Metal material with biological properties and articles made therefrom

文档序号：1803280 发布日期：2021-11-05 浏览：26次中文

阅读说明：本技术 具有生物特性的金属材料及由其制成的制品 (Metal material with biological properties and articles made therefrom ) 是由由佐史江托马斯·J·韦伯斯特小松隆史于 2020-03-20 设计创作，主要内容包括：本发明的方面涉及具有生物特性如抗菌性的金属材料和由所述金属材料制成的制品。(Aspects of the present invention relate to a metal material having biological properties such as antibacterial properties and an article made of the metal material.)

1. A stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.1 μ ι η to 3 μ ι η, configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

2. A stainless steel metallic material comprising a uniform grain nanostructure and having an average grain size of 0.2 μ ι η to 1 μ ι η.

3. A stainless steel metallic material comprising a uniform grain nanostructure and having an average grain size of 0.2 μ ι η to 0.5 μ ι η.

4. The stainless steel metallic material of any one of claims 1 to 3 wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetized.

5. The stainless steel metallic material of any one of claims 1 to 3 wherein the metallic material is type 316 stainless steel metal, and wherein the material is magnetized.

6. A type 304 stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

7. A type 316 stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η, configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

8. The metallic material of any of claims 1-3 or 6-7, wherein the metallic material inhibits adsorption or growth of microorganisms on the metallic material by at least 50%.

9. The metallic material of any of claims 1-3 or 6-7, wherein the metallic material is magnetized.

10. The metallic material of any of claims 1-3 or 6-7, wherein the metallic material reduces inflammatory cell adsorption or growth, reduces bacterial adsorption or growth, increases osteoblast adsorption or growth, increases endothelial cell adsorption or growth, or a combination thereof.

11. The metallic material of claim 9, wherein said metallic material reduces inflammatory cell adsorption or growth, reduces bacterial adsorption or growth, increases osteoblast adsorption or growth, increases endothelial cell adsorption or growth, or a combination thereof.

12. The metallic material of any of claims 1-3 or 6-7, wherein the microorganism is a gram positive bacterium.

13. The metallic material of any of claims 1-3 or 6-7, wherein the microorganism is a gram-negative bacterium.

14. The metallic material according to any one of claims 1-3 or 6-7, wherein the microorganism is one of Staphylococcus aureus (Staphylococcus aureus), Staphylococcus epidermidis (Staphylococcus epidermidis), methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli (E.coli), multidrug-resistant Escherichia coli, Pseudomonas aeruginosa (Pseudomonas aeruginosa).

15. A wire or rod made of the stainless steel metallic material of any one of claims 1-3 or 6-7.

16. A medical device made of the stainless steel metallic material of any one of claims 1-3 or 6-7.

17. A foil made of the stainless steel metal material of any one of claims 1-3 or 6-7.

18. An appliance made of the stainless steel metallic material of any one of claims 1-3 or 6-7.

19. Kitchen appliance made of a stainless steel metal material according to any of claims 1-3 or 6-7.

20. A wire or rod made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η, the stainless steel metallic material configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

21. The wire or rod according to claim 20, wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetized.

22. The wire or rod according to claim 20, wherein the metallic material is type 316 stainless steel metal, and wherein the material is magnetized.

23. A medical device made from a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η, the stainless steel metallic material configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

24. The medical device of claim 23, wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetized.

25. The medical device of claim 23, wherein the metallic material is type 316 stainless steel metal, and wherein the material is magnetized.

26. A foil made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η, the stainless steel metallic material configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

27. A foil according to claim 26, wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetised.

28. A foil according to claim 26, wherein the metallic material is type 316 stainless steel metal, and wherein the material is magnetised.

29. An apparatus made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

30. The apparatus of claim 29, wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetized.

31. The apparatus of claim 29, wherein the metallic material is type 316 stainless steel metal, and wherein the material is magnetized.

32. Stainless steel wire comprising a uniform grain nanostructure and having an average grain size of 0.2 to 0.5 μm, wherein the wire has antibacterial properties.

33. The wire according to claim 32 wherein said metallic material is type 304 stainless steel metal and wherein said material is magnetized.

34. The wire according to claim 32 wherein said metallic material is type 316 stainless steel metal and wherein said material is magnetized.

35. A stainless steel medical device comprising a uniform grain nanostructure and having an average grain size of 0.2 μ ι η to 0.5 μ ι η, wherein the medical device has antimicrobial properties.

36. The metal device of claim 35, wherein the metallic material is type 304 stainless steel metal, and wherein the material is magnetized.

37. The metal device of claim 35, wherein the metal material is type 316 stainless steel metal, and wherein the material is magnetized.

38. The medical device of any one of claims 35-37, wherein the medical device inhibits adsorption or growth of microorganisms on the medical device by at least 50%.

Technical Field

Aspects of the present disclosure relate to metallic materials and metallic devices having nanostructures of uniform average grain size and biological properties.

Background

Approximately 400,000 vascular catheter-related bacteremia and fungemia are reported annually in the united states. Such infections can be life-threatening and are often difficult to treat. The bactericidal effect that reduces or prevents colonization is typically by coating the device with an antibiotic.

Alternatively, it may be desirable for the implantable device to increase or decrease the adhesion and/or growth of eukaryotic cells.

Disclosure of Invention

Aspects of the present disclosure relate to metallic materials having grain sizes that provide enhanced antimicrobial action, improved surface energy for eukaryotic cell growth, or a combination thereof, and articles made therefrom.

Aspects of the present disclosure relate to a metallic material comprising uniform grains having an average grain size of 100nm to 3 μ ι η, more particularly 200nm to 1 μ ι η, and more particularly 200nm to 500nm, configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

In some embodiments, the metal material inhibits the adsorption or growth of microorganisms on the metal material by at least 50%. In some embodiments, the microorganism is a gram-positive bacterium. In some embodiments, the microorganism is a gram-negative bacterium. In some embodiments, the microorganism is one of staphylococcus aureus, staphylococcus epidermidis, methicillin-resistant staphylococcus aureus (MRSA), escherichia coli, multidrug-resistant (MDR) escherichia coli, or pseudomonas aeruginosa.

In some embodiments, the metallic material reduces inflammatory cell adsorption or growth, reduces bacterial adsorption or growth, increases osteoblast adsorption or growth, increases endothelial cell adsorption or growth, or a combination thereof.

In some embodiments, the metallic material has an average grain size that substantially inhibits the adsorption or growth of microorganisms determined from a response profile obtained by culturing microorganisms on a metallic material having grains of different average grain sizes and plotting the number of microorganisms after culturing against the average grain size.

In some embodiments, the grains have an average grain size of 0.1 μm or more and 3 μm or less. In some embodiments, the grains have an average grain size of 0.2 μm or more and 1 μm or less. In some embodiments, the grains have an average grain size of 0.2 μm or more and 0.5 μm or less. In some embodiments, the grains have an average grain size of 0.1 μm or more and 1 μm or less. In some embodiments, the grains have an average grain size of 0.2 μm or more and 0.5 μm or less.

In some embodiments, the metallic material may be stainless steel. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel.

In some embodiments, the metallic material is a wire or rod. In some embodiments, the average grain size is from 0.1 μm or more to 3 μm or less. In some embodiments, the average grain size is from 0.2 μm or more to 1 μm or less. In some embodiments, the average grain size is 0.2 μm to 0.5 μm.

Some aspects of the present disclosure relate to medical devices made from the metallic materials described herein.

Some aspects of the present disclosure relate to foils made from the metallic materials described herein.

Some aspects of the present disclosure relate to instruments made from the metallic materials described herein.

Some aspects of the present disclosure relate to a metal wire comprising crystal grains having an average crystal grain size of 0.2 μm to 1 μm, wherein the metal wire has antibacterial properties. In some embodiments, the average grain size is 0.2 μm to 0.5 μm.

Some aspects of the present disclosure relate to a metallic medical device comprising crystal grains having an average crystal grain size of 100nm to 3 μm, wherein the medical device has antibacterial properties. In some embodiments, the average grain size is 0.2 μm to 1 μm. In some embodiments, the average grain size is 0.2 μm to 0.5 μm.

In some embodiments, the metallic medical device inhibits the adsorption or growth of microorganisms on the metallic medical device by at least 50%.

Aspects of the present disclosure relate to stainless steel metallic materials and articles made therefrom. Aspects of the present disclosure relate to a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.1 μ ι η to 3 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodiments, the stainless steel metallic material comprises a uniform grain nanostructure and has an average grain size of 0.2 μm to 1 μm. In some embodiments, the stainless steel metallic material comprises a uniform grain nanostructure and has an average grain size of 0.2 μm to 0.5 μm. In some embodiments, the metallic material is type 304 stainless steel metal. In some embodiments, the metallic material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal. In some embodiments, the metallic material is magnetized.

Some aspects of the present disclosure relate to a type 304 stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

Some aspects of the present disclosure relate to a type 316 stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

In some embodiments, the metal material inhibits the adsorption or growth of microorganisms on the metal material by at least 50%.

In some embodiments, the metallic material is magnetized.

In some embodiments, the microorganism is a gram-positive bacterium. In some embodiments, the microorganism is a gram-negative bacterium. In some embodiments, the microorganism is one of staphylococcus aureus, staphylococcus epidermidis, methicillin-resistant staphylococcus aureus (MRSA), escherichia coli, multi-drug resistant escherichia coli, pseudomonas aeruginosa.

In some embodiments, the article is a wire or rod. In some embodiments, the article is a medical device. In some embodiments, the article is a stainless steel metallic material. In some embodiments, the article is an appliance. In some embodiments, the article is a kitchen utensil.

Some aspects of the present disclosure relate to a wire or rod made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized. In some embodiments, the wire or rod, wherein the metallic material is type 316 stainless steel metal and the material is magnetized.

Some aspects of the present disclosure relate to medical devices made from stainless steel metallic materials comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of microorganisms, (ii) promote adhesion, growth, or a combination thereof of predetermined eukaryotic cells, or (iii) inhibit adhesion, growth, or a combination thereof of predetermined eukaryotic cells. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal, and the material is magnetized.

Some aspects of the present disclosure relate to foils made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of microorganisms, (ii) promote adhesion, growth, or a combination thereof of predetermined eukaryotic cells, or (iii) inhibit adhesion, growth, or a combination thereof of predetermined eukaryotic cells. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal, and the material is magnetized.

Some aspects of the present disclosure relate to an apparatus made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.2 μ ι η to 0.5 μ ι η configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized.

In some embodiments, the metallic material is type 316 stainless steel metal, and the material is magnetized.

Some aspects of the present disclosure relate to stainless steel wire comprising a uniform grain nanostructure and having an average grain size of 0.2 μm to 0.5 μm, wherein the wire has antimicrobial properties. In some embodiments, the metallic material may be type 304 stainless steel.

In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal, and the material is magnetized.

Some aspects of the present disclosure relate to a stainless steel medical device comprising a uniform grain nanostructure and having an average grain size of 0.2 μm to 0.5 μm, wherein the medical device has antimicrobial properties. In some embodiments, the metallic material may be type 304 stainless steel. In some embodiments, the metallic material may be type 316 stainless steel. In some embodiments, the type 316 stainless steel is 316L stainless steel. In some embodiments, the metallic material is type 304 stainless steel metal, and the material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal, and the material is magnetized. In some embodiments, the medical device inhibits the adsorption or growth of microorganisms on the medical device by at least 50%.

Drawings

Fig. 1A is an example of a response profile obtained by plotting response amounts (CFU/ml) of gram-positive bacteria (staphylococcus aureus, methicillin-resistant staphylococcus aureus, staphylococcus epidermidis) against the average crystallite size of the crystallites, according to some embodiments.

Fig. 1B is an example of a response profile obtained by plotting the response volume (CFU/ml) of gram-negative bacteria (e.coli, pseudomonas aeruginosa) versus the average grain size of the grains, according to some embodiments.

Fig. 2A is an example of a response profile obtained by plotting the response volume (CFU/ml) of gram-positive bacterial MRSA versus the average grain size of polished or unpolished grains, according to some embodiments.

Fig. 2B is an example of a response profile obtained by plotting the response volume (CFU/ml) of the gram-negative bacterium pseudomonas aeruginosa relative to the average polished or unpolished grain size of the grains, according to some embodiments.

Fig. 3A is an example of a response profile obtained by plotting osteoblast viability against average unpolished grain size of grains, according to some embodiments.

Fig. 3B is an example of a response profile obtained by plotting osteoblast viability against average polished grain size of the grains, according to some embodiments.

Fig. 4A-4C illustrate the percent viability of human skin fibroblasts when grown on type 304 stainless steel metal samples having different grain sizes, according to some embodiments.

Fig. 5A-5C illustrate the percent viability of human skin fibroblasts when grown on type 316 stainless steel metal samples having different grain sizes, according to some embodiments.

FIG. 6 is a schematic illustration of a polishing method according to some embodiments.

Fig. 7 is a graph illustrating the percent viability of human fetal osteoblasts on titanium alloy in a cell growth assay, according to some embodiments.

Fig. 8 is a schematic illustration of a vibrating sample magnetometer used in accordance with some embodiments.

FIG. 9 is a response (CFU/cm) normalized to surface area for four bacterial strains (E.coli, MDR E.coli, MRSA, and Staphylococcus epidermidis), according to some embodiments²) An example of a response profile obtained by plotting against the average grain size of type 304 stainless steel.

FIG. 10 is a response (CFU/cm) normalized to surface area for three bacterial strains (MDR E.coli, MRSA, and S.aureus), according to some embodiments²) Examples of response spectra obtained by plotting average grain size and magnetic properties against type 304 stainless steel samples.

Fig. 11 is an example of a response profile obtained by plotting the response quantity (cell number) normalized to surface area of human fetal osteoblasts against the average grain size and magnetic properties of a type 304 stainless steel sample, according to some embodiments.

Fig. 12A-12C are examples of response profiles obtained by plotting the response (number of cells) normalized to surface area of human fetal osteoblasts against the average grain size and magnetic properties of a type 304 stainless steel sample, according to some embodiments. FIG. 12A is a response plot obtained by plotting the response (cell number) normalized to surface area of human fetal osteoblasts against type 304 stainless steel samples with an average grain size of 0.5 μm with different magnetizations (UT: untreated; DM: demagnetized; 0.1T, 0.5T, and 1.1T), according to some embodiments. Fig. 12B is a response plot obtained by plotting the response (cell number) normalized to surface area of human fetal osteoblasts against type 304 stainless steel samples with an average grain size of 5 μm with different magnetizations (UT: untreated; DM: demagnetized; 0.1T, 0.5T, and 1.1T), according to some embodiments. Fig. 12C is a response profile obtained by plotting the response (cell number) normalized to surface area of human fetal osteoblasts against type 304 stainless steel samples with an average grain size of 9 μm with different magnetizations (UT: untreated; DM: demagnetized; 0.1T, 0.5T, and 1.1T), according to some embodiments.

Fig. 13 illustrates a response profile for biofilm formation using safranin obtained by plotting absorbance at 570nm versus the average grain size of type 304 stainless steel, according to some embodiments.

Detailed Description

The metal material having fine grains is superior in characteristics such as strength, toughness, and corrosion resistance, compared to the metal material having coarse grains. Therefore, the metal material is widely used in various industrial applications such as steel plates and medical devices.

Some aspects of the present disclosure relate to metals processed to form recrystallized metallic material having the following average grain size: 0.01 to 3 μm, 0.02 to 3 μm, 0.05 to 3 μm, 0.1 to 3 μm, 0.2 to 3 μm, 0.5 to 3 μm, 1 to 3 μm, 2 to 3 μm, 0.01 to 2 μm, 0.02 to 2 μm, 0.05 to 2 μm, 0.1 to 2 μm, 0.2 to 2 μm, 0.5 to 2 μm, 1 to 2 μm, 0.01 to 1 μm, 0.02 to 1 μm, 0.05 to 1 μm, 0.1 to 1 μm, 0.2 to 1 μm, 0.5 to 1 μm, 0.01 to 0.6 μm, 0.02 to 0.6 μm, 0.05 to 0.6 μm, 0.1 to 6 μm, 0.02 to 0.5 μm, 0.01 to 0.6 μm, 0.02 to 0.06 μm, 0.1 to 0.6 μm, 0.0.01 to 0.0.0.0.5 μm, 0.01 to 0.5 μm, 0.05 to 0.05, 0.05 to 0.5 μm, 0.05 to 0.0.0.05, 0.0.0.0.05 to 0.0.0.0.05, 0.0.0.0.0.0.0.0.6 μm, 0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.05 to 5 to 3, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.3, 0.0.0.0.0.0.0.0.0.0., About 0.1 μm, about 0.2 μm, about 0.3 μm, about 0.4 μm, about 0.5 μm, about 0.6 μm, about 0.7 μm, about 0.8 μm, about 0.9 μm, about 1 μm, about 2 μm, about 3 μm or higher, or any range therebetween. In some embodiments, the metal is processed to form a recrystallized metallic material having an average grain size of 0.2 μm to 0.5 μm.

It is understood that the metallic material may have a uniform average grain size. In some embodiments, the metal comprises an average grain size of about 0.1 μm ± 20%, about 0.2 μm ± 20%, about 0.3 μm ± 20%, about 0.4 μm ± 20%, about 0.5 μm ± 20%, about 0.6 μm ± 20%, about 0.7 μm ± 20%, about 0.8 μm ± 20%, about 0.9 μm ± 20%, about 1 μm ± 20%, about 2 μm ± 20%, about 3 μm ± 20%, or any range therebetween. In some embodiments, the metal comprises an average grain size of about 0.1 μm ± 40%, about 0.2 μm ± 40%, about 0.3 μm ± 40%, about 0.4 μm ± 40%, about 0.5 μm ± 40%, about 0.6 μm ± 40%, about 0.7 μm ± 40%, about 0.8 μm ± 40%, about 0.9 μm ± 40%, about 1 μm ± 40%, about 2 μm ± 40%, about 3 μm ± 40%, or any range therebetween.

In some aspects, the metal is a type 304 stainless steel metal having an average grain size of about 0.10 μm to about 3 μm, such as 0.2 to 0.5 μm. In some embodiments, the type 304 stainless steel metal has a composition as described in table 4.

In some aspects, the metal is a type 316 stainless steel metal having an average grain size of about 0.1 μm to about 3 μm, such as 0.2 μm to 0.5 μm. In some embodiments, the type 316 stainless steel metal has a composition as set forth in table 5.

In some aspects, the metal is titanium or a titanium alloy having a grain size of about 0.8 μm to about 9 μm, such as 0.8 to 8.80 μm. In some embodiments, the titanium alloy is beta-titanium (Ti-15V-3Cr-3Sn-3Al), Ti-6Al-4V, or a combination thereof.

In some embodiments, the metallic material may be processed to adjust grain size to control cell adhesion, cell growth, or a combination thereof. In some embodiments, the metallic material or device may have an average grain size that inhibits adhesion, growth, or a combination thereof of bacteria. In some embodiments, the metallic material or device may have an average grain size that increases adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodiments, the metallic material or device may have an average grain size that inhibits adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodiments, the metallic material or device may have: (i) an average grain size that inhibits adhesion, growth, or a combination thereof of bacteria, (ii) an average grain size that promotes adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, and (iii) an average grain size that inhibits adhesion, growth, or a combination thereof of a predetermined eukaryotic cell.

Antibacterial metal material

The term "antimicrobial" as used herein refers to the property of preventing or reducing the growth or reproduction or adherence of microorganisms (e.g., bacterial and fungal organisms), or killing microorganisms.

The term "bacterial and fungal organism" as used in the present invention means all genera and species of bacteria and fungi, including but not limited to all cocci, bacilli and spirochetes. Non-limiting examples of bacteria include staphylococci (e.g., Staphylococcus epidermidis (Staphylococcus epidermidis), Staphylococcus aureus (Staphylococcus aureus)), enterococcus faecalis (enterococcus faecalis), Pseudomonas aeruginosa (Pseudomonas aeruginosa), escherichia coli (e.coli), clostridium difficile (clostridium difficile), and other gram positive and gram negative bacteria. Non-limiting examples of fungal organisms include Candida albicans (Candida albicans), Candida krusei (Candida krusei), Candida parapsilosis (Candida parapsilosis), Candida pseudotropicalis (Candida pseudotropicalis), Candida glabrata (Candida glabrata), Candida vitis (Candida lucitania), and Candida tropicalis (Candida tropicalis).

In some embodiments, the bacteria are gram positive bacteria including, but not limited to, staphylococcus aureus, staphylococcus epidermidis, methicillin-resistant staphylococcus aureus (MRSA), and the like. In some embodiments, the bacteria are gram-negative bacteria, including, but not limited to, pseudomonas aeruginosa, escherichia coli, Klebsiella pneumoniae (Klebsiella pneumoniae), Legionella pneumophila (Legionella pneumophila), Proteus mirabilis (Proteus mirabilis), Enterobacter cloacae (Enterobacter cloacae), Serratia marcescens (Serratia marcescens), Helicobacter pylori (Helicobacter pylori), Salmonella enteritidis (Salmonella enteritidis), and Salmonella typhi (Salmonella typhi).

Aspects of the invention provide metallic materials and methods for providing effective broad spectrum protection against infection, including but not limited to protection against drug-resistant staphylococci, MDR gram-negative bacteria (e.g., MDR pseudomonas aeruginosa) to metallic materials.

Aspects of the present invention provide a metal material having improved antibacterial properties. In some embodiments, the metallic material may be used in a medical device, such as an implant (e.g., without limitation, an orthopedic implant). In some embodiments, the metallic material may be used in surgical instruments, vascular stents, endoscopic instruments, catheter components, guide wires, K-wires, plates, pins, screws, needles, pacemaker leads, braces, and the like, or implantable medical devices. In some embodiments, the metallic material may be used in surgical instruments. In some embodiments, the metallic material may be used in a biosensor. In some embodiments, the metal material may be used in kitchen utensils. In some embodiments, metallic materials may be used for experimental tools. In some embodiments, a metallic material may be used for the k-wire.

Non-limiting examples of medical devices include vascular catheters, such as peripherally insertable central venous catheters, dialysis catheters, long-term tunneled central venous catheters, peripheral venous catheters, single and multi-lumen short-term central venous catheters, arterial catheters, pulmonary artery Swan-Ganz catheters, and the like, urinary catheters, other long-term urinary devices, tissue-adhesive urinary devices, renal stents, penile prostheses, vascular grafts, vascular access ports, wound drainage tubes, hydrocephalus shunts, ventricular drainage catheters, nerve and epidural catheters, nerve stimulators, peritoneal dialysis catheters, pacemaker capsules, artificial urinary sphincter, facet or temporary joint replacements, dilators, heart valves, orthopedic prostheses, spinal hardware, surgical site repair meshes (e.g., hernia meshes), endotracheal tubes, biliary stents, gastrointestinal tubes, colonic tract implants, male and female genitourinary implants, catheters, other long-term urinary devices, tissue-adhesive urinary devices, and the like, Cosmetic or reconstructive implants, stethoscope drums, orthopedic implants (e.g., joints (knee, hip, elbow, shoulder, ankle), prostheses, external fixation pins, intramedullary rods and nails), spinal implants), cardiac pacemakers, defibrillators, electronic device leads, adapters, lead extensions, implantable infusion devices, implantable pulse generators, implantable physiological monitoring devices, devices for positioning implantable pulse generators or implantable infusion devices under the skin, and devices for refilling implantable infusion devices or other medical and indwelling devices that may be subject to microbial infestation (e.g., refill needles and port access cannulas).

In some embodiments, the device is a stainless steel device and may be used in, but is not limited to, high speed surgical drills, vertebroplasty and kyphoplasty devices, minimally invasive surgical and endoscopic devices, orthopedic implants and surgical instruments.

In some embodiments, the device is a titanium device and may be used in, but is not limited to, orthopedic implants, dental implants, spinal implants, minimally invasive surgical instruments and endoscopic devices, as well as surgical instruments.

In some embodiments, the antimicrobial properties of the metal material are achieved without adding antimicrobial agents into or onto the metal material.

In some embodiments, the metal material inhibits adhesion of bacterial cells by 100%, 95%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or at least 5%, or any value or range therebetween.

Metal material for promoting cell adhesion or inhibiting cell adhesion

In some embodiments, it is desirable to increase the adhesion of cells on a metallic material, such as a metallic implant. In other embodiments, it is desirable to reduce or inhibit the adhesion of cells on a metallic material, such as a metallic implant. For example, it may be desirable to increase osteoblast adhesion on the surface of an orthopedic implant. In other instances, it may be desirable to increase the adhesion of endothelial cells and inhibit the adhesion of fibroblasts to the surface of a vascular stent or implant.

In some embodiments, the metallic materials described herein can improve adhesion and/or growth of eukaryotic cells, such as osteoblasts, fibroblasts, chondrocytes, endothelial cells, keratinocytes, smooth muscle cells, urothelial cells, osteoclasts, osteocytes, stem cells, mesenchymal stem cells, induced pluripotent stem cells, neurons, astrocytes, schwann cells, meningeal cells, epithelial cells, and the like.

In some embodiments, the metallic materials described herein have a surface energy that promotes cell adhesion and/or growth of some eukaryotic cells. In some embodiments, the metallic material increases cell adhesion and/or growth by at least 2%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or any value or range therebetween.

In some embodiments, the metal material described herein has a surface energy that inhibits cell adhesion and/or growth of other eukaryotic cells. In some embodiments, the metallic material reduces cell adhesion and/or growth by at least 2%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or any value or range therebetween.

In some embodiments, the metallic materials described herein have antimicrobial properties and improve the adhesion and/or growth of eukaryotic cells, such as osteoblasts, fibroblasts, endothelial cells, chondrocytes, endothelial cells, keratinocytes, smooth muscle cells, urothelial cells, osteoclasts, osteocytes, stem cells, mesenchymal stem cells, induced pluripotent stem cells, neurons, astrocytes, schwann cells, meninges cells, epithelial cells, and the like.

In some embodiments, the metallic materials described herein are antimicrobial and inhibit the adhesion and/or growth of eukaryotic cells, such as immune cells.

Described herein below are metallic materials and metallic materials for use in (i) reducing or inhibiting bacterial adhesion and/or growth; (ii) improving the adhesion and/or growth of eukaryotic cells such as osteoblasts, fibroblasts, and the like; (iii) reduce or inhibit adhesion and/or growth of immune cells; or (iv) (i) reduces or inhibits bacterial adhesion and/or growth, (ii) improves adhesion and/or growth of eukaryotic cells such as osteoblasts, fibroblasts, (iii) reduces or inhibits adhesion and/or growth of immune cells, or any combination of (i), (ii) and (iii).

Use of metallic materials

In some embodiments, the metallic material may be used in a medical device. For example, the metallic material may be used in vascular stents, endoscopic instruments, surgical instruments, catheter components, guide wires, k-wires, pins, plates, screws, and the like, or implantable medical devices.

In some embodiments, the metallic material may be used in a biosensor. In some embodiments, the metal material may be used in kitchen utensils. In some embodiments, metallic materials may be used for experimental tools.

Non-limiting examples of medical devices include vascular catheters, such as peripherally insertable central venous catheters, dialysis catheters, long-term tunneled central venous catheters, peripheral venous catheters, single and multi-lumen short-term central venous catheters, arterial catheters, pulmonary artery Swan-Ganz catheters, and the like, urinary catheters, other long-term urinary devices, tissue-adhesive urinary devices, renal stents, penile prostheses, vascular grafts, vascular access ports, wound drainage tubes, hydrocephalus shunts, ventricular drainage catheters, nerve and epidural catheters, nerve stimulators, peritoneal dialysis catheters, pacemaker capsules, artificial urinary sphincter, facet or temporary joint replacements, dilators, heart valves, orthopedic prostheses, spinal hardware, surgical site repair meshes (e.g., hernia meshes), endotracheal tubes, biliary stents, gastrointestinal tubes, colonic tract implants, male and female genitourinary implants, catheters, other long-term urinary devices, tissue-adhesive urinary devices, and the like, Cosmetic or reconstructive implants, stethoscope drums, orthopedic implants (e.g., joints (knee, hip, elbow, shoulder, ankle), prostheses, external fixation pins, intramedullary rods and nails, spinal implants), cardiac pacemakers, defibrillators, electronic device leads, adapters, lead extensions, implantable infusion devices, implantable pulse generators, implantable physiological monitoring devices, devices for positioning implantable pulse generators or implantable infusion devices under the skin, and devices for refilling implantable infusion devices or other medical and indwelling devices that may be subject to microbial infestation (e.g., refill needles and port access cannulas).

Some embodiments relate to surgical instruments.

In some embodiments, the device is a K-wire.

In some embodiments, the device is an implantable orthopedic implant.

In some embodiments, the device is a vascular stent.

Composition of

Known metallic materials for medical device applications may be used, and examples of the metallic material include iron, stainless steel, aluminum, silver, copper, titanium, tin, nickel, zinc, chromium, and alloys of these metallic materials. Among them, stainless steel is preferable in view of easy control of the grain size of the grains, versatility, easy availability, workability, and low toxicity. The stainless steel is not particularly limited, and may be any one of martensitic stainless steel, ferritic stainless steel, austenitic/ferritic stainless steel, and precipitation-hardened stainless steel.

In some embodiments, the metallic material is stainless steel or a stainless steel alloy. For example, the metallic material may be type 304 stainless steel or type 316 stainless steel. Type 316 stainless steel differs from type 304 in the presence of molybdenum. In some embodiments, the stainless steel material may comprise 6% to 22% nickel. In some embodiments, the stainless steel material may also contain other alloying elements, such as chromium (16% to 26%) for corrosion resistance. In some embodiments, the stainless steel may include manganese and molybdenum. In some embodiments, type 316 stainless steel may be used for medical devices.

In some embodiments, the metallic material is titanium or a titanium alloy. In some embodiments, the metallic material is cobalt chromium. In some embodiments, the metallic material is cobalt chromium molybdenum. In some embodiments, the metallic material is nitinol.

Nano-structure

According to an aspect of the present invention, the metal material provided herein has a nanostructure not limited to a surface. For example, the metallic material may retain its nanostructure throughout its processing, resulting in a metallic material with a uniform nanostructure.

The metallic material according to some embodiments is made of fine grains, which allows for application to a wide range of devices. The metal material has a uniform nanostructure. The predetermined average grain size nanostructure (e.g., 0.2 μm to 0.5 μm) is uniform throughout the material, i.e., on the surface and within the material.

The grains forming the metallic material according to some embodiments have an average grain size for controlling biological characteristics of the metallic material.

Aspects of the present invention are based on the following phenomena: the biological properties of the metallic material depend on the average grain size of the metallic material.

In some aspects of the present invention, the metallic materials described herein may have a grain size, surface free energy, and roughness such that: (i) reducing or inhibiting bacterial adhesion and/or growth; (ii) improving or increasing the adhesion and/or growth of eukaryotic cells such as osteoblasts, fibroblasts, and the like; (iii) reduce or inhibit adhesion and/or growth of immune cells; or (iv) (i) reduces or inhibits bacterial adhesion and/or growth, (ii) improves adhesion and/or growth of eukaryotic cells such as osteoblasts, fibroblasts, (iii) reduces or inhibits adhesion and/or growth of immune cells or any combination of (i), (ii) and (iii).

In some aspects of the invention, the metallic materials described herein may have a grain size that approximates the size of eukaryotic cells of the tissue of interest and promotes adhesion and/or growth of cells to the metal. Further, the metallic materials described herein can have a surface free energy that promotes cell adhesion and/or growth to the metal. Furthermore, the metallic materials described herein may have a roughness that promotes cell-to-metal adhesion and/or growth.

In some aspects of the invention, the metallic materials described herein can have a grain size that inhibits adhesion and/or growth of cells to the metal. Further, the metallic materials described herein can have a surface free energy that promotes cell adhesion and/or growth to the metal. Furthermore, the metallic materials described herein may have a roughness that inhibits adhesion and/or growth of cells to the metal. In some embodiments, the cell is a prokaryotic cell and/or a eukaryotic cell.

Some aspects of the present invention are based on the following phenomena: the antibacterial property of the metal material depends on the average crystal grain size of the metal material. In some embodiments, the metallic material has a predetermined average grain size of about 0.1 to 3 μm. In some embodiments, the metallic material has a predetermined average grain size of 0.2 to 1 μm. In some embodiments, the metallic material has a predetermined average grain size of 0.2 to 0.5 μm. In some embodiments, the metallic materials provided herein can inhibit the growth of microorganisms and/or improve the growth of osteoblasts and fibroblasts. In some embodiments, the metallic materials provided herein can inhibit the growth, immobilization, or both growth and immobilization (adsorption) of microorganisms. In some embodiments, the metallic materials provided herein can inhibit the growth, immobilization, or both growth and immobilization (adsorption) of immune cells. In some embodiments, the metallic materials provided herein promote the growth, fixation, or growth and fixation (adsorption) of predetermined eukaryotic cells.

Aspects of the invention relate to methods for inhibiting the growth, immobilization, or both growth and immobilization of microorganisms.

In some embodiments, the average grain size for inhibiting growth and/or immobilization of microorganisms may be 0.01 to 3 μm, 0.02 to 3 μm, 0.05 to 3 μm, 0.1 μm to 3 μm, 0.2 to 3 μm, 0.5 μm to 3 μm, 1 μm to 3 μm, 2 μm to 3 μm, 0.01 to 2 μm, 0.02 to 2 μm, 0.05 to 2 μm, 0.1 μm to 2 μm, 0.2 to 2 μm, 0.5 μm to 2 μm, 1 μm to 2 μm, 0.01 to 1 μm, 0.02 to 1 μm, 0.05 to 1 μm, 0.1 μm to 1 μm, 0.2 to 1 μm, 0.5 μm to 1 μm, 0.01 to 0.6 μm, 0.02 to 6, 0.05 to 0.5 μm, 0.5 μm to 0.5 μm, 0.05 to 0.5 μm, 0.5 μm to 2 μm, 0.05 to 1 μm, 0.5 μm to 0.5 μm, 0.5 μm to 2 μm, 0.5 μm, 0.6 μm to 2 μm, 0.5 μm, or more, About 0.02 μm, about 0.03 μm, about 0.04 μm, about 0.05 μm, about 0.06 μm, about 0.07 μm, about 0.08 μm, about 0.09 μm, about 0.1 μm, about 0.2 μm, about 0.3 μm, about 0.4 μm, about 0.5 μm, about 0.6 μm, about 0.7 μm, about 0.8 μm, about 0.9 μm, about 1 μm, about 2 μm, about 3 μm or higher, or any range therebetween. In some embodiments, the average grain size for inhibiting growth and/or immobilization of microorganisms may be 0.2 μm to 0.5 μm.

In some embodiments, the average grain size is greater than 0.1 μm but less than 3 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 1 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 0.7 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 0.5 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 0.4 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 0.3 μm. In some embodiments, the average grain size is greater than 0.3 μm but less than 0.5 μm. In some embodiments, the average grain size is greater than 0.4 μm but less than 0.5 μm.

Grain boundaries can be measured by Electron Back Scattering Diffraction (EBSD) and can show different atoms at low angles. The angular difference may be greater than 5 degrees. Each grain may be defined by an area surrounded by a grain boundary line. When the grain size is large, the shape is a unique and random polygon. As the particles become smaller, the shape becomes a smaller polygon, resembling a circle, cube, or rectangle. The diameter of the short and long or circular shape of the rectangle is about the average grain size.

In some embodiments, the metallic material has an average grain size of about 0.5 μm (+/-20%) or 0.2 μm to 0.5 μm, and inhibits the growth of gram positive and gram negative bacteria.

During the refinement of the grains, the chemical composition of the metallic material is not changed. Therefore, any metal of different chemical composition may be used as long as it is a metallic material having crystals or grains, such as titanium, a titanium-based material, stainless steel, a Co-Cr alloy, Co-Cr-Mo, nitinol, platinum, palladium, or the like.

In some embodiments, the metallic material has improved tensile strength and hardness over conventional stainless steel in addition to its antimicrobial properties.

As a method for adjusting the average grain size of the grains, a thinning method may be employed. Examples of the method include a rolling process, a shearing process, a compression process, a deformation process, and a combination of these processes for the metal raw material before the refinement. In this case, cooling or heating may be performed, or refinement may be performed in an atmosphere in the presence or absence of a specific gas (e.g., oxygen or nitrogen). Generally, the refinement is performed by plastic deformation caused by heating and recrystallization by cooling. The above process is carried out once or repeatedly a plurality of times to obtain the desired average grain size.

According to aspects of the present invention, devices formed of metallic materials provided herein have nanostructures that are not limited to surfaces. For example, the metallic material may retain its nanostructure throughout its processing, resulting in a metallic material with a uniform nanostructure.

According to some embodiments, the magnetic field of the metallic materials provided herein may alter the surface charge and the initial protein adsorption event, in turn altering bacterial attachment and colonization and/or growth of eukaryotic cells.

Polished/unpolished metal material

In some embodiments, the metal material may be polished to alter the surface roughness. In some embodiments, a method of polishing a metal material includes rough polishing using an abrasive film (see example 4).

The surface roughness can be calculated with an Atomic Force Microscope (AFM), and three different parameters-root mean square roughness (Rq), arithmetic roughness (Ra), and maximum height (Rz) can be obtained for metallic materials.

TABLE 1

TABLE 2

In some embodiments, the polished material has a surface roughness on the order of about 0.1nm to 100 μm.

Previous studies have shown that the optimal value of Surface energy for Inhibiting bacterial Growth is about 42mN/m (see Liu et al, "advancement of the Role of Polymer Surface Nanoscale Topograph on Inhibiting Bacteria addition and Growth" Biomaterials Science and Engineering,2016,2(1), pp 122- "130).

In some embodiments, the metal material has a surface energy value of 40 to 45mN/m, 40 to 47mN/m, 40 to 50mN/m, 40 to 55mN/m, 40 to 60mN/m, 35 to 45mN/m, 35 to 50mN/m, 35 to 55mN/m, 35 to 60mN/m, 30 to 45mN/m, 30 to 50mN/m, 30 to 55mN/m, 30 to 60 mN/m.

In some embodiments, the metallic materials described herein have a surface energy that promotes the growth of some eukaryotic cells. In some embodiments, the metal material described herein has a surface energy that inhibits the growth of other eukaryotic cells. For example, surface energy can promote the attachment and growth of endothelial cells and inhibit the attachment and/or growth of fibroblasts.

In some embodiments, the metal material that promotes the growth of eukaryotic cells has a surface energy value of 40 to 45mN/m, 40 to 47mN/m, 40 to 50mN/m, 40 to 55mN/m, 40 to 60mN/m, 35 to 45mN/m, 35 to 50mN/m, 35 to 55mN/m, 35 to 60mN/m, 30 to 45mN/m, 30 to 50mN/m, 30 to 55mN/m, 30 to 60 mN/m.

The optimum Ra, Rq, Rz can be calculated using the Khang equation, and the ideal surface energy is 45 mN/m.

Es＝E0,+ρ×reff

ES (surface energy)

Eo, s ═ base surface energy

roughness (reff)

Rho is coupling constant

In some embodiments, the metallic material may have a surface energy tailored to adsorb proteins that can reduce inflammatory cell function, reduce bacterial function, increase osteocyte function, increase endothelial cell function, or any combination of the foregoing.

In some embodiments, the metallic material may have an average grain size adjusted to adsorb proteins that can reduce inflammatory cell function, reduce bacterial function, increase osteocyte function, increase endothelial cell function, or any combination of the foregoing.

Without being bound by theory, the change in surface energy may in turn alter initial protein adsorption, e.g., for inhibiting bacterial attachment and colonization.

In some embodiments, the metal material may be polished or unpolished. In some embodiments, the polished and/or unpolished metallic material has an average grain size of about 100nm to 10 μm, such as less than 500nm, for example, about 100nm, to reduce attachment or growth of gram-positive and negative bacteria. In some embodiments, the polished and/or unpolished metallic material has an average grain size of preferably about 1 to 3 μm.

Shape/device

The shape of the metal material according to some embodiments is not particularly limited, and may take any shape, for example, a plate shape, a wire shape, a rod shape, a spherical shape, or a cylindrical shape. In some embodiments, the metallic material is in the shape of a wire or a thread.

In some embodiments, the metallic material is in the form of a plate or foil having a thickness of about 0.1mm to 1mm, such as 0.1mm, 0.2mm, 0.3mm, 0.4mm, 0.5mm, 0.6mm, 0.7mm, 0.8mm, 0.9mm, or 1 mm.

In some embodiments, the metallic material is in the form of a rod or wire having a diameter of 0.02mm to 6 mm.

Method for inhibiting microbial growth

A method of inhibiting microbial growth according to some embodiments is a method of using a metallic material that includes a predetermined average grain size.

In some embodiments, a metallic material, such as a stainless steel material, is provided, wherein each average grain size of the grains is adjusted to be within the following range: 0.01 to 3 μm, 0.02 to 3 μm, 0.05 to 3 μm, 0.1 to 3 μm, 0.2 to 3 μm, 0.5 to 3 μm, 1 to 3 μm, 2 to 3 μm, 0.01 to 2 μm, 0.02 to 2 μm, 0.05 to 2 μm, 0.1 to 2 μm, 0.2 to 2 μm, 0.5 to 2 μm, 1 to 2 μm, 0.01 to 1 μm, 0.02 to 1 μm, 0.05 to 1 μm, 0.1 to 1 μm, 0.2 to 1 μm, 0.5 to 1 μm, 0.01 to 0.6 μm, 0.02 to 0.6 μm, 0.05 to 0.6 μm, 0.1 to 6 μm, 0.02 to 0.5 μm, 0.01 to 0.6 μm, 0.02 to 0.06 μm, 0.1 to 0.6 μm, 0.0.01 to 0.0.0.0.5 μm, 0.01 to 0.5 μm, 0.05 to 0.05, 0.05 to 0.5 μm, 0.05 to 0.0.0.05, 0.0.0.0.05 to 0.0.0.0.05, 0.0.0.0.0.0.0.0.6 μm, 0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.05 to 5 μm, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.05 to 5 to 3, 0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.0.3, 0.0.0.0.0.0.0.0.0.0., About 0.1 μm, about 0.2 μm, about 0.3 μm, about 0.4 μm, about 0.5 μm, about 0.6 μm, about 0.7 μm, about 0.8 μm, about 0.9 μm, about 1 μm, about 2 μm, about 3 μm or higher, or any range therebetween. In some embodiments, the average grain size may be 0.2 μm to 0.5 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 1 μm. In some embodiments, the average grain size is greater than 0.1 μm but less than 3 μm. In some embodiments, the average grain size is greater than 0.2 μm but less than 0.5 μm.

In some embodiments, the average grain size of the grains used to impart the best inhibition of microbial growth is determined based on the response profile obtained in the above process.

Fig. 1A shows the antibacterial property of a metal material against gram-positive bacteria. For example, FIG. 1A shows that metallic materials with grain sizes of 0.5 μm, 1 μm, 1.5 μm, 3 μm, and 9 μm inhibit the growth/adhesion of gram-positive bacteria. Fig. 1B shows the antibacterial property of the metal material against gram-negative bacteria. For example, FIG. 1B shows that metallic materials with grain sizes of 0.5 μm, 3 μm, and 9 μm inhibit the growth/adhesion of gram-positive bacteria.

Method for promoting or inhibiting cell adhesion and/or growth

In some embodiments, the device may be implanted in the following anatomical locations: subcutaneous, intraperitoneal, intramuscular, intravascular, intraocular, intracerebral or other suitable sites.

In some embodiments, the nanostructure of the metallic material can be tailored to match proteins on the nanoscale and cells on the microscale. In some embodiments, the grain size may promote adhesion of endothelial cells or osteoblasts.

In some embodiments, an implantable metallic device having two or more surfaces is provided. In some embodiments, the device may include a first metal surface configured to have a surface energy that promotes attachment and/or growth of a first cell type and a second surface configured to have a surface energy that inhibits attachment and/or growth of a second, different cell type. In some embodiments, the device may include a first metal surface configured to have an average grain size that promotes attachment and/or growth of a first cell type and a second surface configured to have an average grain size that inhibits attachment and/or growth of a second, different cell type. For example, the implantable device may be a vascular stent having a first surface configured to promote attachment and/or growth of endothelial cells and a second surface configured to inhibit attachment and/or growth of fibroblasts.

In some embodiments, the metallic material may have an average grain size and/or surface energy that inhibits cell attachment, cell growth, or a combination thereof. For example, the metallic material may inhibit the attachment and/or growth of cells responsible for inflammation, such as immune cells.

Aspects of the present disclosure relate to a metallic material, metallic device, or appliance having a uniform average grain size nanostructure of 0.2 to 0.5 μm, wherein the metallic material, metallic device, or appliance inhibits/reduces bacterial growth and biofilm formation. In some embodiments, the metallic material, metallic device, or apparatus may be used in applications where antimicrobial activity is desired. In some embodiments, the application comprises a surgical instrument, an endoscopic device (e.g., a vertebroplasty needle). In some embodiments, the metal is type 304 stainless steel metal. In some embodiments, the metal is type 316 stainless steel metal. In some embodiments, the metal is type 304 stainless steel metal. In some embodiments, the metal is magnetized.

Aspects of the present disclosure relate to magnetized metallic materials, metallic devices or instruments having uniform average grain size nanostructures of 0.2 to 0.5 μm, wherein the metallic materials, metallic devices or instruments inhibit/reduce bacterial growth and biofilm formation and promote/increase eukaryotic cell adhesion. In some embodiments, the metallic material, metallic device or apparatus may be used in applications requiring antibacterial activity and osteointegration. In some embodiments, the applications include implantable devices and orthopedic implants. In some embodiments, the metal is type 304 stainless steel metal. In some embodiments, the metal is type 316 stainless steel metal. In some embodiments, the metallic material has a thickness of greater than 2.3 x 10⁷emu/m³Magnetic moment of (a).

Aspects of the present disclosure relate to a stainless steel metallic material or an article made of a stainless steel metallic material comprising uniform grain nanostructures and having an average grain size of 0.1 to 3 μ ι η, configured to (i) inhibit adhesion, growth, or a combination thereof of a microorganism, (ii) promote adhesion, growth, or a combination thereof of a predetermined eukaryotic cell, or (iii) inhibit adhesion, growth, or a combination thereof of a predetermined eukaryotic cell. In some embodimentsThe stainless steel metallic material comprises a uniform grain nanostructure and has an average grain size of 0.2 to 1 μm. In some embodiments, the stainless steel metallic material comprises a uniform grain nanostructure and has an average grain size of 0.2 to 0.5 μm. In some embodiments, the metallic material is type 304 stainless steel metal. In some embodiments, the metallic material is magnetized. In some embodiments, the metallic material is type 316 stainless steel metal. In some embodiments, the metallic material is magnetized. In some embodiments, the metallic material is magnetized. In some embodiments, the metallic material has a thickness of greater than 2.3 x 10⁷emu/m³Magnetic moment of (a).

Examples

Hereinafter, the present invention will be described in detail with reference to examples, but the present invention is not limited to the following examples unless it goes beyond the gist of the present invention.

Example 1: manufacture of metallic materials

To provide a metal material, stainless steel (SUS 304) was subjected to rolling treatment and thermal recrystallization to adjust the average grain sizes of the grains to 0.5 μm, 1 μm, 1.5 μm, 2 μm, 3 μm and 9 μm, respectively. The metal material had a plate shape with a length of 10mm, a width of 10mm and a thickness of 0.1 mm. The rolling treatment and the thermal recrystallization were performed according to the following procedures. Specifically, stainless steel (SUS 304) was passed through a rotary mill several times, and cold-rolled to about 40 to 65% (a reduction ratio of about 3 to 15% each time). Then, the resulting stainless steel is annealed at 600 to 850 ℃ for 10 to 100 seconds (heating rate: 200 ℃/sec) to recrystallize the stainless steel. The recrystallized stainless steel is cooled to obtain austenitic stainless steel according to the phase transformation state (cooling rate: 200 to 400 ℃/sec).

It should be understood that type 316 stainless steel and other metals may be treated the same to control the average grain size.

The metallic material obtained using the rolling treatment and thermal recrystallization described herein has a nanostructure (e.g., average grain size) that is not limited to a surface. For example, the metallic material may retain its nanostructure throughout its processing, resulting in a metallic material with a uniform nanostructure.

Example 2 measurement of average grain size

Test samples of the metal materials provided above were polished with argon ions using an ion polisher ("IM 4000", manufactured by Hitachi High-Technologies Corporation). Then, an electron microscope ("SU-70", manufactured by Hitachi High-Technologies Corporation) having a crystal orientation analysis function was used in a vacuum atmosphere (1X 10)^- ³Pa) the average grain size of the metallic material is measured at room temperature. The size of each crystal grain is determined by determining the area of each crystal grain within an arbitrary measurement range (i.e., an observed image; magnification: 1000 times) and calculating the diameter of a circle (assuming that the shape of the crystal grain is a circle having the same area as the area of the crystal grain). The area of the crystal grain and the diameter of a circle having the same area as the area of the crystal grain were calculated using an image processor ("TSL OMI Analysis 7", manufactured by TSL Solutions). Then, the sum of all crystal grain diameters in an arbitrary measurement range is divided by the number of crystal grains, and the resultant value is defined as an average crystal grain size (nm).

Example 3 antibacterial Properties of unpolished Metal Material

The method comprises the following steps:

the bacteria were first incubated overnight. At most 10⁵After the concentration of (c), the bacteria were mixed with a metal material sample (type 304 stainless steel sample) and incubated for 24 hours. The type 304 stainless steel sample was then washed with distilled water and sonicated for 10 minutes. After vortexing the samples for an additional 10 seconds, several dilutions of each sample were placed on agar plates. The agar plates were incubated for 12 hours.

As a result:

fig. 1A and 1B each show an example of a response spectrum obtained by plotting colony forming units of gram-positive bacteria or gram-negative bacteria after culture with respect to the average grain size of the grains.

As shown in fig. 1A, when the average grain sizes were 0.5 μm, 1 μm, 1.5 μm, 3 μm, and 9 μm, the number of gram-positive bacteria adsorbed on the metallic material was relatively reduced, indicating that the tested metallic material inhibited the growth/adhesion of gram-positive bacteria. Specifically, FIG. 1A shows the general reduction in the number of gram-positive bacteria on a metal material having an average grain size of 0.5 μm. As shown in fig. 1B, when the average grain sizes were 0.5 μm, 3 μm, and 9 μm, the number of bacteria adsorbed on the metal material was relatively reduced, indicating that the tested metal material inhibited the growth/adhesion of gram-negative bacteria. FIG. 1A shows the general reduction in the number of gram-negative bacteria, in particular E.coli, on a metal material having an average grain size of 0.5 μm.

Example 4 antibacterial Properties of unpolished Metal Material to polished Material

The antibacterial property of the unpolished metal material with respect to the polished material was measured.

The method comprises the following steps:

sample preparation: type 304 stainless steel as shown in table 3: diameter (phi): 11 mm; thickness: 0.1 mm.

TABLE 3

In the first step, the sample was rough polished using 3M abrasive membrane grindstone alumina.

The samples were first hand polished on five sheets of paper using a 3M abrasive film mesh number 4000(3 μ M) for about 40 seconds. The sample was then polished by hand with a 3M abrasive film mesh number 8000(1 μ M) for about 40 seconds. The sample was then hand polished with 3M abrasive film mesh No. 15000(0.3 μ M) for about 40 seconds.

In the second step, the sample was alumina polished using an alumina solution and polished on a bench using a grinder. The alumina solution used isSolutions of mixtures of alumina and 0.05 μm alumina (1 μm alumina: Buehler Micro Polish II alumina 1.0 μm; 0.05 μm alumina: Buehler MasterPrep polishing suspension 0.05 μm).

The sample was polished for 5 minutes with the first direction a, 5 minutes with the second direction B, 5 minutes with the direction a and 5 minutes with the direction B, for a total of 20 minutes, as shown in fig. 6.

In a third step, the sample was washed with: (1) firstly, using water: gently wash the sample first by hand in diluted neutral detergent and tap water, then rinse with tap water from a tap, then gently wipe the sample to dry the sample; (2) the samples were then kept with ethanol by placing them in ethanol and removing the samples and gently wiping to keep the samples dry.

Fig. 2A and 2B show that the number of bacteria adsorbed on the metal materials having average grain sizes of 0.5 μm, 1 μm, 1.5 μm, 2 μm, 3 μm, and 9 μm is relatively reduced when the metal materials are polished.

Example 5 cytotoxic MTS assay

The method comprises the following steps:

human fetal osteoblasts (HFOb) and a sample of metallic material were seeded in a 12-well plate. Cell culture medium was changed every 2 days. For cell proliferation assays, MTS reagent was added after 3, 5 and 7 days to determine the number of viable cells.

As a result:

fig. 3A and 3B show that the metallic material is not cytotoxic. For most samples, viability was 80% or higher for all three readings (3, 5 and 7 days). For day 7, all readings showed 100% or greater viability. Higher viability indicates that the metallic material sample promotes cell growth.

For the polished samples, all readings showed 90% to 120% cell viability. For day 5, all samples exhibited greater than 100% cell viability, indicating that the metallic material samples promoted cell growth. The unpolished samples exhibited greater cell viability than the polished samples. On the unpolished sample, the metallic material having an average grain size of 9 μm showed the greatest activity.

Examples 6-304 type stainless steels antibacterial Properties of colony-forming units of Escherichia coli, multidrug-resistant Escherichia coli, methicillin-resistant Staphylococcus aureus, and Staphylococcus epidermidis normalized with respect to surface area, in relation to average particle size

Method

Type 304 stainless steel materials having average grain sizes of 0.5 μm, 1.5 μm, 2 μm, 3 μm and 9 μm were used in this example.

To ensure no contamination, the samples were washed by sonication with acetone, then 70% ethanol and DI water for three 10 minutes.

The samples were then placed individually in the wells of a 24-well plate and sterilized under UV light overnight. Prior to cell seeding, the samples were rinsed twice with PBS to remove any possible debris. Then 0.5mL of the prepared bacterial suspension (10)⁶cells/mL) was added to each sample and the plate was placed at 37 ℃ and 5% CO₂In a static incubator. After 24 hours of incubation, the plates were removed from the incubator. The inoculum was aspirated from each well and replaced with 1ml of PBS. The wells of a sterile 24-well plate were filled with 1ml of PBS and the samples were transferred to the plate. The samples were washed three more times with PBS (4 total PBS rinses). The final PBS wash was not aspirated. The plates were sonicated in a cold water bath for 15 minutes.

The plate was briefly vortexed and 10. mu.l of the solution was pipetted in triplicate onto an agar plate. The stock solutions (O-dilutions) were then diluted 1:10, 1:100, 1:1000, 1:10000 and 1:100000 in PBS and each dilution was pipetted onto agar plates. Colony Forming Units (CFU) were then counted manually after incubation for about 12 to 14 hours.

In the method used in this example, the sample was washed (1+3 times) four times to remove planktonic cells, which did not attach to the sample and were free-floating. The samples were carefully washed so as not to destroy sessile bacteria. Sonication for 15 minutes allowed the bacteria to detach from the surface. Counting sessile bacteria allows for the evaluation of biofilm-forming bacteria and biofilm prevention on the surface of the sample. It is understood that bacteria can attach to the sample surface and cause an infection, and biofilm formation is a characteristic of this infection.

Results

Fig. 9 shows that for all tested bacterial strains, reducing the average grain size to 0.5 μm causes a reduction in the adhesion of the bacteria to the surface of the metal material. The bacteria tested included gram negative and gram positive bacteria, drug resistant bacteria and drug sensitive bacteria. This reduction is even more pronounced in the case of multi-drug resistant E.coli (MDR E.coli) bacteria. The growth of MDR e.coli was inhibited 4 to 49 times for an average grain size of 0.5 μm compared to other grain sizes.

Example 7 biofilm formation test Using safranin

Method

To ensure no contamination, the samples were washed by sonication with acetone, then 70% ethanol and DI water for three 10 minutes. The samples were then placed individually in the wells of a 24-well plate and sterilized under UV light overnight. Prior to cell seeding, the samples were rinsed twice with PBS to remove any possible debris. Then 0.5mL of the prepared bacterial suspension (10)⁶cells/mL) was added to each sample and the plate was placed at 37 ℃ and 5% CO₂In a static incubator. After 24 hours of incubation, the plates were removed from the incubator. The inoculum was aspirated from each well and replaced with 1ml of PBS. The wells of a sterile 24-well plate were filled with 1ml of PBS and the samples were transferred to the plate. The wells were washed three more times with PBS (4 total PBS rinses).

The samples were stained with safranin (0.1% w/v) for 5 minutes. Safranin is a cationic dye used in histology and cytology to distinguish and identify different tissues and cells. Safranin is used to stain gram positive and gram negative bacteria.

The sample was washed 3 to 4 times with 1ml of PBS (until PBS was colorless).

The safranin was redissolved with 95% ethanol. The absorbance of the redissolved safranin in each well was measured using a spectrophotometer (SpectraMax M3, Molecular Devices, Sunnyvale, Calif.), where λ_abThe wavelength of absorbance was 570 nm.

Results

Figure 13 shows that for all tested bacterial strains, reducing the average grain size to 0.5 μm causes a reduction in biofilm formation on the surface of the samples. The bacteria tested included gram negative and gram positive bacteria, drug resistant bacteria and drug sensitive bacteria.

It is understood that the biofilm formation process comprises three steps:

1-adsorption or accumulation of organisms on the collector surface (i.e., substrate or sample).

2-attachment or consolidation at the interface between the organism and the collector, typically involving the formation of polymer bridges between the organism and the collector.

3-colonization or growth and division of organisms on the collector surface. (see Garrett, T.R., Bhakoo, M.and Zhang, Z.,2008.Bacterial addition and biolofilms on surfaces. progress in Natural Science,18(9), pp.1049-1056.)

The percentage of biofilm prevention was calculated for the 0.5 μm samples compared to the other grain sizes (i.e. 1.5, 2, 3 μm):

in the case of E.coli, 21 to 34% biofilm prevention was observed compared to 1.5, 2, 3 μm average grain size samples.

In the case of MDR e.coli, 10 to 28% biofilm prevention was observed compared to 1.5, 2, 3 μm average grain size samples.

In the case of MRSA, biofilm prevention was observed at 29 to 44% compared to 1.5, 2, 3 μm average grain size samples.

In the case of staphylococcus epidermidis, biofilm prevention was observed at 30 to 41% compared to the 1.5, 2, 3 μm average grain size samples.

Example 8 Effect of grain size on fibroblast growth

The research on the metal material to human skin fibroblast (HDF) ((HDF))CCL-110^TM) The cytotoxicity of (a).

Method

First, human skin fibroblasts were treated with 5% CO at 37 ℃₂In a moist incubator (Tagla's minimum Essential (EMEM) medium containing 10% fetal bovine serum and 1% penicillin streptomycin).

Then, the cells and the wire sample were seeded at 5,000 cells/well in a 48-well plate at 1000. mu.L of cell cultureIn nutrient medium and 5% CO at 37 deg.C₂Incubate in a humid atmosphere for 3, 5, and 7 days.

After a period of incubation, the medium was removed and replaced with 1000 μ L of fresh medium in which the MTS solution was diluted 1:5 (200 μ L +1000 μ L EMEM). This time, the well plates were incubated for only 3 hours to allow for a color change. The absorbance was measured at 490nm using an absorbance microplate reader (SpectraMax). Data are expressed as a percentage of cell viability.

The metal samples tested were type 316 stainless steel wire and type 304 stainless steel wire with different diameters and different average grain sizes as shown below:

type 304 stainless steel sample: the metal samples tested were of different diameters (In mm) and different average grain sizes (in μm) of type 304 stainless steel wire.

CG304Grain size 21.5 μm

CG304Grain size 12.0 μm

CG304Grain size 15.0 μm

UFGSS 304Grain size 0.27 μm

UFGSS 304Grain size 0.22 μm

UFGSS 304Grain size 0.23μm

The chemical composition of the type 304 stainless steel metal samples used is shown in the following table:

TABLE 4

Type 316 stainless steel sample: the metal samples tested were of different diameters (In mm) and different average grain sizes (in μm) of type 316 stainless steel wire.

CG316Grain size 16.5 μm

CG316Grain size 10.7 μm

CG316Grain size 7.1 μm

UFGSS 316Grain size 0.25 μm

UFGSS 316Grain size 0.22 μm

UFGSS 316Grain size 0.18 μm

The chemical composition of the type 316 stainless steel metal samples used is shown in the following table:

TABLE 5

The diameter and sample area of the samples tested are shown in the following table:

TABLE 6

Sample diameter (mm)	0.2	0.4	0.8
				Area of sample (mm)²)	8.04	16.21	32.92

Figures 4A-4C show the percent viability of human skin fibroblasts when grown on type 304 stainless steel metal samples. Figure 4C shows that ultra-fine type 304 stainless steel metal samples with average grain sizes of 0.22 μm and 0.27 μm promote human skin fibroblasts.

Figures 5A-5C show the percent viability of human skin fibroblasts when grown on type 316 stainless steel metal samples. Fig. 5C shows that conventional metal samples with an average grain size of 16.5 μm promote human skin fibroblasts.

Example 9: effect of type 304 stainless Steel magnetic polarization and grain size on bacterial colony formation

Method

The susceptibility of The samples was studied using a vibrating sample magnetometer (VSM, The LakeShore 7407VSM) (FIG. 8)). The sample volume used is equal to 6.2mm³And is the same at all grain sizes tested.

The magnetic susceptibility of type 304 stainless steel and type 316 stainless steel with various average grain sizes (0.5, 1.5, 2, 3, 9, 10 μm) was measured at room temperature using a vibrating sample magnetometer. The samples were mounted on a sample holder and the magnetic field was-1000 to +1000 Oersted (Oe) to measure their magnetic susceptibility (n.gtoreq.2).

Type 304 stainless steel with different average grain sizes (0.5, 1 and 9 μm) was used in this example.

The samples were magnetized with 0.1 tesla magnet (0.1T), 0.5 tesla magnet (0.5T) and 1.1 tesla magnet (1.1T).

Results

Fig. 10 shows that controlling the magnetic moments (0.1T, 0.8T, 1.1T) of the type 304 stainless steel samples reduces the adhesion of bacteria to the samples compared to Untreated (UT) or Demagnetized (DM) type 304 stainless steel samples. Magnetized samples showed a more significant reduction in bacterial adhesion. Statistical analysis indicates the difference from untreated samples (UT).

Type 304 stainless steel samples with average grain sizes of 0.5 μm and 1 μm showed more reduction in bacterial adhesion/biofilm formation due to their highest magnetic susceptibility compared to samples with 9 μm grains.

Table 7 shows the magnetic moments at 0.1T

304 grain size (mum)	Magnetic moment at 0.1T (memu)
		0.5	144.98±8.90
1.5	6.75±0.070
		2	5.85±0.070
3	5.25±0.70
		9	4.6±0.01

Table 7 shows that the reduction in average grain size increases the magnetic susceptibility of the type 304 stainless steel sample, and that the type 304 stainless steel sample with an average grain size of 0.5 μm exhibits the highest magnetic susceptibility, above 144.98 ± 8.90 memu.

Example 10 bone growth

Method

Each sample was seeded with 26000 individual fetal osteoblasts (. apprxeq.2X 10) in tissue culture treated 24-well plates with 500. mu.L of the appropriate medium containing 10% FBS and 1% penicillin-streptomycin⁴Individual cell cm^-2). The plates were allowed to stand at 37 ℃ under static conditions and enriched in 5% CO₂For 3, 24 and 72 hours. After the incubation period, the metabolic activity of the cells adhered to the surface of each sample was quantified. Briefly, the above medium was carefully aspirated and replaced with the appropriate medium containing 20% MTS reagent. The cells were mixed with MTS in the dark at 37 ℃ and 5% CO₂Incubate for 3 hours. After incubation, the absorbance of each well was measured using a spectrophotometer (SpectraMax M3, Molecular Devices, Sunnyvale, Calif.), where λ_abThe wavelength of absorbance was set to 490 nm. The obtained OD data was subjected to graphical analysis and statistical analysis. Type 304 stainless steels with different average grain sizes (0.5, 1 and 9 μm) were used in this test.

Results

Fig. 11 shows that after 3 hours post-incubation, a higher number of osteoblasts adhered to the magnetized 304 stainless steel sample. Further, fig. 11 shows that magnetization of type 304 stainless steel samples with average grain sizes of 0.5 μm and 1 μm using a 1.1T magnet increased cell adhesion, while magnetization of type 304 stainless steel samples with average grain sizes of 9 μm did not affect cell adhesion.

Fig. 12A to 12C show that all magnetized type 304 stainless steel samples tested with an average grain size of 0.5 μm (fig. 12A), 1 μm (fig. 12B) or 9 μm (fig. 12C) support cell adhesion and growth for up to 72 hours after incubation. In addition, fig. 12A to 12C show that growth was lowest on the Demagnetized (DM) samples 72 hours after incubation.

Example 11: relationship between antibacterial property of 316 type stainless steel to bacterial colony forming unit and average grain size, magnetic property and surface energy

Method

Methicillin-resistant staphylococcus aureus (MRSA) was first incubated overnight. At most 10⁵After the concentration of (a), the bacteria were mixed with the metal material sample and incubated for 24 hours. The samples were then washed with distilled water and sonicated for 10 minutes. After vortexing the samples for an additional 10 seconds, several dilutions of each sample were placed on agar plates. The agar plates were incubated for 12 hours.

Material

The chemical composition of the type 316 stainless steel metal samples used is shown below.

TABLE 9

The metal samples tested were of diameterAnd type 316 stainless steel wires with different average grain sizes: an ultra-fine material having an average grain size of 0.22 μm and a conventional material having an average grain size of 10.7 μm.

The samples were placed on a sample holder of a vibrating sample magnetometer (7400-S Series VSN, Lake Shore). Readings were set at magnetic fields of 5000Oe to-5000 Oe.

Results

The following shows the response amounts (CFU/cm) with respect to the average grain size, magnetic properties and surface energy of the grains²) The result of (1).

And an average grain size of 10.7 μm (520 CFU/cm)²) When the average grain size is 0.22 μm (6 CFU/cm)²) CFU/cm of MRSA on type 316 stainless steel wire²And decreases. The surface energy of the sample was reduced when the average grain size was 0.22 μm (37.39mN/m) as compared with when the average grain size was 10.7 μm (332.14 mN/m). Without being bound by theory, the adhesion of MRSA to type 316L stainless steel metallic materials decreases when the surface energy of the metallic sample approaches 42 mN/m. An increase in magnetic moment by work hardening was observed on type 316 stainless steel metallic material wire with an average grain size of 0.22 μm compared to conventional type 316 stainless steel wire with an average grain size of 10.7 μm.

Examples 12-316 stainless steels having antibacterial properties against bacterial colony forming units as a function of average grain size, magnetic properties and surface energy

Method

Pseudomonas aeruginosa was first incubated overnight. At most 10⁵After the concentration of (a), the bacteria were mixed with the metal material sample and incubated for 24 hours. The samples were then washed with distilled water and sonicated for 10 minutes. After vortexing the samples for an additional 10 seconds, several dilutions of each sample were placed on agar plates. The agar plates were incubated for 12 hours.

Material

The chemical composition of the type 316 stainless steel metal samples used is shown in the table below.

Watch 10

The metal samples tested were of diameterAnd type 316 stainless steel wires with different grain sizes: ultra-fine material samples with an average grain size of 0.25 μm and conventional material samples with an average grain size of 16.5 μm. The sample was placed on a sample holder of a vibrating sample magnetometer (7400-SSeries VSN, Lake Shore). Readings were set at magnetic fields of 5000Oe to-5000 Oe.

Results

The following shows the response amounts (CFU/cm) with respect to the average grain size, magnetic properties and surface energy of the grains²) The result of (1).

And an average grain size of 16.5 μm (73 CFU/cm)²) When the average grain size is 0.25 μm (24 CFU/cm)²) In time, CFU/cm of Pseudomonas aeruginosa on type 316 stainless steel wire²And decreases. The surface energy of the type 316 stainless steel metallic material wire was reduced when the average grain size was 0.25 μm (1.87mN/m) as compared with that when the average grain size was 16.5 μm (971 mN/m). Without being bound by theory, the adhesion of pseudomonas aeruginosa to type 316L stainless steel metal materials is reduced when the surface energy of the metal sample approaches about 42 mN/m. An increase in magnetic moment by work hardening was observed on type 316 stainless steel metallic material wire with an average grain size of 0.25 μm compared to conventional type 316 stainless steel wire with an average grain size of 16.5 μm.

Examples 13-304 stainless steels antimicrobial Properties on bacterial colony Forming units versus average grain size, magnetic Properties and surface energy

Method

MRSA was first incubated overnight. At most 10⁵After the concentration of (a), the bacteria were mixed with the metal material sample and incubated for 24 hours. The samples were then washed with distilled water and sonicated for 10 minutes. After vortexing the samples for an additional 10 seconds, several dilutions of each sample were placed on agar plates. The agar plates were incubated for 12 hours.

Material

The chemical composition of the type 304 stainless steel metal samples used is shown in the table below.

TABLE 11

The metal samples tested were of diameterAnd type 304 stainless steel wire with different grain sizes: ultra-fine material samples having an average grain size of 0.27 μm and conventional material samples having an average grain size of 21.5 μm.

The samples were placed on a sample holder of a vibrating sample magnetometer (7400-S Series VSN, Lake Shore). Readings were set at magnetic fields of 5000Oe to-5000 Oe.

Results

The response amounts (CFU/cm) with respect to the average grain size, magnetic properties and surface energy of the grains are shown in the following table²) The result of (1).

And when the average grain size is 21.5 μm (82 CFU/cm)²) When the average grain size is 0.27 μm (23 CFU/cm)²) CFU/cm of MRSA on type 304 stainless steel wire²And decreases. The surface energy of the type 304 stainless steel metal material wire was 324.27mN/m when the average grain size was 0.27 μm, compared to 15.08mN/m when the average grain size was 21.5 μm. An increase in magnetic moment by work hardening was also observed on type 304 stainless steel metallic material wire with an average grain size of 0.27 μm compared to conventional type 304 stainless steel wire with an average grain size of 21.5 μm. Without being bound by theory, for type 304 stainless steel metal, the surface energy effect weakens as the magnetic moment increases.

Example 14: osteoblast growth data using titanium alloys

Cytotoxicity Studies of human fetal osteoblasts (hFOB) (CRL-11372, ATCC)

Human fetal osteoblasts containing 5% CO at 37 deg.C₂In a humidified incubator (Takara Shuzo) was cultured alone in complete medium (Dulbecco's Modified Eagle Medium (DMEM/F12) containing 10% fetal bovine serum and 1% penicillin streptomycin). Then, the cells and wire samples were seeded at 5,000 cells/well in 1000. mu.L of cell culture medium in 48-well plates and at 37 ℃ in 5% CO₂Incubate in a humid atmosphere for 3, 5, and 7 days. After a period of incubation, the medium was removed and replaced with 1000. mu.L of fresh medium, in which Presoblue solution (100. mu.L + 900. mu.L DEMEM/F12) was diluted 1: 10. This time, the well plates were incubated for 45 minutes to allow for a color change. Fluorescence was measured with a microplate reader (SpectraMax) at an excitation wavelength of 560nm and an emission wavelength of 590 nm. Data are expressed as a percentage of cell viability.

Table 12 shows the titanium alloy samples used.

TABLE 12

(x) unknown grain size

Table 13 shows the composition of the titanium alloy control sample

Watch 13

ASTM F136 titanium

Cell growth assays using human fetal osteoblasts (hFOB).

For all samples tested, the percentage of viability was higher than 80% during the 7 day incubation period, indicating relatively low cytotoxicity to hFOB cells (see fig. 7).

Furthermore, the percentage value of viability was higher than 100% in most cases, indicating that the number of cells grown in the presence of the sample was higher compared to the number of cells grown in contact with fresh medium only. Without being bound by theory, cells grow on top of the titanium samples, which promotes their proliferation.

After day 5, the percentage of cell viability was slightly reduced for samples with grain sizes of 3.1 μm, 2 μm and 0.8 μm.

Although the disclosure has been described herein with reference to particular embodiments, it is to be understood that these embodiments are merely illustrative of the principles and applications of the present disclosure. It is therefore to be understood that numerous modifications may be made to the illustrative embodiments and that other arrangements may be devised without departing from the spirit and scope of the present disclosure as defined by the appended claims.

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