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Title:
METHOD FOR PRODUCING HYPERBRANCHED POLYGLYCEROL COATINGS
Document Type and Number:
WIPO Patent Application WO/2023/035046
Kind Code:
A1
Abstract:
Provided is a method of coating a substrate with a hyperbranched polyglycerol. The hyperbranched polyglycerol is formed by activating the surface of the substrate followed by polymerisation of monomers of glycidol, or derivatives thereof. Also provided are substrates coated by the methods, methods for reducing thrombosis and/or fouling of products using the coatings, methods for increasing hydrophilicity using the coatings, and products with reduced thrombosis, fouling and/or increased hydrophilicity.

Inventors:
MOORE ELI (AU)
AL-BATAINEH SAMEER (AU)
SMITH LOUISE ELIZABETH (AU)
Application Number:
PCT/AU2022/051104
Publication Date:
March 16, 2023
Filing Date:
September 13, 2022
Export Citation:
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Assignee:
TEKCYTE LTD (AU)
International Classes:
A61L27/34; A61L29/08; A61L31/10; A61L33/06; A61P7/02; B01D65/08; C08G65/22; C08J7/056; C09D5/16; C09D171/02
Domestic Patent References:
WO2017156592A12017-09-21
WO2017156593A12017-09-21
Foreign References:
EP2123731A12009-11-25
US6169127B12001-01-02
US7201935B12007-04-10
US20130095999A12013-04-18
Other References:
BURZAVA ANOUCK L. S., JASIENIAK MAREK, COCKSHELL MICHAELIA P., VOELCKER NICOLAS H., BONDER CLAUDINE S., GRIESSER HANS J., MOORE EL: "Surface-Grafted Hyperbranched Polyglycerol Coating: Varying Extents of Fouling Resistance across a Range of Proteins and Cells", ACS APPLIED BIO MATERIALS, AMERICAN CHEMICAL SOCIETY, US, vol. 3, no. 6, 15 June 2020 (2020-06-15), US , pages 3718 - 3730, XP093047201, ISSN: 2576-6422, DOI: 10.1021/acsabm.0c00336
WANG C.X., REN Y., LV J.C., ZHOU Q.Q., MA Z.P., QI Z.M., CHEN J.Y., LIU G.L., GAO D.W., LU Z.Q., ZHANG W., JIN L.M.: "In situ synthesis of silver nanoparticles on the cotton fabrics modified by plasma induced vapor phase graft polymerization of acrylic acid for durable multifunction", APPLIED SURFACE SCIENCE, ELSEVIER, AMSTERDAM , NL, vol. 396, 1 February 2017 (2017-02-01), Amsterdam , NL , pages 1840 - 1848, XP093047204, ISSN: 0169-4332, DOI: 10.1016/j.apsusc.2016.11.173
TALEMI PEJMAN, DELAIGUE MARINE, MURPHY PETER, FABRETTO MANRICO: "Flexible Polymer-on-Polymer Architecture for Piezo/Pyroelectric Energy Harvesting", APPLIED MATERIALS & INTERFACES, AMERICAN CHEMICAL SOCIETY, US, vol. 7, no. 16, 29 April 2015 (2015-04-29), US , pages 8465 - 8471, XP093047205, ISSN: 1944-8244, DOI: 10.1021/am5089082
Attorney, Agent or Firm:
PHILLIPS ORMONDE FITZPATRICK (AU)
Download PDF:
Claims:
59

CLAIMS:

1. A method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising: exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof, and polymerising the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the substrate.

2. The method according to claim 1, wherein the substrate comprises a metal, a metal-alloy, a metal oxide, a polymer, a glass, a ceramic, or a combination of any one or more of the aforementioned substrates.

3. The method according to claims 1 or 2, wherein the vapour substantially comprises glycidol monomers.

4. The method according to any one of claims 1 to 3, wherein the vapour comprises gaseous monomers of glycidol and/or a derivative thereof.

5. The method according to any one of claims 1 to 4, wherein the surface is activated by plasma activation.

6. The method according to any one of claims 1 to 4, wherein the surface is activated by chemical activation.

7. The method according to any one of claims 1 to 6, wherein the vapour does not comprise any further chemicals which initiate polymerisation.

8. The method according to any one of claims 1 to 7, wherein the monomers are produced from a source of liquid glycidol and/or a derivative thereof in a reaction vessel.

9. The method according to claim 8, wherein the monomers are produced by heating a source of liquid glycidol and/or a derivative thereof in the reaction vessel. 60

10. The method according to any one of claims 1 to 7, wherein the monomers are produced in a reaction vessel where the polymerisation of the monomers on the activated surface occurs.

11. The method according to any one of claims 1 to 7, wherein the vapour is introduced from an external source into a reaction vessel where the polymerisation of the monomers on the activated surface occurs.

12. The method according to any one of claims 1 to 11, wherein the surface is activated and the polymerisation of the monomers occurs in the same reaction vessel.

13. The method according to any one of claims 1 to 12, wherein the exposing of the activated surface to the vapour is continuous exposure to the vapour.

14. The method according to any one of claims 1 to 13, wherein the exposing of the activated surface to the vapour comprises a temperature in the range from 50°C to 140°C.

15. The method according to any one of claims 1 to 14, wherein the exposing of the activated surface to the vapour comprises a time of 72 hours or less.

16. The method according to any one of claims 1 to 15, wherein the hyperbranched polyglycerol coating produced on the substrate comprises a thickness of at least 1 nm.

17. The method according to any one of claims 1 to 16, wherein the coating produced by the method has a characteristic of reduced fouling to biological materials or a reduced fouling and/or thrombosis associated with use of the coating in a medical setting.

18. A method according to any one of claims 1 to 17, comprising the step of: activating a surface of the substrate prior to exposing the activated surface to the vapour comprising monomers of glycidol and/or a derivative thereof.

19. A method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising reacting a vapour comprising monomers of glycidol and/or a derivative thereof with an activated surface of the substrate and producing the 61 hyperbranched polyglycerol coating by polymerisation of the monomers on the activated surface.

20. A method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising polymerising monomers of glycidol and/a derivative thereof in a vapour on an activated surface of the substrate and thereby coating the substrate with the hyperbranched poly glycerol.

21. The method according to any one of claims 1 to 20, wherein the method is used: to reduce fouling of the substrate; to decrease attachment of proteins, microbial matter, organic matter and/or cells to the substrate; to decrease fouling of a medical device; to reduce formation of clots; to decrease fouling of filtration membranes; to decrease fouling of tubing; to decrease fouling of liquid handling equipment; to increase hydrophilicity of the substrate; and/or to decrease fouling of a product comprising the substrate.

22. A substrate coated with a hyperbranched polyglycerol produced by the method of any one of claims 1 to 21.

23. The substate according to claim 22, wherein the coated substrate is an antifouling and/or anti-thrombotic substrate.

24. A substrate coated with a hyperbranched polyglycerol, the coating produced by exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or derivative thereof and polymerisation of the monomers on the activated surface.

25. A product comprising a substrate according to any one of claims 22 to 24.

26. The product according to claim 25, wherein the product is a medical device, a stent, an implantable product, a vascular device, a graft, a valve, tubing, a water handling device, a membrane, or a filter.

27. A method of reducing fouling of a substrate, the method comprises exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a 62 derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

28. A substrate with reduced fouling produced by the method according to claim 27.

29. A method of reducing fouling and/or thrombosis of a substrate for use in a medical setting, the method comprises exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

30. A method of producing a substrate with reduced fouling and/or thrombosis for use in a medical setting, the method comprises exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers and thereby producing a substrate with reduced fouling.

31. A substrate with reduced fouling and/or thrombosis produced by the method according to claim 29 or claim 30.

32. A method of producing a product with reduced fouling, the method comprising exposing one or more activated surfaces of the product to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched poly glycerol coating on the one or more activated surfaces of the product by polymerisation of the monomers, thereby producing a product with reduced fouling.

33. A product with reduced fouling produced by the method according to claim 32.

34. A method of producing a medical product with reduced fouling and/or thrombosis associated with the product, the method comprising exposing one or more activated surfaces of the product to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the one or more activated surfaces of the product by polymerisation of the monomers, thereby producing a medical product with reduced fouling and/or thrombosis. 63

35. A medical product with reduced fouling and/or thrombosis produced by the method according to claim 34.

36. A system for coating a substrate with a hyperbranched polyglycerol, the system comprising a plasma activation system comprising a reaction vessel and a means to provide a vaporised liquid to the reaction vessel.

37. The system according to claim 36, wherein the liquid comprises glycidol and/or a derivative thereof.

38. The system according to claim 36 or claim 37, wherein the liquid is vaporised in the reaction vessel.

39. A method of coating a substrate with a hyperbranched polyglycerol, the method comprising using a system according to any one of claims 36 to 38 to activate the substrate and coat the activated substrate with the hyperbranched polyglycerol by polymerisation of gaseous monomers of glycidol and/or a derivative thereof from the source of liquid glycidol.

Description:
METHOD FOR PRODUCING HYPERBRANCHED POLYGLYCEROL COATINGS.

[0001] This application claims priority to Australian provisional application 2021902950 filed on 13 September 2021, the entire content of which is incorporated herein.

FIELD

[0002] The present disclosure relates to methods for producing a hyperbranched polyglycerol coating on a substrate. The present disclosure also relates to substrates coated by the methods, methods for reducing thrombosis and/or fouling of products using the coatings, methods for increasing hydrophilicity using the coatings, and products with reduced thrombosis, fouling and/or increased hydrophilicity.

BACKGROUND

[0003] There are a variety of situations where it would be advantageous to reduce the accumulation of biological materials on surfaces. For example, medical devices implanted into a subject typically suffer from a loss of performance, functionality and/or longevity after implantation, in part due to interactions of the device with the tissue and/or fluids of the subject. A similar consideration applies to medical equipment used in contact with biological fluids.

[0004] The use of membranes for filtering aqueous based materials is another situation where there is a loss of performance over time due to accumulation of biological and non- biological material on filters, necessitating the cleaning or replacement of the filters to address the fouling.

[0005] In the case of medical devices, there are a number of types of devices that are subject to reduced functionality overtime. For example, synthetic vascular grafts are used in a variety of peripheral, aortic and vascular access procedures. However, such devices often suffer a loss of performance or function over time from the effects of cell attachment, hyperplasia and thrombus formation associated with the grafts.

[0006] Similarly, stents or stent grafts are a commonly used medical device for the treatment of a number of conditions, such as their use in angioplasty to improve blood flow to narrowed or blocked coronary arteries, their use for peripheral artery angioplasty to treat atherosclerotic narrowing of the abdominal, leg and renal arteries and veins caused by peripheral artery disease, and their use to assist in the treatment of aneurysms. However, not only do stents and grafts suffer a loss of function over time, but they also carry a risk of stent associated thrombosis due to clots forming in the stent or graft.

[0007] The ability of medical devices such as grafts and stents to resist one or more of platelet attachment, cell attachment and fouling may have important effects on their usable lifespan and to reduce the possibility of adverse effects occurring in a patient.

[0008] Device failure also often occurs as a result of complication through the build-up of fibrotic tissue around the implanted device, which results from a reaction to the foreign material. Similarly, inflammation may result from the introduction of medical devices to various parts of the body.

[0009] As such, there is a need to improve the surface properties of many substrates to reduce fouling. Specifically, there is a need to improve medical devices to reduce biological material and microbial biofilm attaching to the surface and causing unwanted side-effects or reducing the performance of the medical device. In addition, there is a need to improve the surface properties of filters and similar types of membranes to reduce fouling by microorganisms and organic matter.

SUMMARY

[0010] The present disclosure relates to methods for producing a hyperbranched polyglycerol coating on a substrate. The present disclosure also relates to substrates coated by the methods, methods for reducing thrombosis and/or fouling of products using the coatings, methods for increasing hydrophilicity using the coatings, and products with reduced thrombosis, fouling and/or increased hydrophilicity.

[0011] Certain embodiments of the present disclosure provide a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof, and polymerizing the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the substrate.

[0012] Certain embodiments of the present disclosure provide a method of producing a hyperbranched polyglycerol coating on a substrate, including activating the surface of the substrate. Accordingly, such embodiments of the present disclosure provide a method comprising: activating a surface of the substrate; exposing the activated surface to a vapour comprising monomers of glycidol and/or a derivative thereof; and polymerising the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the substrate.

[0013] Certain embodiments of the present disclosure provide a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising reacting a vapour comprising monomers of glycidol and/or a derivative thereof with an activated surface of the substrate and producing the hyperbranched polyglycerol coating by polymerisation of the monomers on the activated surface.

[0014] Certain embodiments of the present disclosure provide a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising polymerising monomers of glycidol and/a derivative thereof in a vapour on an activated surface of the substrate and thereby coating the substrate with the hyperbranched polyglycerol.

[0015] Certain embodiments of the present disclosure provide a substrate coated with a hyperbranched polyglycerol produced by a method as described herein.

[0016] Certain embodiments of the present disclosure provide a substrate coated with a hyperbranched polyglycerol, the coating produced by exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or derivative thereof and polymerisation of the monomers on the activated surface.

[0017] Certain embodiments of the present disclosure provide a product comprising a coated substrate as described herein. [0018] Certain embodiments of the present disclosure provide a method of reducing fouling of a substrate, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

[0019] Certain embodiments of the present disclosure provide a method of producing a substrate with reduced fouling, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers and thereby producing a substrate with reduced fouling.

[0020] Certain embodiments of the present disclosure provide a substrate with reduced fouling produced by a method as described herein.

[0021] Certain embodiments of the present disclosure provide a method of reducing fouling and/or thrombosis of a substrate for use in a medical setting, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

[0022] Certain embodiments of the present disclosure provide a method of producing a substrate with reduced fouling and/or thrombosis for use in a medical setting, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers and thereby producing a substrate with reduced fouling.

[0023] Certain embodiments of the present disclosure provide a substrate with reduced fouling and/or thrombosis produced by a method as described herein.

[0024] Certain embodiments of the present disclosure provide a method of producing a product with reduced fouling, the method comprising exposing one or more activated surfaces of the product to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the one or more activated surfaces of the product by polymerisation of the monomers, thereby producing a product with reduced fouling.

[0025] Certain embodiments of the present disclosure provide a product with reduced fouling produced by a method as described herein.

[0026] Certain embodiments of the present disclosure provide a method of producing a medical product with reduced fouling and/or thrombosis associated with the product, the method comprising exposing one or more activated surfaces of the product to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the one or more activated surfaces of the product by polymerisation of the monomers, thereby producing a medical product with reduced fouling and/or thrombosis.

[0027] Certain embodiments of the present disclosure provide a medical product with reduced fouling and/or thrombosis produced by a method as described herein.

[0028] Certain embodiments of the present disclosure provide a system for coating a substrate with a hyperbranched polyglycerol, the system comprising a plasma activation system comprising a reaction vessel and a means to vaporise a liquid in the reaction vessel.

[0029] In certain embodiments, the system comprises an operation unit which initiates the means to vaporise the liquid in the reaction vessel after the plasma activation system has activated the substrate, and preferably after the plasma activation system has ceased.

[0030] Certain embodiments of the present disclosure provide a method of coating a substrate with a hyperbranched polyglycerol, the method comprising using a system as described herein to activate the substrate followed by coating the activated substrate with the hyperbranched polyglycerol by polymerising gaseous monomers of glycidol and/or a derivative thereof from the source of liquid glycidol.

[0031] Other embodiments are described herein. BRIEF DESCRIPTION OF THE FIGURES

[0032] Certain embodiments are illustrated by the following figures. It is to be understood that the following description is for the purpose of describing particular embodiments only and is not intended to be limiting with respect to the description.

[0033] Figure 1 shows a summary of XPS results collected on the hyperbranched polyglycerol (HPG) coated PTFE cannulas (A) and PTFE sheets (B) as a function of plasma process gas mixing ratio. The other HPG coating conditions were: gas flow rate- high, plasma power- 100W, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h, remained constant. The bars from left to right represent - 0% B: 100% A, 25% B: 75% A; 50% B: 50% A; 75% B: 25% A; 100% B: 0% A.

[0034] Figure 2 shows a summary of XPS C is curve-fitting results of the HPG coated PTFE cannulas (A) and PTFE sheets (B) as a function of plasma process gas mixing ratio. The other HPG coating conditions: gas flow rate-high, plasma power-lOOW, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h, remained constant. The bars from left to right represent - 0% B: 100% A, 25% B: 75% A; 50% B: 50% A; 75% B: 25% A; 100% B: 0% A.

[0035] Figure 3 shows a summary of WCA results collected on HPG coated PTFE sheets as a function of plasma process gas mixing ratio. Other HPG coating conditions: gas flow rate-high 4 seem), plasma power- 100W, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h.

[0036] Figure 4 shows a summary of WCA results collected on HPG coated PTFE cannulas as a function of plasma process gas mixing ratio. Other HPG coating conditions: gas flow rate-high (4 seem), plasma power- 100W, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h.

[0037] Figure 5 shows a summary of XPS results collected on HPG coated PTFE cannula (A) and PTFE sheets (B) as a function of plasma process gas flow rate. Other HPG coating conditions: Process gas-Argon, plasma power- 100W, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; high flow rate (4 seem); and low flow rate (2 seem).

[0038] Figure 6 shows a summary of XPS C Is curve-fitting results of HPG coated PTFE cannulas (A) and PTFE sheets (B) as a function of plasma process gas flow rate. Other HPG coating conditions: Process gas-Argon, plasma power-lOOW, plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; high flow rate (4 seem); and low flow rate (2 seem).

[0039] Figure 7 shows a summary of XPS results collected on HPG coated PTFE cannula (A) and PTFE sheets (B) as a function of applied power. Other HPG coating conditions: Process gas-Argon, low flow rate (2 seem), plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; 50W; 100W; 200W; and 400W.

[0040] Figure 8 shows a summary of XPS C Is curve-fitting results of HPG coated PTFE cannulas (A) and PTFE sheets (B) as a function of applied power. Other HPG coating conditions: Process gas-Argon, low flow rate (2 seem), plasma activation time- 20min, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; 50W; 100W; 200W; and 400W.

[0041] Figure 9 shows a summary of WCA results collected on HPG coated PTFE sheets as a function of applied power. Other HPG coating conditions: Process gas- Argon, low flow rate (2 seem), plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h.

[0042] Figure 10 shows a summary of WCA results collected on HPG coated PTFE cannula as a function of applied power. Other HPG coating conditions: Process gas- Argon, low flow rate (2 seem), plasma activation time-20min, HPG grafting temperature- 100°C and HPG grafting time-24h.

[0043] Figure 11 shows a summary of XPS results collected on HPG coated PTFE cannulas (A and B) and PTFE sheets (C and D) as a function of plasma activation time. Other HPG coating conditions: Process gas-Argon, low flow rate (2 seem), RF power: 200W and 400W, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; Imin; 5min; lOmin; and 20min.

[0044] Figure 12 shows a summary of XPS C Is curve-fitting results of HPG coated PTFE cannula (A and B) and PTFE sheets (C and D) as a function of plasma activation time. Other HPG coating conditions: Process gas-Argon, low flow rate (2 seem), RF power: 200W and 400W, HPG grafting temperature- 100°C and HPG grafting time-24h. The bars from left to right represent - untreated; Imin; 5min; lOmin; and 20min.

[0045] Figure 13 shows XPS spectrum data for HPG polymerisation on woven PET.

[0046] Figure 14 shows XPS spectrum data for HPG polymerisation on woven PET using oxygen or argon plasma conditions.

[0047] Figure 15 shows SEM images of untreated PET fibre as compared to argon plasma treated PET fibre and oxygen plasma treated PET Fibre.

[0048] Figure 16 shows in panel A SEM images of argon plasma treated HPG polymerisation of PET fibres after 2 hrs (left), 4 hrs (right) and 24 hrs (bottom) as compared to oxygen plasma treated PET Fibre (Panel B) and 24 h HPG polymerisation.

[0049] Figure 17 shows XPS data for different sample volumes of glycidol.

[0050] Figure 18 shows static blood assay using fluoresce microscopy. Figure 18A shows static blood assay on control PET. Left image - edge of the blood drop and right image - centre of the blood drop. Bright green is fibrin and cells stained with CFSE. Static blood assay on 2h HPG initiated with argon plasma (left) and oxygen plasma (right). Figure 18B shows static blood assay on 2h HPG initiated with argon plasma (left) and oxygen plasma (right). Figure 18C shows static blood assay on 4h HPG, initiated with argon plasma (left) and oxygen plasma (right). Figure 18D shows static blood assay on 24h HPG at 65°C; initiated with oxygen plasma on the bottom of shelf (left and top shelf (right). [0051] Figure 19 shows SEM analysis of static blood on various PET substrates. Panel A shows static blood on control PET (1000X and 5000X magnification). Panel B shows static blood on oxygen plasma initiated; 24h polymerisation in 2.5L SS box (lOOOx and 5000x magnification). Panel C shows static blood on oxygen plasma initiated; 24h polymerisation in 10 mL plastic tube (lOOOx and 5000x magnification). Panel D shows static blood on oxygen plasma initiated; 24h polymerisation in 25 mL glass tube (lOOOx and 5000x magnification). Panel E shows static blood on oxygen plasma initiated; 4h polymerisation in 2.5 L SS box (lOOOx and 5000x magnification). Panel F shows static blood on oxygen plasma initiated; 4h polymerisation in 10 mL plastic tube (lOOOx and 5000x magnification). Panel G shows static blood on oxygen plasma initiated; 4h polymerisation in 25 mL glass tube (lOOOx and 5000x magnification).

[0052] Figure 20 shows a summary of XPS elemental concentrations measured on the HPG coated PET grafts. The grafts were plasma activated under A: condition 1 (oxygen plasma activation) and B: condition 2 (argon plasma activation) parameters. XPS analyses were performed on the inner and outer surface of the grafts. Three separate regions were analysed on each side: namely edge, middle and edge. The bars of figure 20A, from left to right represent - PET graft (APEX), untreated; HPG/PET graft (APEX), inner surface, edge, condition 1; HPG/PET graft (APEX), outer surface, edge, condition 1 : HPG/PET graft (APEX), inner surface, middle, condition 1; HPG/PET graft (APEX), outer surface, middle, condition 1; HPG/PET graft (APEX), inner surface, edge, condition 1; HPG/PET graft (APEX), outer surface, edge, condition 1. The bars of figure 20B, from left to right represent - PET graft (APEX), untreated; HPG/PET graft (APEX), inner surface, edge, condition 2; HPG/PET graft (APEX), outer surface, edge, condition 2: HPG/PET graft (APEX), inner surface, middle, condition 2; HPG/PET graft (APEX), outer surface, middle, condition 2; HPG/PET graft (APEX), inner surface, edge, condition 2; HPG/PET graft (APEX), outer surface, edge, condition 2.

[0053] Figure 21 shows a summary of curve-fitting analyses results of the XPS high- resolution C is spectra of HPG coated PET grafts. The grafts were plasma activated under condition 1 - oxygen (A) or condition 2 - argon (B) parameters. XPS analyses were performed on the inner and outer surface of the grafts. Three separate regions were analysed on each side: namely edge, middle and edge. The bars of figure 21A, from left to right represent - PET graft (APEX), untreated; HPG/PET graft (APEX), inner surface, edge, condition 1; HPG/PET graft (APEX), outer surface, edge, condition 1 : HPG/PET graft (APEX), inner surface, middle, condition 1; HPG/PET graft (APEX), outer surface, middle, condition 1; HPG/PET graft (APEX), inner surface, edge, condition 1; HPG/PET graft (APEX), outer surface, edge, condition 1. The bars of figure 2 IB, from left to right represent - PET graft (APEX), untreated; HPG/PET graft (APEX), inner surface, edge, condition 2; HPG/PET graft (APEX), outer surface, edge, condition 2: HPG/PET graft (APEX), inner surface, middle, condition 2; HPG/PET graft (APEX), outer surface, middle, condition 2; HPG/PET graft (APEX), inner surface, edge, condition 2; HPG/PET graft (APEX), outer surface, edge, condition 2.

[0054] Figure 22 shows the force-displacement curves for the tensile testing of the PET graft. Displacement rate = 25.4 mm/min, sample length 100 mm, gauge length 40 mm. Control: Dotted lines (n = 4), Condition 1 - oxygen: dashed lines (n = 5), Condition 2 - argon: solid lines (n = 5).

[0055] Figure 23 shows representative images of fibrin networks and bound blood cells across uncoated PET samples and differing magnifications.

[0056] Figure 24 shows representative images of blood components bound to HPG- coated PET produced using condition 1 - oxygen (top) and condition 2 - argon (bottom).

[0057] Figure 25 shows a summary of XPS results of elemental proportions on the surface of uncoated and HPG-coated silicone (PDMS) sheet, that were activated at either 100 W (A) or 200 W (B) power during plasma activation. Data was collected from samples that were freshly coated and aged for up to 4 weeks at room temperature. The bars from left to right represent - Silicone sheet, untreated; HPG/Silicone sheet (fresh); HPG/Silicone sheet (1 week old); HPG/Silicone sheet (2 weeks’ old); and HPG/Silicone sheet (4 weeks’ old).

[0058] Figure 26 shows the XPS Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated PDMS, that were activated with either 100 W (A) or 200 W (B) power during plasma activation. Data was collected from samples that were freshly coated and aged for up to 4 weeks at room temperature. The bars from left to right represent - Silicone sheet, untreated; HPG/Silicone sheet (fresh); HPG/Silicone sheet (1 week old); HPG/Silicone sheet (2 weeks’ old); and HPG/Silicone sheet (4 weeks’ old).

[0059] Figure 27 shows the WCA values for uncoated and HPG-coated PDMS, that were activated with either 100 W (A) or 200 W (B) power during plasma activation. Data was collected from samples that were freshly coated and aged for up to 4 weeks at room temperature.

[0060] Figure 28 shows the XPS results of elemental proportions on the surface of uncoated and HPG-coated stainless steel sheet (A) and the Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated stainless steel sheet (B). The bars from left to right represent - SS sheet, untreated; HPG/SS sheet, 80°C; HPG/SS sheet, 100°C.

[0061] Figure 29 shows the XPS results of elemental proportions on the surface of uncoated and HPG-coated cobalt/chromium stent (A) and the Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated cobalt/chromium stent (B). The bars from left to right represent - CoCr stent, untreated; HPG/CoCr stent, 80°C; HPG/CoCr stent, 100°C.

[0062] Figure 30 shows the XPS results of elemental proportions on the surface of uncoated and HPG-coated polyvinylidene difluoride (PVDF) filtration membranes (A) and the Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated PVDF membranes (B). The bars from left to right represent - PVDF membrane (0.45pm), untreated; HPG/PVDF membrane (0.45pm), 100W; and HPG/PVDF membrane (0.45pm), 200W.

[0063] Figure 31 shows XPS results of elemental proportions on the surface of uncoated and HPG-coated PTFE filtration membranes (A) and the Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated PTFE membranes (B). The bars from left to right represent - PTFE membrane (0.45pm), untreated; HPG/ PTFE membrane (0.45pm), 100W; and HPG/ PTFE membrane (0.45pm), 200W.

[0064] Figure 32 shows XPS results of elemental proportions on the surface of uncoated and HPG-coated polypropylene (PP) filtration membranes (A) and the Cis spectrum analysis of the functional groups on the surface of uncoated and HPG-coated PP membranes (B). The bars from left to right represent - PP membrane (0.45 pm), untreated; HPG/PP membrane (0.2pm), lOmins; HPG/PP membrane (0.2pm), 5mins; and HPG/PP membrane (0.2pm), 2mins.

[0065] Figure 33 shows the transmembrane pressure profile of uncoated PVDF (Pristine membrane - upper line) and HPG-coated PVDF membrane (Coated membrane - lower line) under long-term filtration using humic acid (A) and sodium alginate (B).

[0066] Figure 34 shows the resistance to fouling after the uncoated and HPG-coated membranes are cleaned with sodium hydroxide (NaOH) and citric acid after long-term filtration with humic acid (A) or sodium alginate (B). The bars from left to right represent - End of filtration; after NaOH cleaning; and after citric acid cleaning.

DETAILED DESCRIPTION

[0067] The present disclosure relates to methods for producing a hyperbranched polyglycerol coating on a substrate. The present disclosure also relates to substrates coated by the methods, methods for reducing fouling and/or thrombosis using the coatings, and products with reduced fouling and/or thrombosis.

[0068] Certain embodiments of the present disclosure provide a method of producing a hyperbranched polyglycerol coating on a substrate.

[0069] In certain embodiments, the present disclosure provides a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof, and polymerising the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the substrate.

[0070] The term “hyperbranched polyglycerol” as used herein refers to a branched aliphatic polyether with hydroxyl end groups. It will be appreciated that the term also includes a branched polyether in which a proportion of the hydroxyl end groups have been derivatised and/or replaced with a suitable group. [0071] In certain embodiments, the substrate comprises a metal substrate, a metal-alloy substrate, a metal oxide, a polymer substrate, a glass substrate, a ceramic substrate, or a combination of any one or more of the aforementioned substrates. Other types of substrates are contemplated.

[0072] The term “metal” as used herein refers to a substrate comprising a metallic material, such as a pure metal, a metal alloy, or a mixture of one or more metals and/or other materials. It will be understood that metals, when exposed to an environment containing oxygen, will result in the formation of a metal oxide. Hence, it will be understood to a person skilled in the art that the methods and examples disclosed herein inherently encompass metal oxides. For example, a metal substrate may be composed entirely of a metal, metal oxide or a metal alloy, or may be composed in part of a metallic material and one or more other materials. Such other materials may include fluoropolymers, such as those listed herein.

[0073] Examples of metals include titanium, nickel, cobalt, chromium, lithium, iron, aluminium, manganese, niobium and tantalum. Other types of metals are contemplated.

[0074] In certain embodiments, the substrate comprises, consists essentially of, or consists of a substantially pure metal.

[0075] In certain embodiments, the substrate comprises, consists essentially of, or consists of a metal oxide.

[0076] In certain embodiments, the substate comprises a metal alloy or an alloy of a metal oxide.

[0077] Examples of metal alloys include an iron containing alloy, a nickel containing alloy, a titanium containing alloy, a cobalt contain alloy, or a chromium containing alloy. In certain embodiments, the substrate comprises an iron chromium alloy, a nickel titanium alloy or a cobalt chromium alloy. Other types of alloys are contemplated.

[0078] The term "polymer” and the related terms such as “polymeric” or “polymer- based”, as used herein in relation to the substrate, refers to a substrate that comprises one or more chemical compounds made up of a plurality of repeating similar structural units. Examples of polymeric materials include synthetic materials made of organic polymers (such as plastics and resins), and natural materials such as silk, wool, cellulose, rubber and biological macromolecules. The polymer may also contain one or more non- polymeric materials.

[0079] In certain embodiments, the polymeric substrate comprises a thermoplastic, an elastomer, a thermoset, or a fibre.

[0080] In certain embodiments, the polymeric substrate comprises one or more of a polysil oxane, a fluoropolymer, a polyester and/or a polyurethane. Other types of polymers are contemplated.

[0081] In certain embodiments, the polymeric substrate comprises one or more polysilicones.

[0082] Examples of polysilicones include polydimethylsiloxane, polydi(trifluoropropyl)siloxane, poly di vinyl siloxane, polydiphenylsiloxane, and copolymers of the aforementioned polysilicones. Polysilicones are commercially available or may be produced by a method known in the art.

[0083] In certain embodiments, the polymeric substrate comprises one or more fluoropolymers. Fluoropolymers are commercially available or may be synthesized by a method known in the art.

[0084] Examples of fluoropolymers comprise one or more a PVF (polyvinylfluoride), a PVDF (polyvinylidene fluoride), a PTFE (polytetrafluoroethylene), a PCTFE (polychlorotrifluoroethylene), a PFA/MFA (perfluoroalkoxy polymer), a FEP (fluorinated ethyl ene-propylene), an ETFE (polyethylenetetrafluoroethylene), an ECTFE (polyethylenechlorotrifluoroethylene), a FFPM/FFKM (perfluorinated elastomer), a FPM/FKM (fluorocarbon [chlorotrifluoroethylenevinylidene fluoride]), a FEPM (tetrafluoroethylene-propylene), a PFPE (perfluoropoly ether), and a PF SA (perfluorosulfonic acid) and a perfluoropolyoxetane. Other fluoropolymers are contemplated.

[0085] In certain embodiments, the polymeric substrate comprises a polyester. Polyesters are commercially available or may be synthesized by a method known in the art.

[0086] Examples of polyesters comprise one or more of a polyglycolide or polyglycolic acid (PGA), a polylactic acid (PLA), a poly caprolactone (PCL), a polyhydroxyalkanoate (PHA), a polyhydroxybutyrate (PHB), a polyethylene adipate (PEA), a polybutylene succinate (PBS), a poly(3-hydroxybutyrate-co-3 -hydroxy valerate) (PHBV), a polyethylene terephthalate (PET), a polybutylene terephthalate (PBT), a polytrimethylene terephthalate (PTT), a polyethylene naphthalate (PEN), and Vectran. Other polyesters are contemplated.

[0087] In certain embodiments, the polymeric substate comprises a polyurethane. Examples of polyurethanes include one or more of thermoplastic polyurethane, thermoplastic polycarbonate-urethane (PCU), segmented polyurethane (SPU), thermoplastic silicone-polycarbonate-urethane (TSPCU), thermoplastic polyetherurethane (TPU), and thermoplastic Sili cone-Poly ether-urethane (TSPU). Other polyurethanes are contemplated.

[0088] The term “glycidol” as used herein refers to the chemical compound oxiranylmethanol, otherwise referred to by other chemical names such as 2,3-epoxy-l- propanol, 3 -hydroxypropylene oxide, epoxypropyl alcohol, hydroxymethyl ethylene oxide or 2-hydroxymethyl oxirane. Glycidol is commercially available or may be synthesized by a method known in the art such as the epoxidation of allyl alcohol. Derivatives of glycidol include substituted derivatives at one or more of the 1, 2, and/or 3 positions of the alkane chain, such as hydroxy, halogen, alkyl, alkenyl, alkynal, aryl, acyl, nitro, amino, ether or ester substituted derivatives, which are either commercially available or may be synthesized by a method known in the art.

[0089] The term “vapour” as used herein refers to a substance in a gaseous form, a substance in an evaporated form, a liquid substance heated to be in a gaseous form, or a substance normally in liquid or solid form which is mixed, diffused or suspended in another gas, and may be present in substantially pure form or be mixed, diffused or suspended in one or more other gaseous materials. [0090] In certain embodiments, the vapour comprises substantially pure glycidol monomers and/or a derivative thereof. In certain embodiments, the vapour comprises substantially pure glycidol monomers. In certain embodiments, the vapour comprises monomers of glycidol and/or a derivative thereof in the presence of one or more other substances. In certain embodiments, the vapour comprises glycidol monomers and an inert vapour or gas.

[0091] In certain embodiments, the vapour comprises 100%, at least 99%, at least 98%, at least 97%, at least 95%, at least 93%, at least 90%, at least 80%, at least 70%, at least 60%, or at least 50% monomer of glycidol and/or a derivative thereof. In certain embodiments, the vapour comprises 50% or greater, 60% or greater, 70% or greater, 80% or greater, 90% or greater, or 95% or greater monomers of glycidol and/or a derivative thereof. Other amounts are contemplated.

[0092] In certain embodiments, the hyperbranched polyglycerol coating is formed on the activated surface of the substrate, the activated substrate produced by a method comprising one or more of activation by plasma treatment, covalent bonding directly to the substrate, or covalent bonding indirectly to the substrate.

[0093] In some embodiments, activation of the surface comprises the formation of a surface with an oxygen containing group or a nitrogen containing group. Accordingly, in some embodiments, the process of activation may encompass any treatment (such as a chemical treatment) that results in an activated surface, such as those described. In some embodiments, providing and oxygen containing group or nitrogen containing group comprises providing a hydroxyl functional group or amine functional group.

[0094] In certain embodiments, the hyperbranched polyglycerol coating is formed directly on the activated surface of the substrate.

[0095] In certain embodiments, the hyperbranched polyglycerol coating is formed directly on a plasma activated surface of the substrate. In certain embodiments, the hyperbranched polyglycerol coating is formed indirectly on a plasma activated surface of the substrate. In certain embodiments, a surface of the substrate polymeric material is activated by plasma treatment and the coating is formed on the activated surface. [0096] In certain embodiments, the hyperbranched polyglycerol coating is formed on a functionalised surface of the substrate. In certain embodiments, a surface of the substrate is functionalised and the coating is formed on the functionalised substrate.

[0097] Methods for functionalisation of surfaces/substrates are known in the art. For example, a polyurethane substrate may be functionalised by treatment with a diisocyanate to introduce free isocyanate groups for coupling.

[0098] Chemical methods for activating a substrate to allow formation of a hyperbranched polyglycerol coating (directly or indirectly) on the material are known in the art.

[0099] In certain embodiments, the coating is formed from a method involving chemical activation of the substrate. Other methods are contemplated.

[0100] In certain embodiments, the hyperbranched polyglycerol coating is formed on a surface of the substrate activated by plasma treatment.

[0101] In certain embodiments, the substrate is activated by plasma treatment and the coating is formed (directly or indirectly) on the activated substrate. In certain embodiments, the substrate is activated by plasma treatment and the coating is formed directly on the activated substrate.

[0102] Examples of plasma treatment include radio frequency induced plasma treatment, corona plasma treatment, glow discharge plasma treatment, plasma immersion ion implantation, low pressure plasma treatment, and atmospheric pressure plasma treatment. Methods for plasma treatment of materials or substrates to form plasma modified/activated surfaces are known in the art.

[0103] In certain embodiments, the method comprises activating the surface of the substrate by plasma treatment and forming the hyperbranched polyglycerol coating by contacting the activated substrate with glycidol monomers in vapour to initiate polymerisation of the monomers.

[0104] In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of a gas. Examples of envisaged gases comprise one of more of oxygen, argon, nitrogen, and air and combinations thereof. Other gases are contemplated.

[0105] In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of oxygen. In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of nitrogen. In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of argon. In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of air.

[0106] In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of one or more non-depositing gases. In certain embodiments, the non-depositing gas comprises argon.

[0107] In certain embodiments, the surface of the substrate is activated by plasma treatment in the presence of one or more depositing gases. In certain embodiments, the depositing gas comprises oxygen or air. In certain embodiments, the surface of the substate is activated by depositing a gas that provides activable groups, for example plasma treatment with an organic alcohol which may be used to produce a high level of - OH groups for subsequent engraftment of the HPG.

[0108] In certain embodiments, the surface of the substrate is activated by plasma treatment with a gas and then the gas deposited polymer is subsequently activated by a further plasma treatment. For example, plasma treatment may be used to deposit a hydrocarbon, and the deposited hydrocarbon polymer subsequently activated by treatment with an oxygen or argon plasma treatment, followed by engraftment with HPG.

[0109] In certain embodiments, the plasma treatment comprises radio frequency induced plasma treatment. Other types of plasma treatment are contemplated.

[0110] In certain embodiments, the plasma treatment comprises treatment using a power in the range of 10 W to 400W.

[0111] In certain embodiments, the plasma treatment comprises treatment using a power in the range of 10W to 400W, 10W to 200W, 10W to 50W, 50W to 400W, 50W to 200 W, 50W to 100 W, 100W to 400 W, 100W to 400 W, or 100 to 200 W. As will be understood, to a person skilled in the art, the degree of activation of the surface of the substrate by plasma treatment will be on a spectrum. Therefore, it will be appreciated that plasma treatment even at low power (for example less than 10W) can be used to activate a surface. The appropriate power can be determined by a person skilled in the art based on the disclosure herein and routine trial and error. As such, other power ranges are contemplated. In some embodiments, the power is 400W or less, 200W or less, 50W or less, 20W or less, or 10W or less.

[0112] In certain embodiments, the time of plasma activation is 1 hour or less, 30 minutes or less, 20 minutes or less, 10 minutes or less, or 5 minutes or less. Other times are contemplated.

[0113] In certain embodiments, the hyperbranched polyglycerol coating comprises a thickness of 1 nm or more, 2 nm or more, 3 nm or more, 4 nm or more, 5 nm or more, 6 nm or more, 7 nm or more, 8 nm or more, 9 nm or more, 10 nm or more, 15 nm or more, 20 nm or more, 50 nm or more or 100 nm or more. Other thicknesses are contemplated. A suitable thickness relevant to the application may be selected. Methods for determining the thickness of a coating are known in the art.

[0114] In certain embodiments, the hyperbranched polyglycerol coating comprises a thickness of at least Inm, at least 2 nm, at least 3 nm, at least 4 nm, at least 5 nm, at least 6 nm, at least 7 nm, at least 8 nm, at least 9 nm, at least 10 nm, at least 20 nm, at least 50 nm or at least 100 nm.

[0115] In certain embodiments, the exposing of the activated surface to the vapour comprises a temperature in the range from 50°C to 140°C.

[0116] In certain embodiments, the exposing of the activated surface to the vapour comprises a temperature in the range from 50°C to 130°C, 50°C to 120°C, 50°C to 110°C, 50°C to 100°C, 50°C to 90°C, 50°C to 80°C, 50°C to 70°C, 50°C to 60°C, 60°C to 140°C, 60°C to 130°C, 60°C to 120°C, 60°C to 110°C, 60°C to 100°C, 60°C to 90°C, 60°C to 80°C, 60°C to 70°C, 70°C to 140°C, 70°C to 130°C, 70°C to 120°C, 70°C to 110°C, 70°C to 100°C, 70°C to 90°C, 70°C to 80°C, 80°C to 140°C, 80°C to 130°C, 80°C to 120°C, 80°C to 110°C, 80°C to 100°C, 80°C to 90°C, 90°C to 140°C, 90°C to 130°C, 90°C to 120°C, 90°C to 110°C, 90°C to 100°C, 100°C to 140°C, 100°C to 130°C, 100°C to 120°C, 100°C to 110°C, 110°C to 140°C, 110°C to 130°C, 110°C to 120°C, 120°C to 140°C, 120°C to 130°C, or 130°C to 140°C. Other ranges are contemplated.

[0117] In certain embodiments, the exposing of the activated surface to the vapour comprises a temperature of 50°C or greater, 60°C or greater, 70°C or greater, 80°C or greater, 90°C or greater, or 100°C or greater.

[0118] In certain embodiments, the exposing of the activated surface to the vapour comprises a time of 72 hours or less, 48 hours or less, 24 hours or less, 12 hours or less, 6 hours or less, 4 hours or less, 2 hours or less, or 1 hour or less.

[0119] In certain embodiments, the exposing of the activated surface to the vapour comprises a time in the range from 4 hours to 72 hours, 4 hours to 48 hours, 4 hours to 24 hours, 4 hours to 18 hours, 4 hours to 12 hours, 4 hours to 6 hours, 6 hours to 72 hours, 6 hours to 48 hours, 6 hours to 24 hours, 6 hours to 18 hours, 6 hours to 12 hours, 12 hours to 72 hours, 12 hours to 48 hours, 12 hours to 24 hours, or 12 hours to 18 hours, 24 hours to 72 hours, 24 hours to 48 hours, or 48 hours to 72 hours.

[0120] In certain embodiments, the coating is formed by a reaction comprising a single (non-iterative) reaction synthesis of monomers. In certain embodiments, the coating is formed by reactions comprising multiple (iterative) reaction syntheses of monomers.

[0121] In certain embodiments, the vapour does not comprise any further chemical initiators. In certain embodiments, the vapour comprises one or more further chemical initiators.

[0122] In certain embodiments, the method comprises providing energy to polymerise the monomers on the activated surface of the substrate. In certain embodiments, this energy is provided subsequent to completion of activation. In certain embodiments, this energy is not provided by plasma.

[0123] In certain embodiments, the energy for polymerising the monomers is heat energy, and the method comprises providing heat to polymerise the monomers. It is to be understood that providing heat means increasing the temperature above ambient temperature.

[0124] In certain embodiments, the heat is provided in a temperature range above 50°C. In a preferred embodiment the temperature range is from 50°C to 140°C.

[0125] In certain embodiments, the heat is provided in a temperature range from 50°C to 130°C, 50°C to 120°C, 50°C to 110°C, 50°C to 100°C, 50°C to 90°C, 50°C to 80°C, 50°C to 70°C, 50°C to 60°C, 60°C to 140°C, 60°C to 130°C, 60°C to 120°C, 60°C to 110°C, 60°C to 100°C, 60°C to 90°C, 60°C to 80°C, 60°C to 70°C, 70°C to 140°C, 70°C to 130°C, 70°C to 120°C, 70°C to 110°C, 70°C to 100°C, 70°C to 90°C, 70°C to 80°C, 80°C to 140°C, 80°C to 130°C, 80°C to 120°C, 80°C to 110°C, 80°C to 100°C, 80°C to 90°C, 90°C to 140°C, 90°C to 130°C, 90°C to 120°C, 90°C to 110°C, 90°C to 100°C, 100°C to 140°C, 100°C to 130°C, 100°C to 120°C, 100°C to 110°C, 110°C to 140°C, 110°C to 130°C, 110°C to 120°C, 120°C to 140°C, 120°C to 130°C, or 130°C to 140°C. Other ranges are contemplated.

[0126] In some embodiments, exposing the activated surface to the vapour is done simultaneously with polymerisation. In some such embodiments - where heat is provided to polymerise the monomers - the heat energy for polymerisation is the heat from the temperature used for exposing the activated surface to the vapour.

[0127] In certain embodiments, the monomers are produced in a reaction vessel where the polymerisation of the monomers on the activated surface occurs.

[0128] In certain embodiments, the monomers are produced from a source of liquid glycidol (and/or a derivative thereof) located in a reaction vessel.

[0129] In certain embodiments, the monomers are produced by heating a source of liquid glycidol and/or a derivative thereof in the reaction vessel.

[0130] In certain embodiments, the vapour is introduced from an external source into a reaction vessel where the polymerisation of the monomers on the activated surface occurs. [0131] In certain embodiments, the surface is activated, and the polymerisation of the monomers occurs in the same reaction vessel, as two separate steps in the process. In certain embodiments, the surface is activated by plasma activation and subsequently the polymerisation of the monomers occurs in the same reaction vessel.

[0132] In certain embodiments, the exposing of the activated surface to the vapour is a continuous exposure to the vapour over a period of time. In certain embodiments, the exposing of the activated surface to the vapour is a discontinuous exposure to the vapour over a period of time.

[0133] In certain embodiments, the hyperbranched polyglycerol coating produced has a characteristic of reduced fouling to biological materials, or a reduced fouling or thrombosis associated with use of the coating in a medical setting.

[0134] Biological materials include cells, cell debris, proteins, platelets, microbial matter, and organic matter. Other types of biological materials are contemplated.

[0135] In certain embodiments, the methods as described herein are used to reduce fouling of a substrate, to reduce attachment of proteins to a substrate, to reduce attachment of microbial matter, to reduce attachment of organic matter and/or cells to the substrate; to reduce fouling of a medical device, to reduce thrombosis, to reduce fouling of filtration membranes, to reduce fouling of tubing, or to reduce fouling of liquid handling equipment.

[0136] In certain embodiments, the reduction of one or more of the aforementioned characteristics comprises a reduction by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90%, as compared to an uncoated substrate.

[0137] In certain embodiments, the methods as described herein are used to increase hydrophilicity of the substrate. Methods for assessing hydrophilicity are described herein.

[0138] Examples of products utilising a coated substrate as described herein include medical devices such as a graft, a stent, a cannula, a catheter, a guide wire, a patch, a sheath, a suture, a valve, tubing or a part of a device in contact with a biological fluid. Other types of medical devices are contemplated.

[0139] In certain embodiments, the product is a device such as a water handling device, a membrane, or a filter.

[0140] Methods for manufacturing products from, or incorporating, a substrate as known in the art and as described herein. It will be appreciated that the coated substrate may be first coated by a method as described herein and then incorporated into the product, and/or the substrate may be coated in situ in the product.

[0141] In certain embodiments, the method includes the step of activating the surface of the substrate or providing a substrate with an activated surface. Accordingly, in such embodiments the method of producing a hyperbranched polyglycerol coating on a substrate, comprises: activating a surface of the substrate, or providing a substrate with an activated surface; then exposing the activated surface to a vapour comprising monomers of glycidol and/or a derivative thereof; and polymerising the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the substrate.

[0142] In certain embodiments, the present disclosure provides a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising reacting a vapour comprising monomers of glycidol and/or a derivative thereof with an activated surface of the substrate and producing the hyperbranched polyglycerol coating by polymerisation of the monomers on the activated surface.

[0143] In certain embodiments, the present disclosure provides a method of producing a hyperbranched polyglycerol coating on a substrate, the method comprising polymerising monomers of glycidol and/a derivative thereof in a vapour on an activated surface of the substrate and thereby coating the substrate with the hyperbranched polyglycerol.

[0144] In certain embodiments, the methods as described herein are used to reduce fouling of a substrate, to reduce thrombosis associated with the use of a medical device; to produce an anti-fouling substrate; to produce an anti -thrombotic substrate; to reduce attachment of proteins, microbial matter, organic matter and/or cells to a substrate; to reduce fouling of a medical device, to reduce fouling of a filtration membrane; to reduce fouling of tubing; to reduce fouling of liquid handling equipment; and/or to increase hydrophilicity of a substrate.

[0145] Certain embodiments of the present disclosure provide a substrate coated with a hyperbranched polyglycerol produced by a method as described herein.

[0146] In certain embodiments, the substrate is used in a medical setting. Examples of products or devices which are used in a medical setting include medical devices such as a graft, a stent, a cannula, a catheter, a guide wire, a patch, a sheath, a suture, a valve, tubing or a part of device in contact with a biological fluid. Other types of medical devices are contemplated.

[0147] In certain embodiments, the substrate is used in a non-medical setting. In certain embodiments, the substrate is used in a water or liquid handling setting. Examples of such products or devices include filters, pumps, valves, or pipes.

[0148] In certain embodiments, the coated substrate is an anti-fouling substrate. In certain embodiments, the coated substrate is an anti-fouling and/or anti -thrombotic substrate.

[0149] Certain embodiments of the present disclosure provide a substrate coated with a hyperbranched polyglycerol, the coating produced by exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or derivative thereof and polymerisation of the monomers on the activated surface.

[0150] Certain embodiments of the present disclosure provide a product comprising a coated substrate as described herein.

[0151] In certain embodiments, the product is used in a medical setting. Examples of products or devices include medical devices such as a graft, a stent, a cannula, a catheter, a guide wire, a patch, a sheath, a suture, a valve, tubing or a part of device in contact with a biological fluid. Other types of medical devices are contemplated. [0152] In certain embodiments, the product is used in a non-medical setting. In certain embodiments, the substrate is used in a water or liquid handling setting. Examples of such products or devices include filters, pumps, valves, or pipes.

[0153] Methods for producing products or devices incorporating substrates are known in the art.

[0154] Certain embodiments of the present disclosure provide a method of reducing fouling of a substrate.

[0155] In certain embodiments, the present disclosure provides a method of reducing fouling of a substrate, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

[0156] In certain embodiments, the fouling comprises fouling with one or more cells, cell debris, proteins, platelets, microbial matter, and organic matter.

[0157] Certain embodiments of the present disclosure provide a substrate with reduced fouling produced by a method as described herein.

[0158] In certain embodiments, the present disclosure provides a method of reducing fouling and/or thrombosis of a substrate for use in a medical setting, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof and forming a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers.

[0159] In certain embodiments, the present disclosure provides a method of producing a substrate with reduced fouling and/or thrombosis for use in a medical setting, the method comprising exposing an activated surface of the substrate to a vapour comprising monomers of glycidol and/or a derivative thereof to form a hyperbranched polyglycerol coating on the activated surface of the substrate by polymerisation of the monomers. [0160] Certain embodiments of the present disclosure provide a substrate with reduced fouling and/or thrombosis produced by a method as described herein.

[0161] Certain embodiments of the present disclosure provide a method of producing a product with reduced fouling.

[0162] In certain embodiments, one or more materials in the product are coated as described herein prior to production of the product.

[0163] In certain embodiments, one or more materials in the product are coated as described herein after production of the product.

[0164] Certain embodiments of the present disclosure provide a product with reduced fouling produced by a method as described herein.

[0165] In certain embodiments, the product is used in a medical setting.

[0166] Examples of products or devices which are used in a medical include medical devices such as a graft, a stent, a cannula, a catheter, a guide wire, a patch, a sheath, a suture, a valve, tubing or a part of device in contact with a biological fluid. Other types of medical devices are contemplated.

[0167] In certain embodiments, the product is used in a non-medical setting.

[0168] In certain embodiments, the substrate is used in a water or liquid handling setting. Examples of such products or devices include filters, pumps, valves, or pipes.

[0169] Certain embodiments of the present disclosure provide a medical product with reduced fouling and/or thrombosis produced by a method as described herein.

[0170] In certain embodiments, one or more materials in the medical device are coated prior to production of the medical device. For example, a medical device may be produced from metal or polymeric materials that have been pre-coated with a hyperbranched polyglycerol. [0171] In certain embodiments, one or more materials in the medical device are coated after production of the medical device. For example, a medical device may be produced and the metal or polymeric materials in the device subsequently coated with a hyperbranched polyglycerol.

[0172] In certain embodiments, a medical device as described herein comprises one or more characteristics in use selected from reduced attachment of platelets to the coated polymeric material, reduced attachment of cells (such as inflammatory cells) and/or proteins to the coated polymeric material, reduced fouling, reduced clotting, reduced thrombosis, reduced restenosis and reduced anastomotic hyperplasia.

[0173] In certain embodiments, the reduction of one or more of the aforementioned characteristics comprises a reduction by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90%, as compared to uncoated polymeric material.

[0174] Certain embodiments of the present disclosure provide a medical device with one or more characteristics of reduced platelet attachment, reduced cell attachment, reduced fouling, reduced clotting, reduced thrombosis and reduced anastomotic hyperplasia produced by coating the device by a method as described herein.

[0175] Methods for assessing platelet and cell attachment to materials are known in the art. For example, cells may be stained with specific cell stains/markers and these used to identify cells associated with a material. Methods for assessing fouling are known in the art, and include for example, visualisation of the material for attached matter (e.g., proteins, cells, platelets) by light microscopy. Methods for assessing anastomotic hyperplasia are known in the art, and include for example, histologic assessment of implanted materials or assessment of hyperplasia in animal models using flow analysis. Methods for assessing clotting or thrombosis are known in the art, and include for example, assessment of implanted materials for the presence of a clot/thrombus and/or in vitro studies as described herein.

[0176] Certain embodiments of the present disclosure provide a system for coating a substrate with a hyperbranched poly glycerol. [0177] In certain embodiments, the present disclosure provides a system for coating a substrate with a hyperbranched polyglycerol, the system comprising a plasma activation system comprising a reaction vessel and a means to vaporise a source of a liquid in the reaction vessel.

[0178] Methods and equipment for coating a substrate are as described herein.

[0179] Certain embodiments of the present disclosure provide a method of coating a substrate with a hyperbranched polyglycerol with a system as described herein.

[0180] In certain embodiments, the present disclosure provides a method of coating a substrate with a hyperbranched polyglycerol, the method comprising using a system as described herein to activate the substrate and coat the activated substrate with the hyperbranched polyglycerol by polymerisation of gaseous monomers of glycidol and/or a derivative thereof from the source of liquid glycidol.

[0181] Preferably, for the methods provided herein, when plasma activation of the substrate is performed, the activation is completed before polymerisation of the gaseous monomers.

[0182] Certain embodiments of the present disclosure provide use of product comprising a coated substrate as described herein.

[0183] In certain embodiments, the product is a medical device.

[0184] In certain embodiments, the present disclosure provides use of a medical device as described herein to prevent and/or treat a condition, such as a vascular condition, arterial or venous narrowing, ischemia, angina, an aneurysm, or to repair or support an artery or vein. Other diseases, conditions or states are contemplated. Methods for treating conditions using medical devices are known in the art.

[0185] In certain embodiments, the present disclosure provides a method of treating a condition in a subject that would benefit from the introduction of a medical device as described herein. In certain embodiments, the condition is a vascular condition. In certain embodiments, the vascular condition comprises arterial or venous narrowing, angina, an aneurysm, or repair or support of an artery or vein.

[0186] In certain embodiments, the substrate is a membrane. Accordingly, in certain embodiments the present disclosure provides a membrane comprising a hyperbranched polyglycerol coating and methods for producing such a membrane.

[0187] In some embodiments, the membrane is a filter membrane, such as a water filter membrane or other liquid filter membrane.

[0188] In certain embodiments, the present disclosure provides a method of producing a hyperbranched polyglycerol coating on a membrane, such as a filter membrane, the method comprising: exposing an activated surface of the membrane to a vapour comprising monomers of glycidol and/or a derivative thereof, and polymerising the monomers on the activated surface to produce the hyperbranched polyglycerol coating on the membrane.

[0189] The present disclosure also provides a method of improving the cleanability of a substrate by coating the substrate with a poly glycerol coating using the methods described herein. Also provided, in certain embodiments, are substrates having improved cleanability. In certain embodiments, the substrate having improved cleanability is a membrane, such as a filter membrane.

[0190] The present disclosure is further described by the following examples. It is to be understood that the following description is for the purpose of describing particular embodiments only and is not intended to be limiting with respect to the above description.

[0191] EXAMPLE 1- Coating of PTFE Cannula

[0192] This example presents the results from the optimization of a coating process for PTFE cannulas. The results show the successful application of a HPG coating on the outer surface of the cannula. The thickness of the coating produced under the optimized conditions was close to the analysis depth of the XPS (10-15nm). In addition, the coated cannulas displayed highly hydrophilic properties compared with the hydrophobic, untreated PTFE cannula. The optimisation of the coating process is presented, along with the supporting analytical data. [0193] Abbreviations

HPG: Hyperbranched polyglycerol

PTFE: Polytetrafluoroethylene

XPS: X-ray photoelectron spectroscopy

MFC: Mass flow controller

RF: Radio frequency

WCAs: Water contact angles

[0194] Background

[0195] PTFE (polytetrafluoroethylene) is commonly used in many medical device applications due to its chemical resistance and low coefficient of friction. Examples of medical products containing PTFE include cannulas, catheters, tubing, filters, surgical grafts, and endovascular grafts. Cannulate are used extensively with subcutaneous drug infusion pumps used in the treatment and management of diabetes, primary immunodeficiencies and pain management. The small soft PTFE cannula is subcutaneously implanted and left indwelling for a period of several days.

[0196] The aim of this study was to show that a HPG coating would render the infusion cannula more hydrophilic and more biocompatible, with an extended duration of greater than 7 days when subcutaneously implanted.

[0197] Reagents

[0198] Glycidol (96%) was purchased from Sigma- Aldrich and used without further purification. PTFE cannulas purchased from Solmed (Aust), and PTFE sheets purchased from Goodfellow (UK) were used. Absolute Ethanol (ChemSupply) and RO water were used as solvents.

[0199] Plasma activation process

[0200] The plasma activation step was performed using a plasma reactor with an aluminium chamber (Dimensions of the chamber 250mm x 240mm x 605mm) (Nano, Diner electronic GmbH, Germany). The reactor was equipped with a power generator delivering a power range 0-500W at 100kHz. PTFE cannulas and small pieces (approximately 1 ,5cm x 1cm) of PTFE sheets were mounted on the ground electrode. The chamber was pumped down to reach a base pressure below 1 x 10' 3 mbar. The processing gas was then introduced at a specific flow rate controlled by a mass flow controller (MFC). Finally, the plasma was ignited in the chamber at a certain power and continued for a certain time. Once the plasma activation run was completed, the samples were transferred into a box for the subsequent HPG grafting step.

[0201 ] HPG grafting process

[0202] HPG grafting process was performed by placing the plasma activated samples into a sealed container. Into the same container was added glycidol (4 mL glycidol per litre of container volume) at the bottom of the container. The samples to be coated were not in contact with the glycidol. The sealed container was then placed in an oven at 100°C for 24h. At temperatures above 50°C glycidol will evaporate into a vapour, so the samples in this study were exposed to a vapour of glycidol for the 24 hrs. The samples were then taken out and washed thoroughly with RO water and then ethanol for 15min each. Finally, the samples were left to dry in a laminar flow cabinet before stored in a clean container.

[0203] Analytical methods

[0204] X-Ray Photoelectron Spectroscopy (XPS)

[0205] XPS analyses were performed on a Kratos Axis Ultra DLD X-ray photoelectron spectrometer equipped with a monochromatic AlKa radiation source (hv = 1486.6 eV) operating at 225 W (15 kV x 15 mA). Spectra on PTFE flat samples were acquired using an analysis area of 300 x 700 pm, and for the PTFE cannulas an aperture size of 110 pm was used. High resolution spectra were corrected for charging effects during analysis using a reference value of 285 eV, the binding energy of the C-C component from neutral hydrocarbons.

[0206] Atomic percentage values were quantified from survey spectra using CasaXPS software (www.casaxps.com). CasaXPS software was also used to fit component peaks under the C Is high-resolution spectra. The line shape of the curves was assumed to be Gaussian-Lorentzian (G/L) with a 30% Lorentzian component. The peak width and binding energy were constrained for all components to obtain the best fit possible.

[0207] Water contact angle measurements

[0208] WCA is commonly used as a method of characterising the wettability of surfaces. This technique is highly sensitive to changes in surface chemistry. A 3 pL droplet of Milli-Q water was carefully dispensed on the surface of the flat HPG/PTFE samples. After 5 min, the contact angle was measured over a time period of 20 s (one measurement was taken every 2 s with a total of 10 measurements taken). The average and standard deviation values were then computed.

[0209] For the HPG/PTFE cannulas, samples were dipped in water and an image of the meniscus was captured. Four images were taken along each cannula (from the tip to the base) and a 3x magnification was used. The contact angles were calculated by processing the images using a plugin drop analysis: LB-ADSA on ImageJ.

[0210] Results and Discussion

[0211] Plasma process gas used for the plasma activation step

[0212] Plasma process gas was investigated as a process variable using a range of mixing ratios of gas A (Oxygen) and gas B (Argon), while maintaining the gas flowrate at 4 standard cubic centimetres per minute (seem) throughout. The results show that increasing the percentage of gas B gradually in the mix from 0% to 100% resulted in a gradual increase in the proportion of oxygen and carbon in the XPS spectra; associated with the presence of the HPG coating being formed on the surface (Figure 1). At the same time, a gradual decrease in the concentration of fluorine (associated with the substrate) is observed. Only carbon and fluorine were observed on the surface of untreated PTFE cannulas and PTFE sheets.

[0213] A higher proportion of oxygen in the XPS spectra is indicative of a thicker HPG coating grafted on the surface. This means that using 100% gas B is more effective at activating the surface compared with gas A or a mix of both, when trying to engraft an HPG coating on PTFE. Differences in the proportion of oxygen measured on the surface of HPG/PTFE sheets compared with the HPG/PTFE cannulas under all conditions, is likely due to the difference in properties of the starting materials; cannulas and sheets were manufactured differently. This is likely to be related to differences in the ratio of carbon and fluorine on the respective untreated substrates, which can account for differences in the XPS results observed.

[0214] Curve-fitting analysis of the high resolution XPS C is spectra was performed to measure the relative concentrations of various functional groups present on the surface. A summary of the curve-fitting results is presented in Figure 2.

[0215] The functional groups present on the surface include C-C/C-H, C-O, C=O, CF, CF2 and CF3. The focus is on the percentage of C-0 and CF2, which are the major functional groups present in the HPG polymer coating and PFTE substrate, respectively. The percentage of C-0 increases gradually with the increase in the proportion of gas B in the mix, to reach a maximum value for C-0 of 38.2% at 100% of gas B. At the same time, there is a corresponding decrease in the percentage of CF2 detected. At 100% gas B, the percentage of CF2 decreased from 93.8% to 44.7% and from 88.4% to 49.4% for the PTFE cannulas and PTFE sheets, respectively.

[0216] The wettability was determined by measuring the water contact angles (WCAs) on the surface of samples after the HPG engraftment. A summary of WCA results is presented in Figure 3. The WCA measured on the surface of untreated PTFE sheet was approximately 127°, consistent with a very hydrophobic polymer. A drop in WCA of approximately 20° at 0% of gas B and approximately 30° at 100% of gas B, in the plasma process gas mix, was observed for the PTFE sheet.

[0217] To measure the WCA on untreated and HPG treated PTFE cannulas, images were captured of the meniscus formed when a sample is dipped in water and analysed. A summary of the WCAs results for cannulas is given in Figure 4. The WCA of the untreated PTFE cannula is comparable to that measured for the PTFE sheet. The same reduction in WCA is observed on HPG coated cannulas, except the WCA values are lower than those measured on the PTFE sheet. The lower WCAs indicates higher oxygen (in the form of C-O) as measured by XPS on the modified cannulas when compared with the PTFE sheets after HPG grafting. [0218] Plasma process gas flow rate

[0219] To investigate the impact of the flow rate of the plasma process gas, PTFE cannulas and PTFE sheets were plasma activated at a higher (4 seem) and lower (2 seem) flow rate of gas B. XPS results presented in Figure 5 show that the low gas B flow rate gave higher oxygen levels on the surface, as measured by XPS, compared with the higher flow rate. This would indicate that using a low flow rate of gas B is more effective at activating the surface of both PTFE materials for HPG engraftment, compared with the high flow rate. Similarly, curve-fitting analysis of the high-resolution C is spectra show higher percentages of the C-0 functionality on the surface at the lower flow rate of plasma process gas compared to high flow rate (Figure 6), which is indicative of a thicker HPG coating grafted on the surfaces.

[0220] Applied power

[0221] Using gas B at a flow rate of 2 seem, and keeping the remaining parameters constant, the effect of applied power (50, 100, 200 and 400W) for plasma activation was investigated. The XPS results collected on the activated samples after HPG grafting is presented in Figure 7. On both PTFE cannulas and PTFE sheets a gradual increase in the oxygen and carbon concentrations by XPS we observe, as power was increased, accompanied by a gradual decrease in the concentration of fluorine. This indicates that higher activation powers lead to a thicker engrafted HPG layer.

[0222] The small concentration of fluorine measured on the surface at the highest applied power indicates that the thickness of the grafted HPG layer is close to the XPS analysis depth (approximately 10-15nm). The results from curve-fitting analysis in Figure 8 show similar observations, with the percentage of C-0 functionality measuring approximately 90% at the highest applied RF power.

[0223] The wettability of the HPG coatings at various applied powers is presented in Figure 9. The WCA values dropped as the applied power was increased, indicating that the thicker the HPG coating the more hydrophilic the surface. The WCA values at 200W and 400W were comparable, which may suggest that a maximum effective coating was achieved at around 200W. [0224] The results from the wettability and XPS studies show a good correlation between the WCAs and the surface chemistry of the corresponding coatings.

[0225] Plasma activation time

[0226] The final parameter investigated was the plasma activation time, which was assessed using the highest applied powers (i.e. 200W and 400W), as their XPS results were comparable. While keeping all other plasma activation parameters constant, PTFE cannulas and PTFE sheets were activated with gas B plasma for 1, 5, 10 and 20 min.

[0227] The XPS results for the HPG coatings show small differences between the measured elemental concentrations over the tested plasma activation times (Figure 11). In addition, there were no significant differences between the elemental concentrations of HPG coatings produced at 200W and 400W. This would suggest shorter activation times may be used to generate suitable HPG coatings on the surface of PTFE cannulas. On the other hand, a gradual increase in the oxygen concentration on PTFE sheets was observed, which reached a saturation level after lOmin of plasma activation. For the 200W and 400W applied power, the oxygen concentrations are comparable on the coated PTFE cannulas and PTFE sheets for the plasma activation times of lOmin and 20min. Similar observations are obtained after curve fitting analysis of the high-resolution C is spectra, as shown in Figure 12.

[0228] This confirms that a maximum effective coating is being generated at 200W and at lOmin activation time, under the specific conditions used in this particular reactor. Other reactor configurations may require different process parameters to achieve the same HPG coating. Interestingly, when cannulas were coated using 400W and 20 min activation, the level of oxygen fell and fluorine increased, compared with 10 min. This may suggest that these extreme activation conditions tested (400W and 20 min), in this reactor, may be having a detrimental effect on the quality of the HPG coating being generated.

[0229] Conclusions [0230] The experiments were successful in confirming an HPG coating can be engrafted on PTFE, when exposed to a vapour of glycidol during the engraftment step. XPS and WCA analyses were used throughout to confirm the presence of an HPG coating on PTFE. Several key parameters were investigated while optimizing the HPG coating process; including plasma gas, plasma gas flow rate, applied power and plasma activation time. The results show that a broad range of activation parameters can be used to generate a suitable surface from which to engraft an HPG coatings. The observed concentration of fluorine of around 5% on coated surfaces formed using the best conditions, suggests that the thickness of HPG coating we were achieving was in the range of 10-15nm.

EXAMPLE 2 - HPG Coating on woven PET Grafts

[0231] The aim of this study was to optimise conditions for the grafting of hyperbranched polyglycerol (HPG) from plasma activated polyethylene terephthalate (PET) woven materials. Some grafting conditions suitable for metallic substrates may result in shrinkage of woven PET graft material. Woven PET is used extensively in medical and surgical procedures and can be found in stent grafts and endovascular grafts. So shrinkage of this material during the HPG coating process would not be desirable.

[0232] This example summarises experiments aiming to minimise PET graft shrinkage, while still engrafting a functional HPG polymer. A number of variables were investigated in the process conditions including plasma gas (argon or oxygen), polymerisation time, polymerisation temperature and monomer volume to polymerisation vessel volume ratio.

[0233] Abbreviations

HPG - Hyperbranched polyglycerol

PET - Polyethylene terephthalate

PBS - Phosphate buffered saline

XPS - X-ray photoelectron spectroscopy

SEM - Scanning electron microscopy

PFA - Paraformaldehyde

HDMS - Hexamethyldisilazane

DAPI - 4’,6-diamidoino-2-phenylindole

CFSE - Carboxyfluorescein succinimidyl ester [0234] Background

[0235] Exposure of woven PET grafts to glycidol under the conditions used for PTFE, above, has been observed to cause damage and shrinkage of the PET. Furthermore, immersion of PET in liquid glycidol to engraft HPG to the surface, causes the PET to become brittle and lose its mechanical properties. The aim in this example was to optimise the coating conditions to overcome the PET shrinkage.

[0236] Previous studies to explore HPG engraftment conditions on metallic substrates like stents, indicated that shorter polymerisation times may yield a robust anti- thrombogenic HPG coating. Polymerisation times as short as 1 hour have historically yielded HPG coatings on stainless steel and silicon wafer that had been chemically activated and polymerised at 100°C. Other studies performed on silicon wafers coated with a cyclohexane plasma polymer as an interlayer, suggested that HPG polymerisation would not occur below 70°C (Anouck et al. 2020 ACS Appl. Bio. Mater. 3: 3718-3730). However, these samples were surface activated under very different conditions of power, pressure and gas.

[0237] In this example, we attempted to coat PET grafts by exposing the material to a vapour of glycidol monomer, while being heated to elevated temperatures.

[0238] It was also desirable to explore polymerisation temperatures below the glass transition temperature of PET (69°C). Therefore, an engraftment temperature of 65°C was tested.

[0239] The example had two main objectives:

1. Demonstrate that an HPG coating could be engrafted by exposure of the PET to a vapour of glycidol.

2. Optimise the process conditions for the grafting of HPG from woven PET fabric to reduce or eliminate shrinkage of PET grafts.

[0240] Reagents [0241] Glycidol: 96% (Cat#G5809) Sigma Aldrich (Saint Louis, Missouri, USA), Argon and oxygen: Industrial Grade (Gas Code: 061) BOC Aust, PET graft material (ATEX Technologies, Inc.)

[0242] Plasma Activation of Substrates

[0243] PET grafts were activated with either an argon or oxygen plasma for 20 mins at 100 W and 0.06 mbar in a Diener Femto pilot reactor. Following plasma treatment with either gas, the vacuum chamber was backfilled with argon gas.

[0244] The only variable explored in the plasma activation step in the process was the gas used (argon or oxygen). All other conditions were unchanged - 20 mins plasma activation at 100 W and 0.06 mbar of gas pressure.

[0245] HPG Engraftment

[0246] Activated samples were taken from the plasma chamber and placed directly into containers of varying size containing a range of liquid glycidol volumes, to investigate monomer volume to chamber volume ratios.

[0247] Containers investigated included: a glass desiccator (1.4 L), a stainless-steel sealed container (2.5 L), capped glass vials (25 mL), 10 mL capped plastic tubes, 15 mL and 50 mL capped Falcon tubes and 250 mL plastic storage box. For the desiccator, stainless steel container and 250 mL storage box, glycidol was added to a separate plastic dish that was placed inside the chamber. For the tubes, glycidol was placed at the bottom of the tubes and the samples to be coated were placed in the tubes, avoiding direct contact with the glycidol liquid.

[0248] Once secured inside their respective containers, they were placed in an 80°C oven containing a sand bath to support the various containers. The 250 mL plastic boxes were placed in an oven at 65°C and atmospheric pressure.

[0249] Samples were incubated at 80°C for 1, 2, 3, 4, 5, 18 and 24 hr, with additional samples tested at 4 and 24 hr. A time course was not carried out for the 65°C engraftment samples, which were held for 24 hr. Following the engraftment step, containers were removed from the ovens and allowed to cool to room temperature before being opened. Small samples were removed from the containers with forceps and placed into 6- or 12- well plates for washing with 80% (w/v) ethanol. Larger samples were placed in clean Falcon tubes for washing. Samples were washed three times with sufficient ethanol to cover the sample. For each wash samples were lightly agitated for 30 seconds before aspirating the ethanol. Samples were then dried under a stream of nitrogen gas and stored in a vacuum desiccator.

[0250] Static Blood Assay

[0251] Approximately 1 cm 2 samples of HPG-coated and uncoated woven PET fabric were placed into separate wells of a 24-well plate and 10 pL of freshly collected human blood, with no anti-coagulant, was placed on each sample to be tested. The lid was placed on the well plate and samples left undisturbed at room temperature for 20 min. Room temperature PBS (~1 mL per well) was added and the well plate agitated in a swirling motion to wash the samples. The PBS was aspirated after 30 s and more PBS added. This was repeated until the PBS stopped turning red for all samples, usually achieved after 4- 5 washes. One mL of PF A (4% in PBS) was added to each sample and left at room temperature for 20 min. Samples were then washed once with 1 mL PBS and either prepared for scanning electron microscopy (SEM) or fluorescently stained with CFSE and DAPI.

[0252] Sample Preparation for SEM

[0253] Samples not exposed to blood were mounted on SEM stubs and sputter coated with 10 nm of platinum. Samples from the static blood assay were first dehydrated through a series of ethanol washes; 80% for 30 min, 90% for 30 min then 100% for 1 hr. Samples were then placed in a 50/50 mixture of ethanol and hexamethyldisilizane (HDMS) for 20 min then 100% HDMS for 20 min. Samples were removed from the 100% HDMS and left to air dry. Dry samples were then attached to SEM stubs and sputter coated with 10 nm of platinum.

[0254] Chemical Analysis [0255] XPS was performed with a Kratos AXIS Ultra DLD spectrometer, using monochromatic AlKa radiation source (hv = 1486.7 eV). Spectra were recorded using the X-rays produced upon bombardment of the Al anode surface with thermal electrons of power 225W (15 mA flux of electrons accelerated by voltage of 15 keV). The analysed area was rectangular (0.3 x 0.7 mm). Survey spectra (-10-1110 eV binding energy range) were collected at pass energy (resolution) of 160 eV with a step size of 0.5 eV. To determine the chemical functionalities, high resolution spectra of the Cis core level peak were collected at pass energy (resolution) of 20 eV with a step size of 0.1 eV. Data analysis was performed with CasaXPS software (Casa Software Ltd.). All binding energies were referenced to the low energy, aliphatic Cis peak at 285.0 eV.

[0256] Visual Analysis of Surface Changes and Static Blood Assay

[0257] Zeiss Merlin FEG SEM was used to capture high resolution images of the surface of pre- and post-HPG-coated PET. Images were also captured of the blood components bound following the static blood assay.

[0258] Zeiss LSM 710 with 10X objective was used to capture fluorescence images.

[0259] Results

[0260] HPG-Coating Conditions for Grafts

[0261] A full-length graft (102 mm) was surface activated using argon plasma conditions of 20 min at 100 W and 0.06 mbar then placed into a glass desiccator (1.4 L) with 4 mL of glycidol placed in a glass dish under the stage supporting the graft, to allow the glycidol to vaporise. The desiccator was incubated at 80°C for 24 hr for the engraftment step.

[0262] Under these conditions of engraftment from the glycidol vapour, the graft showed a slight reduction in length from 102 mm to 95 mm (7%). There were no visible signs of damage or degradation to the material.

[0263] Grafting Time Optimisation [0264] Time is a critical variable in the growth of the hyperbranched polyglycerol polymer on an activated surface and in the case of woven PET, length of exposure to glycidol vapour appears to be a key factor in determining the degree of shrinkage. Specifically, the combination of duration of exposure to glycidol and elevated temperatures. To minimise exposure to glycidol monomer vapour, short engraftment times of one to four hours were assessed to determine if a suitable HPG coating could be generated with minimal or no measurable shrinkage of the PET.

[0265] For these studies, 6 mm samples of woven PET were surface activated with standard argon plasma conditions (see above under heading “Plasma Activation of Substrate”), then individually placed in 25 mL glass vials containing 100 pL glycidol and held at an angle to avoid direct contact of PET with liquid glycidol. The vials were capped and placed at 80°C to allow engraftment from the glycidol vapour.

[0266] XPS

[0267] HPG engraftment on the woven PET was detected by the high resolution Cis spectrum after just 1 hour, as an increase in the 286.6 eV peak (light blue peak in Figure 13) above the baseline levels of untreated PET. The peak at 286.6 eV was observed to increase as engraftment time increased. Inconsistencies were observed with the 24 hrs samples, where the 286.6 eV peak appeared lower at this time point than in samples after 4 hr engraftment. Polymeric materials can undergo polymer chain reorganisation at temperatures above their glass transition temperature. This observation at 24 hr may be a result of PET undergoing polymer chain reorganisation since the PET samples were exposed to temperatures above the glass transition temperature of PET. The data is shown in Figure 13, and the XPS summary data in Table 1. The engrafted HPG is detectable by XPS after just one hour of polymerisation activated PET at 80°C, as evidenced by the increase in the C-0 peak.

Table 1

[0268] Plasma Gas Optimisation

[0269] Oxygen plasma was tested as an alternative to argon, in an attempt to achieve a higher density of active sites on the PET material. This may reduce the time required to form a suitably dense HPG polymer and possibly aid in the growth of HPG at lower temperatures, such as at 65°C.

[0270] After activation samples were coated over a range of engraftment times at 80°C, to allow the comparison of argon with oxygen gas during the plasma activation step. All other plasma conditions were kept the same: Time - 20 minutes, Power - 100 W, Pressure - 0.06 mbar. Both plasma gasses were also compared at 65 and 80°C for 24 hr.

[0271] XPS

[0272] At each time point, the oxygen plasma appeared to generate a stronger XPS signal at 286.6 eV, suggesting the HPG polymer grew faster from PET when activated with the oxygen plasma than with the argon plasma. HPG was also seen on samples activated with the oxygen plasma with engraftment at 65°C.

[0273] The XPS date is shown in Figure 14. A summary of the XPS data is shown Table 2. The coated sample data in this table can be compared with the corresponding functional groups on uncoated PET listed in Table 1.

Table 2 Summary of XPS Data

[0274] SEM

[0275] Due to the nanometre scale thickness of the HPG-coating, observation of the polymer layer is not possible by SEM. However, changes in PET fibre topography following plasma treatment and/or HPG engraftment can be observed (Figure 15).

[0276] Argon plasma does not appear to cause any observable changes to the topography of the PET fibres by SEM (Figure 15). However, the oxygen plasma treatment has an obvious effect on the surface topography, causing a rippling effect on the fibres at right angles to the direction of extrusion (Figure 15). This rippling creates a greater surface area and therefore, may create more active sites for the subsequent engraftment of HPG following plasma activation. This may explain the increase in the 286.6 eV peak by XPS, compared with argon plasma activation.

[0277] As engraftment time is extended from 4 to 24 hours after argon plasma treatment of PET fibres, a slight increase in surface roughness is observed, likely to be due to the HPG-coating on the surface of the PET fibres (Figure 16A). Cracks along the length of the fibres are also noticeable as early as 2 hours and become more pronounced at 4 hours. [0278] Due to the rippling caused by the oxygen plasma alone, visualisation of the more subtle surface roughness observed on the HPG-coating at 24 hrs for argon-activated samples, is less noticeable for oxygen-activated samples (Figure 16B).

[0279] Outcome: Oxygen plasma leads to a thicker engrafted HPG coating on PET than with argon plasma activation, under the same engraftment conditions. Oxygen plasma can help produce HPG coatings at lower engraftment times and lower temperatures than argon plasma. Reducing either or both parameters have the potential to eliminate PET shrinkage or any other changes to the mechanical properties of PET on grafts.

[0280] Glycidol volume for the formation of glycidol vapour

[0281 ] In this glycidol vapour engraftment process, a volume of glycidol is added to the container in which the substrates are to be coated, avoiding direct contact of the substrate samples with the liquid. The sealed container is then placed in an oven at a specified temperature. The elevated temperatures in the oven cause the glycidol in the sealed containers to vaporise creating an atmosphere of glycidol. It was of interest to determine if varying the volume of glycidol in the container, thereby varying the vapour pressure of glycidol, would alter the rate of HPG polymer engraftment on the woven PET fabric.

[0282] In this experiment, 25 mL glass vials were used with glycidol added at 10, 25, 50 and 100 pL. All PET samples were argon-plasma activated and samples were engrafted at 80°C for 24 hours.

[0283] XPS

[0284] XPS analysis (Figure 17) confirmed that the 10 pL sample formed a considerable amount of HPG after 24 hr, evidenced by a peak at 286.6 eV in the high resolution Cis spectra. A higher peak but very similar peak profile was obtained for 25 pL. However, with 50 pL of glycidol added, the Cis peak showed a significantly lower concentration of etheric carbon at 286.6 eV, while the 100 pL sample showed a high concentration of etheric carbon peak. Notwithstanding the anomalous result from the 50 pL sample, the trend is a slight decrease in the proportion of the C-0 functional group from HPG, with increasing monomer volume added. [0285] A summary of the XPS data us shown in Table 3.

[0286] Table 3

Summary of XPS Data

[0287] Static blood assay for fluorescent microscopy

[0288] A static blood assay was combined with fluorescence staining to check for signs of thrombosis on the HPG coated samples. For this experiment, argon and oxygen plasma were compared at 2 and 4 hr at 80°C. Additionally, oxygen plasma was investigated at 65°C on the top and bottom shelf of the oven.

[0289] All samples exhibited background fluorescence from the PET and all blood components are stained with the fluorescent dye. The key features examined in this experiment are the fibrin strands spreading across the surface, represented as bright green elongated structures. The presence and quantity of these structures was used to distinguish between the different engraftment conditions tested.

[0290] The results are shown in Figure 18. Figure 18A shows static blood assay on control PET. Left image - edge of the blood drop and right image - centre of the blood drop. Bright green is fibrin and cells stained with CFSE. Static blood assay on 2hr HPG initiated with argon plasma (left) and oxygen plasma (right). Figure 18B shows static blood assay on 2hr HPG initiated with argon plasma (left) and oxygen plasma (right). Figure 18C shows Static blood assay on 4hr HPG, initiated with argon plasma (left) and oxygen plasma (right). Figure 18D shows Static blood assay on 24hr HPG at 65°C; initiated with oxygen plasma on the bottom of shelf (left and top shelf (right).

[0291 ] This experiment showed that 4 hours at 80°C for HPG engraftment, initiated with oxygen plasma, showed the best anti -thrombotic result. An engraftment time of 24 hr at a reduced temperature of 65°C also showed good anti -thrombotic performance. [0292] Outcomes: The HPG coating formed after 2 hours at 80°C was less efficient in reducing thrombosis compared with the 4 hr polymerisation samples. It appears that 4 hours at 80°C is suitable to produce an anti-thrombotic HPG coating, if the surface is activated with an oxygen plasma. The argon activated, 4-hour engraftment samples showed some signs of thrombosis, but was still significantly better than the 2 hour samples.

[0293] Static blood assay and SEM analysis

[0294] Fresh blood was collected from a healthy donor and quickly placed on strips of the samples to be studied, suspended over 4-well tissue culture plates for glass slides.

[0295] The SEM data is shown in Figure 19. Panel A shows static blood on control PET (1000X and 5000X magnification). Panel B shows static blood on oxygen plasma activated; 24 hr engraftment in 2.5 L SS box (lOOOx and 5000x magnification). Panel C shows static blood on oxygen plasma activated; 24 hr engraftment in 10 mL plastic tube (lOOOx and 5000x magnification). Panel D shows static blood on oxygen plasma activated; 24 hr engraftment in 25 mL glass tube (lOOOx and 5000x magnification). Panel E shows static blood on oxygen plasma activated; 4 hr engraftment in 2.5 L SS box (lOOOx and 5000x magnification). Panel F shows static blood on oxygen plasma activated; 4 hr engraftment in 10 mL plastic tube (lOOOx and 5000x magnification). Panel G shows static blood on oxygen plasma activated; 4 hr engraftment in 25 mL glass tube (lOOOx and 5000x magnification).

[0296] Outcome: In this study a 4 hr HPG engraftment time with oxygen plasma activation appears to produce a functional coating that is anti -thrombotic.

[0297] Conclusions

[0298] The focus of this study was to identify process conditions to produce a suitable HPG coating on woven PET devices. Engraftment time and temperature were the two main parameters investigated in this study, with the expectation that reducing either or both would reduce shrinkage and any other impact on the physical and mechanical properties of woven PET. [0299] The study found that 1) HPG engraftment times for PET can be reduced to 4 hours at 80°C, and 2) engraftment temperature can be reduced from 80°C to at least 65°C and still produce an HPG coating.

[0300] An oxygen plasma, to activate the PET surface before HPG engraftment, was found to be superior to argon plasma.

EXAMPLE 3 - Tensile strength of HPG Coated woven PET Graft

[0301] To further investigate the impact of the HPG vapour coating process on PET graft material, another study used the lower HPG engraftment temperature of 60°C and various plasma activation parameters to determine if a functional HPG coating could be applied that did not impact on the mechanical properties of woven PET.

[0302] Reagents and materials

[0303] Glycidol (96%, Lot# MKCC2224) was purchased from Sigma-Aldrich and used without further purification. Absolute ethanol (ChemSupply) and RO water were used as solvents. Woven PET grafts were supplied by ATEX (diameter: 10 mm, length: 265 mm).

[0304] Plasma activation process

[0305] The plasma activation step was performed using a commercial plasma reactor (Dimensions of the chamber: 250mm x 240mm x 605mm) (Nano, Diner electronic GmbH, Germany). The reactor is equipped with a power generator delivering a power range 0-500W at 100kHz. Samples to be activated were mounted on the ground electrode. The chamber was pumped down to reach a base pressure below 1 x 10' 3 mbar. The following plasma conditions were kept constant - plasma gas flow rate of 4 seem, 100 W power for 20 min. In this example the main variable was the plasma gas used: Condition 1 - oxygen and Condition 2 - air.

[0306] HPG grafting process

[0307] HPG grafting process was performed by placing the plasma activated samples into a sealed container together with a certain volume of glycidol, at a ratio of 2-4 mL of glycidol per litre of container volume. Containers were then placed in an oven at a 60°C for 48h. The samples were then taken out and washed thoroughly with RO water (3 x 5min per cycle) followed by ethanol (3 x 5min per cycle). Finally, the samples were left to dry in a laminar flow cabinet for 60 min before being stored in a clean container.

[0308] Analytical Methods

[0309] XPS was as described above.

[0310] Blood spot analysis - HPG-coated PET graft was also assessed for antithrombotic properties using fresh human blood. The coated and uncoated PET graft samples were placed on HPG-coated glass slides in a 4-well polystyrene plate. 10 pL of freshly collected human blood, from healthy donors and containing no anti-coagulant, was placed on each sample and incubated at room temperature for 10 min. Washed samples were fixed in 2 mL of 4% paraformaldehyde (PF A) in PBS for 20 min and prepared for visual analysis by scanning electron microscopy (SEM). A Zeiss Merlin FEG SEM with SDD EDS was used to image the samples with the InLens detector and 3 kV beam energy at a working distance of approximately 2.5 mm. Samples were analysed for differences in platelet binding and fibrin networks between the HPG-coated and control PET.

[0311] Tensile testing - samples were tested on an MTS Exceed Series 40 Electromechanical Universal Test System with a 100 kN load cell. Samples were loaded in uniaxial tension and tested at a displacement rate of 25.4 mm/min. A gauge length of 40 mm was used.

[0312] Results

[0313] XPS analysis of HPG coating - A summary of the XPS elemental concentrations measured on the inner and outer surfaces of the untreated and HPG coated PET grafts is presented in Figures 20 & 21. The surface chemistry of the untreated and HPG coated PET grafts, for both conditions 1 & 2, confirmed the presence of the HPG coating by the reduction in the C signal and increase in the O signal (Figure 20) and by the reduction in the C-C/C-H signal and increase in the C-0 signal (Figure 21). [0314] Tensile testing - the PET samples become slightly less elastic when coated with HPG, as indicated by the reduction in the gradient of the elastic region of the forcedisplacement curve (Figure 22). The PET graft with HPG applied under condition 2 is slightly stiffer than the PET with HPG applied under condition 1. All samples withstood a displacement of greater than 10 mm (25% of their original length) and fracture when forces of 300 N or greater were applied. The force-displacement curves for the tensile testing are also shown in Figure 22. Application of the HPG coating had no significant impact on the yield force and all the samples appeared to yield between 90 and 100 N of applied force.

[0315] Blood Analysis - HPG-coated PET grafts were also tested for blood compatibility using an in-house blood spot analysis technique. Uncoated control PET consistently showed medium to high levels of thrombosis across multiple donors after just 10 min of exposure to blood. Large areas covered in fibrin networks could be found across the surface along with high numbers of bound and activated platelets (Figure 23). Following HPG coating, the PET samples demonstrated a resistance to platelet binding and activation along with greatly reduced fibrin network formation (Figures 24). Conditions 1 & 2 for applying the HPG coating did not display significantly different antithrombotic responses in this test system. It should also be noted that the fractures observed in the PET fibres, using the original HPG process (Example 2, above), were absent on the surface of the HPG-treated samples using the optimised process. This further suggests that the new conditions 1 & 2 are less damaging to the PET fibres.

[0316] Conclusions

[0317] This study shows that it is possible to apply the HPG coating on PET grafts using the vapour method, without significant impact on the tensile strength of the material. The in vitro blood test performed in this study also shows that the optimised HPG processes (condition 1 & 2) produce a coating with anti -thrombotic properties.

EXAMPLE 4 - HPG COATING OF MEDICAL GRADE SILICONE

[0318] Poly dimethylsiloxane (PDMS) is commonly used in a number of medical devices such as catheters, tubing and breast implants. The stability of HPG on a more flexible and highly mobile polymer was investigated in this example. PDMS sheets were coated with an HPG polymer by exposure to a glycidol vapour, as described in the previous examples.

[0319] Reagents and materials

[0320] Glycidol (96%, Lot# MKCC2224) was purchased from Sigma-Aldrich and used without further purification. Absolute ethanol (ChemSupply) and RO water were used as solvents. PDMS was purchased from Polymer Systems Technology (UK).

[0321] Plasma activation process

[0322] The plasma activation step was performed using a commercial plasma reactor (Dimensions of the chamber: 250mm x 240mm x 605mm) (Nano, Diner electronic GmbH, Germany). The reactor is equipped with a power generator delivering a power range 0-500W at 100kHz. Samples to be activated were mounted on the ground electrode. The chamber was pumped down to reach a base pressure below 1 x 10' 3 mbar. The following plasma conditions were used - Argon flow rate of 4 seem, 100 or 200 W power for 20 min.

[0323] HPG grafting process

[0324] HPG grafting process was performed by placing the plasma activated PDMS samples into a 250 mL sealed container together with 1 mL glycidol, avoiding direct contact of PDMS with the liquid glycidol. Containers were then placed in an oven at 80°C for 24 h. The samples were then taken out and washed thoroughly with RO water (3 x 5min per cycle) followed by ethanol (3 x 5min per cycle). Finally, the samples were left to dry in a laminar flow cabinet for 60 min before being stored in a clean container.

[0325] To assess the stability of the HPG polymer coating on PDMS, samples were analysed for their surface chemistry and water contact angle up to 4 weeks after the HPG was engrafted.

[0326] Analytical Methods [0327] XPS was as described above.

[0328] Water contact angle (WCA) measurements were made according to the method described above.

[0329] Results

[0330] In this study the impact of the power applied to the argon gas during the plasma activation step was tested as a variable for the engraftment of HPG onto the PDMS.

[0331] The XPS analysis of freshly coated PDMS (Figure 25A & B - orange bar), showed a reduction in the proportion of carbon and silicon in the HPG coated samples compared with the uncoated PDMS (Figure 25 A & B - dark blue bar). At the same time, there was a corresponding increase in the level of oxygen on the surface (Figure 25A & B). This is consistent with the formation of an HPG polymer coating from the activated PDMS surface. Comparing 100 W (Figure 25 A) with 200 W of power (Figure 25B) in the activation step, there did not appear to be a significant difference when looking at the elemental proportions.

[0332] When the high resolution Cis spectrum was analysed, there was an analogous reduction in the C-Si functional group with a large increase in the C-0 functional group (Figure 26A & B). This is characteristic of HPG polymer engraftment on the surface. Comparing 100 W (Figure 26 A) with 200 W (Figure 26B) for activation, there was a slight but not significant improvement in the HPG coating at the higher power, as evidenced by the level of the C-0 peak in the fresh samples (orange bars).

[0333] Coated PDMS samples were left to age for up to 4 weeks at room temperature and analysed by XPS at 1, 2 and 4 weeks after coating. The data in Figures 25 and 26 show that the chemistry of the surface does not substantially change over that time, indicating that the bulk silicone has not undergone any rearrangement with the molecules at the surface. This is indicative of a stable HPG coating on the PDMS. Comparing the results for 100 W samples with the 200 W samples, shows there is no difference in the overall results, but activation powers below 100 W or above 200 W may produce different results. [0334] To confirm the chemical profile of the surface of the coated PDMS, WCA was measured on fresh and aged samples out to 4 weeks, compared with uncoated PDMS. PDMS is very hydrophobic and an HPG coating, which is hydrophilic, would cause a reduction in the WCA. The results in Figure 27 show that the WCA for the 100 W (A) or 200 W (B) samples were significantly lower than uncoated PDMS. Over the 4-week storage the WCA values did not appear to change over time, supporting the ascertain from the XPS data (Figures 25 & 26) that the HPG coating is stable over that period. Taking the average of the WCA values for 100 W samples (Figure 27 A) across the 4 weeks and comparing that with the corresponding average WCA values for the 200 W samples (Figure 27b), the 200 W samples had on average a lower WCA than the 100 W samples. The WCA results would indicate that the HPG engraftment may be improved by using higher powers during the plasma activation step.

[0335] Conclusion

[0336] This example confirms that it is possible to engraft an HPG polymer directly from silicone after it has been activated with a non-depositing plasma gas (argon), by exposing activated PDMS to a vapour of glycidol. The results also indicate that the HPG polymer is stable on PDMS.

EXAMPLE 5 - HPG COATING OF STAINLESS STEEL AND COBALT /CHROMIUM ALLOYS

[0337] Many implantable devices comprise metal alloy components such as stainless steel and cobalt/chromium. This example was designed to test the glycidol vapour coating process on these metal alloys.

[0338] Reagents and materials

[0339] Glycidol (96%, Lot# MKCC2224) was purchased from Sigma-Aldrich and used without further purification. Absolute ethanol (ChemSupply) and RO water were used as solvents. Stainless steel sheet was purchased from Goodfellows (UK) and cobalt/chromium stent from eSutures, Inc. (USA).

[0340] Plasma activation process [0341] The plasma activation step was performed using a commercial plasma reactor (Dimensions of the chamber: 250mm x 240mm x 605mm) (Nano, Diner electronic GmbH, Germany). The reactor is equipped with a power generator delivering a power range 0-500W at 100kHz. Samples to be activated were mounted on the ground electrode. The chamber was pumped down to reach a base pressure below 1 x 10' 3 mbar. The following plasma conditions were used for both metal alloys - oxygen at a flow rate of 4 seem, 200 W power for 20 min.

[0342] HPG grafting process

[0343] HPG grafting process was performed by placing the plasma activated metal alloy samples into a 250 mL sealed container together with 1 mL glycidol, avoiding direct contact of samples with the liquid glycidol. Containers were then placed in an oven at 80°C or 100°C for 24 h. The samples were then taken out and washed thoroughly with RO water (3 x 5min per cycle) followed by ethanol (3 x 5min per cycle). Finally, the samples were left to dry in a laminar flow cabinet for 60 min before being stored in a clean container.

[0344] Analysis

[0345] XPS surface analysis of the sample was conducted as described above.

[0346] Results

[0347] The XPS analysis of the stainless steel sheet showed (Figure 28) a slight increase in the carbon and oxygen signal on the surface. This was consistent with a fall in the signal for the metallic elements on the surface (A). This was matched with a correspondingly significant increase in the C-0 functional group on the surface (B); corresponding to the presence of the HPG polymer on the surface.

[0348] The XPS analysis of the cobalt/chromium stent showed (Figure 29) a slight increase in the carbon and oxygen signal on the surface (A). This was matched with a correspondingly significant increase in the C-0 functional group on the surface (B); corresponding to the presence of the HPG polymer on the surface. EXAMPLE 6 - HPG COATING OF WATER TREATMENT MEMBRANES (PVDF, PTFE and PP)

[0349] Plasma activation process

[0350] PVDF and PTFE membranes were activated with a commercial plasma reactor (Nano, Diner electronic GmbH, Germany - Dimensions of the chamber: 250mm x 240mm x 605mm), equipped with a power generator delivering a power range 0-500W at 100kHz.

[0351] Membranes were mounted on the ground electrode and the chamber was pumped down to reach a base pressure below 7 x 10' 3 mbar. The following plasma conditions were used for both the PVDF and PTFE membranes - Argon at a flow rate of 4 seem, 200 W power for 10 min (PVDF) and for 15min (PTFE).

[0352] The plasma activation step for polypropylene membranes was performed using a custom-built stainless steel plasma reactor (Dimensions of the chamber: 650mm x 650mm x 150mm). The reactor was equipped with an RF power generator with a frequency of 13.65mHz and a matching network. Membranes were mounted on the ground electrode prior to the chamber being pumped down to a base pressure below 5 x 10' 3 mbar. The following plasma conditions were used for activating the polypropylene membranes - Air at a pressure of O.Olmbar, 100 W power for 5 min.

[0353] HPG grafting process

[0354] HPG grafting was performed by placing each plasma activated membrane sample into a 250 mL sealed container together with 2 mL glycidol, avoiding direct contact of samples with the liquid glycidol. Containers were then placed in an oven at 100°C for 24 h. The samples were taken out and washed thoroughly with irrigation water (3 x 5min per cycle) followed by ethanol (3 x 5min per cycle). Finally, the samples were dried in a laminar flow cabinet for 60 min before being stored in a clean container.

[0355] Analysis

[0356] XPS surface analysis of the sample was conducted as described above and the results are provided in Figures 30 to 32. [0357] Membrane filtration performance was analysed using 5cm circular pieces of uncoated and coated PVDF membranes and the results are provided in Figures 33 and 34.

[0358] Membrane filtration analysis with humic acid

[0359] The clean membrane resistances of the HPG-coated (Coated) and uncoated (Pristine) membranes before filtration were similar at a value of 2.2 x 10 11 m’ 1 . This was checked by filtering with DI water and monitoring the flux and transmembrane pressure. This confirmed that the coating did not affect the filtration performance.

[0360] A filtration test was performed with humic acid (100 mg/L) as foulant. To assess transmembrane pressure, a supra-critical flux of 80 L/m 2 h was applied to induce fouling and provide an assessment of severity, with a lower transmembrane pressure indicating less severe fouling.

[0361] As shown in Figure 33(A), the HPG-coated (Coated) membrane clearly demonstrated less fouling (anti-fouling) compared to the Pristine (uncoated) membrane. The permeate flux was stable at 80 L/m 2 h for both cases throughout the filtration period, enabling a fair comparison of the membranes.

[0362] To assess if HPG coating improved the ability to clean a fouled membrane, cleaning of the membrane was performed at the end of filtration by soaking the membranes in 0.1 M NaOH solution followed by 0.1 M citric acid solution for 1 hr. The fouled membrane resistance was checked after each cleaning step through filtration with DI water.

[0363] As shown in Figure 34(A), the fouling resistances (total resistance subtracted by clean membrane resistance) of the uncoated (Pristine) and HPG-coated (Coated) membranes after filtration were 2.0 x 10 12 m' 1 and 1.2 x 10 12 m' 1 respectively. For the uncoated (Pristine) membrane, cleaning with NaOH and citric acid removed 76% of the fouling resistance. For the HPG-coated (Coated) membrane, the fouling resistance is almost fully removed (99% reduction) after NaOH and citric acid cleaning. A substantial amount of fouling resistance (91%) was removed from the HPG-coated membrane with just NaOH cleaning. These results show that the HPG-coated membrane has less fouling than the untreated membrane and can be cleaned more easily than the uncoated membrane.

[0364] Membrane filtration analysis with sodium alginate

[0365] The clean membrane resistances of the HPG-coated (Coated) and uncoated (Pristine) membranes before filtration were checked to be similar with a value of 2.2 x 10 11 m’ 1 . This was checked by filtering with DI water and monitoring the flux and transmembrane pressure, indicating that the coating did not impact the original membrane filtration performance.

[0366] A filtration test was performed with sodium alginate (100 mg/L) as a foulant. To assess transmembrane pressure, a constant permeate flux of 80 L/m 2 h was applied for 4.5 hrs. As shown in Figure 33(B), a low transmembrane pressure was maintained for the HPG-coated (Coated) membrane throughout the filtration period - indicating low fouling. In comparison, the uncoated (Pristine) membrane showed a clear increase in transmembrane pressure up to 4 hrs, after which it increased rapidly. This illustrates the ability of the HPG coating to reduce membrane fouling.

[0367] To assess if HPG coating improved the ability to clean a fouled membrane, cleaning of the membrane was performed at the end of filtration by soaking the membranes in 0.1 M NaOH solution followed by 0.1 M citric acid solution for 1 hr each. The fouled membrane resistance was checked after each cleaning step through filtration with DI water.

[0368] As shown in Figure 34(B), the fouling resistances of the uncoated (Pristine) and HPG-coated (Coated) membranes at the end of filtration were 1.9 x 10 12 m' 1 and 3.0 x 10 11 m’ 1 , respectively. The majority of the foulants on both the Uncoated (Pristine) and HPG-coated (Coated) membranes were removed with NaOH cleaning, corresponding to 87% and 84% removal in fouling resistance respectively. The fouling resistance remaining after citric acid cleaning was 6.2 x 10 10 m' 1 for the uncoated (Pristine) membrane, and 8.9 x 10 9 m' 1 for the HPG-coated (Coated) membrane. These results show that the HPG-coated membrane has less fouling than the untreated membrane and that the HPG-coated membrane can be cleaned of sodium alginate as a foulant. [0369] Although the present disclosure has been described with reference to particular embodiments, it will be appreciated that the disclosure may be embodied in many other forms. It will also be appreciated that the disclosure described herein is susceptible to variations and modifications other than those specifically described. It is to be understood that the disclosure includes all such variations and modifications. The disclosure also includes all of the steps, features, compositions and compounds referred to, or indicated in this specification, individually or collectively, and any and all combinations of any two or more of the steps or features.

[0370] Also, it is to be noted that, as used herein, the singular forms “a”, “an” and “the” include plural aspects unless the context already dictates otherwise.

[0371] Throughout this specification, unless the context requires otherwise, the word “comprise”, or variations such as “comprises” or “comprising”, will be understood to imply the inclusion of a stated element or integer or group of elements or integers but not the exclusion of any other element or integer or group of elements or integers.

[0372] Reference to any prior art in this specification is not, and should not be taken as, an acknowledgment or any form of suggestion that this prior art forms part of the common general knowledge in any country.

[0373] The subject headings used herein are included only for the ease of reference of the reader and should not be used to limit the subject matter found throughout the disclosure or the claims. The subject headings should not be used in construing the scope of the claims or the claim limitations.

[0374] The description provided herein is in relation to several embodiments which may share common characteristics and features. It is to be understood that one or more features of one embodiment may be combinable with one or more features of the other embodiments. In addition, a single feature or combination of features of the embodiments may constitute additional embodiments.

[0375] All methods described herein can be performed in any suitable order unless indicated otherwise herein or clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein, is intended merely to better illuminate the example embodiments and does not pose a limitation on the scope of the claimed invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential.

[00333] Future patent applications may be filed on the basis of the present application, for example by claiming priority from the present application, by claiming a divisional status and/or by claiming a continuation status. It is to be understood that the following claims are not intended to limit the scope of what may be claimed in any such future application. Nor should the claims be considered to limit the understanding of (or exclude other understandings of) the disclosed invention.