granulation tissue formation, collagen disorganisation and neutrophil filtration stages of wound healing.

Description

[0001] NANOFIBRE WOUND DRESSING CONTAINING RUTIN

[0002] Technical Field

[0003] The invention relates to a nanofibre wound dressing containing rutin. In said nanofibre wound dressing, the rutin hydrate active ingredient is trapped in the methacrylic acid - methyl methacrylate copolymer (1 :1 ). The success in the angiogenesis, granulation tissue formation, collagen disorganisation and neutrophil filtration stages of wound healing is increased by means of the rutin hydrate active ingredient contained in said nanofibre wound dressing.

[0004] State of the Art

[0005] The disruption of the integrity and functions of tissues or organs by various factors is called wound, and the restoration of this integrity through a series of intertwined processes is called wound healing

[0001] . Wound healing is a dynamic and complex process consisting of successive periods. In acute wounds, the tissue healing process occurs regularly and on time. In chronic wounds, healing takes longer than normal. The use of appropriate wound dressings plays an important role in the wound healing process [2]. Wound dressings cover the wound area, protect the damaged tissue from external effects, and contribute to the healing process by activating cell production if their structure is suitable. A wide variety of wound care products are used in wound care today. These include composite dressings, transparent film dressings, hydrocolloids, alginate dressings, wound fillers, antibacterial dressings and hydrogel dressings.

[0006] The electrospinning process, which allows the production of fibres from many synthetic and natural polymers, can be used in many ways. It is a process that is easy to apply and can be used to produce nanofibres at low costs. The materials produced have advantages in biomedical use due to their high surface / volume ratio and porous structure. In recent years, the production of nanofibrous wound dressings by electrospinning has rapidly gained importance. The similarity of nanofibres to the extracellular matrix structure serves as a support structure for cell proliferation. In addition, their high porosity ensures oxygen permeability and their small porous structure prevents bacterial penetration [3].

[0007] Among the materials examined for application in the electrospinning method are polyvinyl alcohol, polyethylene oxide, polyvinyl pyrrolidone, polyacrylamide, polyurethane, polycarbonate, polytetrafluoroethylene, polyethylene, polypropylene, polyacrylate, and polyacrylate for synthetic polymer systems. Hydroxybutyrate, polyaniline, aramid and the like are included, and natural polymers include deoxyribonucleic acid, collagen, a-lactalbumin, invertase, silk fibroin and the like.

[0008] In the state of the art, wound dressings called passive dressings are used. Passive dressings are products that protect the wound by only covering it, such as known gauze and bandage, and do not have an active effect on the healing process. The intended function of these dressings is to prevent the growth of bacteria in the wound environment by drying the wound to allow the evaporation of wound exudate [4]. However, today, it is understood that growth factors, cytokines and extracellular matrix play an active role in wound healing, that healing is a dynamic process and that traditional wound dressings are not sufficient for effective wound healing. Therefore, when traditional passive wound dressings are used in wound healing, there is a risk that wounds will not heal effectively.

[0009] In another case in the state of the art, wound dressings that are not biocompatible are used. However, if there is no biocompatibility in the wound dressings, the structure of the platelets coming to the area deteriorates and causes some molecules to be released. With this secretion, other cells also migrate to the area, coagulation is provided in the area and thrombus is formed. The formed thrombus can also break away from the surface and cause embolism. This situation can cause death in some advanced tissue damages [5]. For this reason, the use of these materials that are not biocompatible prolongs the treatment process and complicates healing. In this case, the risk of non-biocompatible wound dressings causing tissue damage and prolonging the treatment process and enhancing healing cannot be prevented.

[0010] In another case in the state of the art, biodegradable hydrogel wound dressings are produced from chemically or physically cross-linked hyaluronic acid, chitosan, cellulose, alginate, collagen and gelatin macromolecules [6]. These systems can be formed without the need for chemical modification or the use of a cross-linker, therefore they are considered safe in in vivo applications. However, since they have poor mechanical properties and disintegrate rapidly, it is difficult to control structural variables including pore sizes, gelation times and disintegration profiles. This also limits the design flexibility of the products. Therefore, the risk of design difficulties caused by wound dressings with poor mechanical properties cannot be prevented.

[0011] The limitations and inadequacies of the solutions in the state of the art, the problem of effective wound healing when traditional passive wound dressings are used, the risk of non-biocompatible wound dressings causing tissue damage and strengthening healing by prolonging the treatment process, and the disadvantages of design difficulties caused by wound dressings with poor mechanical properties have made it necessary to make a development in this area.

[0012] Brief Description and Aims of the Invention

[0013] The invention relates to a nanofibre wound dressing containing rutin hydrate active ingredient. In the wound dressing of the invention, rutin hydrate active ingredient is trapped in methacrylic acid - methyl methacrylate copolymer (1 :1 ). Said wound dressing comprises methacrylic acid - methyl methacrylate copolymer (1 :1 ) and rutin hydrate active ingredient.

[0014] One aim of the invention is to increase the stability and effectiveness of the rutin hydrate active ingredient used in the wound dressing. The stability and effectiveness of the rutin hydrate active ingredient used as a flavonoid in the nanofibre wound dressing of the invention is increased by using it in various formulations in the nanofibre wound dressing.

[0015] Another aim of the invention is to obtain a drug carrier system that releases to the wound in a customised manner. The normal pH value of the skin is 5.5. However, this value increases to pH 8 as a result of an injury to the skin. The methacrylic acid - methyl methacrylate copolymer (1 :1 ) from which said formulation is prepared is a pH sensitive polymer and begins to swell and dissolve at values above pH 6. A carrier system that is customised to the wound and sensitive to increasing pH is obtained by using methacrylic acid - methyl methacrylate copolymer (1 :1 ) in said nanofibre wound dressing.

[0016] Another aim of the invention is to ensure that the wound is protected from environmental pollution in the area where the wound dressing is used. The wound can be protected from environmental pollution and environmental factors by means of the nanofibre structure of said wound dressing.

[0017] Another aim of the invention is to accelerate the treatment period of the wounded area in the use of the wound dressing. The treatment period in the wounded area is accelerated by means of the skeletal structure in which the granulation tissue of said nanofibre wound dressing can advance.

[0018] Another aim of the invention is to prevent the use of wound dressings in dermatological treatments from being restricted due to their interactions with environmental factors. By means of the nanofibre structure of said wound dressing, the wound does not interact with environmental factors such as pH, oxygen, humidity, light and heat.

[0019] Another aim of the invention is to regulate the inflammation stage by improving neutrophil infiltration in the wounded area. By means of both the nanofibre structure of said nanofibre wound dressing and the fact that it contains the active ingredient rutin hydrate, the inflammation stage is regulated and neutrophil infiltration is also improved.

[0020] It is aimed to use this nanofibre wound dressing as a carrier system that increases the effectiveness and stability of the active ingredient rutin hydrate it contains, to minimise environmental factors that will negatively affect wound healing, to increase success in wound healing stages, to accelerate the treatment period in the wounded area by means of the skeletal structure where granulation tissue can advance, and to increase the release rate of the active ingredient by being pH sensitive.

[0021] Description of Drawings

[0022] Figure 1. Nanofibre of methacrylic acid - methyl methacrylate copolymer (1 :1 ) -rutin hydrate formulation 30,000x Magnification Figure 2. Nanofibre of methacrylic acid - methyl methacrylate copolymer (1 :1 ) -rutin hydrate formulation 220,000x Magnification

[0023] Figure 3. Schematic representation of nanofibre production by electrospinning method

[0024] Description of Reference Numbers:

[0025] 1- Pump

[0026] 2- Syringe

[0027] 3- Electrospinning solution

[0028] 4- Metal nozzle

[0029] 5- Taylor cone

[0030] 6- Collector (Rotary drum)

[0031] 7- Electrospinning solution jet

[0032] 8- Voltage generator

[0033] Detailed Description of the Invention

[0034] The invention relates to a nanofibre wound dressing containing rutin hydrate active ingredient. In the wound dressing of the invention, rutin hydrate active ingredient is trapped in methacrylic acid - methyl methacrylate copolymer (1 :1 ). By means of the rutin hydrate active ingredient contained in said wound dressing, success is increased in the angiogenesis, granulation tissue formation, collagen disorganisation and neutrophil filtration stages of wound healing, and the formation properties of the collagen structure in the final phase of wound healing are also improved. By means of this nanofibre wound dressing, the effectiveness and stability of the rutin hydrate are increased, the interactions of the wound dressing with environmental factors are reduced, neutrophil infiltration is improved in the wound area and the inflammation stage is regulated, success is increased in the wound healing stages, the treatment period is accelerated in the wound area by means of the skeletal structure where the granulation tissue can advance, and it is used as a carrier system that increases the release rate of the active ingredient as pH sensitive.

[0035] The nanofibre wound dressing of the invention comprises methacrylic acid - methyl methacrylate copolymer (1 :1 ) and rutin hydrate active ingredient. In one embodiment of the invention, the nanofibre wound dressing comprises 98- 90.5% methacrylic acid - methyl methacrylate copolymer (1 :1 ) and 2-9.5% rutin hydrate.

[0036] The production method of the nanofibre wound dressing by electrospinning, which is the subject of the invention, comprises process steps of: i. preparing the solvent consisting of dimethylformamide (DMF) and methyl alcohol (MeOH) by simple mixing method, ii. adding methacrylic acid - methyl methacrylate copolymer (1 :1) to the solvent mixture containing DMF and MeOH, iii. beginning mixing the solution on a magnetic stirrer, iv. adding rutin hydrate active ingredient to the solution that is being mixed, v. keeping the solutions in an ultrasonic bath to remove air bubbles and making them suitable for electrospinning, and vi. spinning the solutions by electrospinning method.

[0037] In one embodiment of the invention, the production method of the nanofibre wound dressing by electrospinning comprises process steps of: i. preparing 10 mL solvent consisting of dimethylformamide (DMF) and methyl alcohol (MeOH) by simple mixing method, ii. adding methacrylic acid - methyl methacrylate copolymer (1 :1) to the solvent mixture containing DMF and MeOH, iii. beginning mixing the solution on the magnetic stirrer by means of a 20 mm long magnetic stir bar at a speed of 750 rpm and room temperature, iv. adding rutin hydrate active ingredient to the solution that has been being mixed for 23 hours and stirring for another 1 hour, v. keeping the solutions in an ultrasonic bath for 20 minutes to remove air bubbles and making them suitable for electrospinning, and vi. spinning the solutions by electrospinning method.

[0038] In the preparation of the nanofibre wound dressing by electrospinning method, which is the subject of the invention, the electrospinning solution of said nanofibre wound dressing comprises methacrylic acid - methyl methacrylate copolymer (1 :1 ), methyl alcohol (MeOH), dimethylformamide (DMF) and rutin hydrate active ingredient.

[0039] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 140 mg rutin hydrate active ingredient.

[0040] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 140 mg rutin hydrate active ingredient.

[0041] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 140 mg rutin hydrate active ingredient.

[0042] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 140 mg rutin hydrate active ingredient.

[0043] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 135 mg rutin hydrate active ingredient.

[0044] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 135 mg rutin hydrate active ingredient.

[0045] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 135 mg rutin hydrate active ingredient.

[0046] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 135 mg rutin hydrate active ingredient. In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 130 mg rutin hydrate active ingredient.

[0047] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 130 mg rutin hydrate active ingredient.

[0048] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 130 mg rutin hydrate active ingredient.

[0049] In one embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 130 mg rutin hydrate active ingredient.

[0050] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 125 mg rutin hydrate active ingredient.

[0051] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 125 mg rutin hydrate active ingredient.

[0052] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 125 mg rutin hydrate active ingredient.

[0053] In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 125 mg rutin hydrate active ingredient. In another embodiment of the invention, the electrospinning solution of said nanofibre wound dressing comprises 1375 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 137.5 mg rutin hydrate active ingredient.

[0054] The solvent used in the preparation of the electrospinning solution of the wound dressing of the invention comprises dimethylformamide (DMF) and methyl alcohol (MeOH). The total solvent volume consisting of DMF and MeOH is 10 mL.

[0055] In one embodiment of the invention, the solvent used in the preparation of said nanofibre wound dressing comprises 6 mL MeOH and 4 mL DMF.

[0056] In another embodiment of the invention, the solvent used in the preparation of said nanofibre wound dressing comprises 6.5 mL MeOH and 3.5 mL DMF.

[0057] In another embodiment of the invention, the solvent used in the preparation of said nanofibre wound dressing comprises 7 mL MeOH and 3 mL DMF.

[0058] In another embodiment of the invention, the solvent used in the preparation of said nanofibre wound dressing comprises 7.5 mL MeOH and 2.5 mL DMF.

[0059] The wound dressing of the invention provides pH-sensitive release. The wound dressing of the invention provides controlled release in accordance with the 1 st degree in in vitro studies in pH 7.4 environment. Thus, the active substance release is ensured throughout the treatment period and the treatment success is increased. In the production of the wound dressing of the invention, the effects of randomly selected experimental conditions on nanofibre formation were investigated. For this purpose, the methacrylic acid - methyl methacrylate copolymer (1 :1 ), which was selected to produce nanofibres containing rutin hydrate active substance, was dissolved in solvents at certain ratios and spun. In the experiments conducted, the presence of nanofibre formation was first examined under light microscope and then the samples suspected of nanofibre formation were examined under electron microscope. In the light of the information obtained as a result of these experiments, the levels of the factors determined to create the Taguchi experimental design that will not prevent nanofibre formation are shown in Table 1 . Table 1. Factors and levels for Taguchi design of nanofibres prepared with methacrylic acid - methyl methacrylate copolymer (1 :1) The experimental pattern prepared according to the factors and levels determined for the Taguchi method is shown in Hata! Ba§vuru kaynagi bulunamadi.. The amount o f rutin added to the formulations was calculated to be 10% of the polymer.

[0060] Table 2. methacrylic acid - methyl methacrylate copolymer (1 :1 ), ingredient quantities for formulations used

[0061] Electrospinning solutions were prepared in the production of the nanofibre wound dressing that is the subject of the invention. Simple dissolution method was used in the preparation of the solutions to be spun via electrospinning device. After weighing the polymers in the ratios specified in Hata! Ba§vuru kaynagi bulunamadi., they were p laced in a 40 mL volume, screw-capped, glass vial. Immediately after the solvent mixture was added to the polymers, mixing was started at 750 rpm on the magnetic stirrer with a 20 mm long magnetic stir bar. Weighed rutin hydrate was added to the solutions that were still mixing at the 23rdhour of mixing. The mixing process was continued for another 1 hour to dissolve the rutin hydrate and electrospinning solutions were prepared at the end of the total 24-hour period. The solutions were kept in an ultrasonic bath for 20 minutes to remove air bubbles and make them suitable for electrospinning.

[0062] The electrospinning solutions were transferred into a suitable syringe and placed in the pump to be transferred to the nozzle with a polyurethane tube. The solution, which was advanced only with the help of a pump until the nozzle exit facing the collector, was formed at the nozzle tip, and then the flow and high voltage were started to ensure that the polymer reached the collector in the form of fibres. The flow rate and voltage values were adjusted according to the values that would create a visible and continuous Taylor cone. The drum collector was adjusted to move at a speed of 3 mm.sec’1in a linear orbit of 80 mm and to rotate around its own axis at a speed of 300 rpm. The average fibre diameters of the nanofibres obtained as a result of the experiments carried out according to the Taguchi method are shown in Table 3 and the bead numbers in Table 4. Table 3. Fibre diameters of nanofibres

[0063] Table 4. Number of beads in nanofibres

[0064] Table 5. Response table for S / N ratios of nanofibre diameter Taguchi analysis

[0065] Table 6. Nanofibre diameter variance analysis results

[0066] The determination coefficient (r2) of the model was calculated as 95.52.

[0067] Table 7. Response table for S / N ratios of number of beads Taguchi analysis

[0068] Table 8. Variance analysis results for number of bead

[0069] The water volume, distance and needle diameter contained in the solvent were kept constant at the most appropriate levels suggested by Taguchi analysis results and the amount of methacrylic acid - methyl methacrylate copolymer (1 :1 ) was changed and the experiments were repeated with the ratios given in Table 5.

[0070] It is seen that the most effective and statistically significant (p<0.05) single parameter in terms of diameter thickness of nanofibres prepared with methacrylic acid - methyl methacrylate copolymer (1 :1 ) is its concentration. It was observed that there were two parameters responsible for the formation of beads and shown to be statistically effective (p<0.05), namely the amount of methacrylic acid - methyl methacrylate copolymer (1 :1 ) and the volume of DMF in the solvent. Although it was observed that the lower the amount of methacrylic acid - methyl methacrylate copolymer (1 :1 ) in the electrospinning solution, the thinner the fibre diameters were obtained, it was noticed that bead structures were formed despite the optimum DMF volume at the values where the amount of methacrylic acid - methyl methacrylate copolymer (1 :1 ) was 1300 mg and below. In addition, although there was no bead structure at 1350 mg and above, the levels of methacrylic acid - methyl methacrylate copolymer (1 :1 ) between 1300 mg and 1350 mg were tested again to obtain nanofibres with the thinnest fibre diameter without beads. Considering that the volume of DMF in the solvent has no statistical effect on the fibre thickness (p>0.05), the solution volume ratio, which is the most effective value in reducing the number of beads, and the other parameters were kept constant at the levels where the nanofibre diameter was smallest, and the experiments were repeated with the ratios in Table 9 by changing the amount of methacrylic acid - methyl methacrylate copolymer (1 :1).

[0071] Table 9. Parameters for examining fibre diameter changes

[0072] As a result of the formulation studies, the electrospinning parameters to obtain optimum wound dressings with the finest fibres without bead structure were determined as in Table 10.

[0073] Table 10. Optimum electrospinning parameters of nanofibres Electrospinning of the methacrylic acid - methyl methacrylate copolymer (1 :1 )-Rutin formulation was carried out at a flow rate of 3 mL / h and a voltage of 20 kV for 72 minutes and 48 seconds. Thus, wound dressings containing 500 mg polymer and 50 mg active ingredient were obtained from both formulations.

[0074] Table 11. Content of nanofibres (%w / w)

[0075] REFERENCES

[0076] [1] Braiman-Wiksman L, Solomonik I, Spira R, Tennenbaum T. Novel insights into wound healing sequence of events. Toxicol Pathol 2007;35(6):767-79.

[0077] [2] Tursen U. [Wound dressings for ulcer treatment], Turk J Dermatol 2013;7(1 ):61 -71 .

[0078] [3] Tort, S., & Acarturk, F. (2015). Yara Tedavisi ve Nanolif Yapismdaki Yara Ortuleri. Turkiye Klinikleri J Pharm Sci, 4(2), 68-78. https: / / doi.org / 10.5336 / pharmsci.2015- 45768

[0079] [4] Boateng, J.S., Matthew, s K.H., Stevens, H.N.E., Eccleston, G.M. “Wound Healing Dressings and Drug Delivery Systems: A Review” J. Pharm. Sci., 97(8), 2892-2923 (2007).

[0080] [5] Qakal C. Polihema Bazli Yara / Yamk Ortu Materyalleri. [Yuksek Lisans Tezi], Ankara, Turkiye: Hacettepe Universitesi Fen Bilimleri Enstitusu Biyomuhendislik Anabilim Dali; 2004.

[0081] [6] Francesko, A., Petkova, P., & Tzanov, T. (2018). Hydrogel dressings for advanced wound management. Current medicinal chemistry, 25(41 ), 5782- 5797.

Claims

CLAIMS1. Method of producing a nanofibre wound dressing by electrospinning for use in wound treatment, comprising the process steps of: i. preparing the solvent consisting of dimethylformamide (DMF) and methyl alcohol (MeOH) by simple mixing method, ii. adding methacrylic acid - methyl methacrylate copolymer (1 :1 ) to the solvent mixture containing DMF and MeOH, iii. beginning mixing the solution on a magnetic stirrer, iv. adding rutin hydrate active ingredient to the solution that is being mixed, v. keeping the solutions in an ultrasonic bath to remove air bubbles and making them suitable for electrospinning, and vi. spinning the solutions by electrospinning method.

2. A method according to claim 1 , comprising the steps of: i. preparing 10 mL of solvent consisting of DMF and MeOH by cold dissolution method, ii. adding methacrylic acid - methyl methacrylate copolymer (1 :1 ) to the solvent mixture containing DMF and MeOH, iii. beginning mixing the solution on the magnetic stirrer by means of a 20 mm long magnetic stir bar at a speed of 750 rpm and room temperature, iv. adding rutin hydrate active ingredient to the solution that is being mixed, v. keeping the solutions in an ultrasonic bath for 20 minutes to remove air bubbles and making them suitable for electrospinning, and vi. spinning the solutions by electrospinning method.

3. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises methacrylic acid - methyl methacrylate copolymer (1 :1 ), methyl alcohol (MeOH), DMF and rutin hydrate active ingredient.

4. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methylmethacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 140 mg rutin hydrate active ingredient.

5. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 140 mg rutin hydrate active ingredient.

6. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 140 mg rutin hydrate active ingredient.

7. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1400 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 140 mg rutin hydrate active ingredient.

8. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 135 mg rutin hydrate active ingredient.

9. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 135 mg rutin hydrate active ingredient.

10. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 135 mg rutin hydrate active ingredient.

11. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1350 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 135 mg rutin hydrate active ingredient.

12. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 130 mg rutin hydrate active ingredient.

13. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 130 mg rutin hydrate active ingredient.

14. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 130 mg rutin hydrate active ingredient.

15. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1300 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 130 mg rutin hydrate active ingredient.

16. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6 mL MeOH, 4 mL DMF and 125 mg rutin hydrate active ingredient.

17. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 6.5 mL MeOH, 3.5 mL DMF and 125 mg rutin hydrate active ingredient.

18. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7 mL MeOH, 3 mL DMF and 125 mg rutin hydrate active ingredient.

19. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1250 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 125 mg rutin hydrate active ingredient.

20. A method according to Claim 1 or Claim 2, wherein the electrospinning solution of said nanofibre wound dressing comprises 1375 mg methacrylic acid - methyl methacrylate copolymer (1 :1 ), 7.5 mL MeOH, 2.5 mL DMF and 137.5 mg rutin hydrate active ingredient.

21. A method according to Claims 1 or 2, wherein the solvent in step (i) comprises 6 mL of MeOH and 4 mL of DMF.

22. A method according to Claims 1 or 2, wherein the solvent in step (i) comprises6.5 mL of MeOH and 3.5 mL of DMF.

23. A method according to Claims 1 or 2, wherein the solvent in step (i) comprises 7 mL of MeOH and 3 mL of DMF.

24. A method according to Claims 1 or 2, wherein the solvent in step (i) comprises7.5 mL of MeOH and 2.5 mL of DMF.

25. Nanofibre wound dressing produced by a method according to any one of claims 20-24.

26. A nanofibre wound dressing according to Claim 25, comprising methacrylic acid - methyl methacrylate copolymer (1 :1) and rutin hydrate active ingredient.

27. A nanofibre wound dressing according to Claim 25, comprising 98-90.5% methacrylic acid - methyl methacrylate copolymer (1 :1 ) and 2-9.5% rutin hydrate.

28. A nanofibre wound dressing according to Claim 25, comprising a skeletal structure into which granulation tissue can grow.

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