Diamond Like Carbon Films Kind Emerging Antibacterial Biomaterial Biology Essay

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Abstract

Diamond-like carbon (DLC) films is a kind of emerging antibacterial biomaterial used for artificial implant to the human body. However, because the antibacterial property of DLC is not good enough, scientists try to improve the antibacterial property of the DLC. And the amorphous nature of DLC opens the possibility of introducing additional elements, such as Si, F, N, O and their combinations into the biomaterial [1]. Scientists always try to plate ion-implanted DLC on the surface of stainless steel as coating, and then test the antibacterial capability. This review concludes the research results of ion-implant DLC for this decade and the comparison between DLC and PTFE. It includes not only the empirical methods but also the results of the experiments.

Key words: biomaterial, ion-implant, DLC films, molecular surface energy, antibacterial

3. Introduction

With the development of biomaterial used in the human body, new questions just came out constantly. The most important question calls our attentions is the corrosion and infection of the biomaterial implanted in the human body such as the Cardiac valve prosthesis, Bone plates implanted into bone when it is broken and Artificial ligaments and tendons. These artificial implant biomaterials help people come back to normal life, however, corrosion and infection can't be treated by drugs as the implanted biomaterial is protected by Phagocytosis and antibiotics [2].At the same time the implanted biomaterial can't be easily taken out from the human body when it is corrosion or it won't work any longer, because the surgery will increase the patients suffering or even cause mortal injury. So the best solution is choosing the biomaterial which is antibacterial and can be used for longer time.

Then the question comes to how to make biomaterial be antibacterial .The substantial reason for the corrosion and the infection is bacteria adhesion. The substantial solution is to alter the material's surface properties and make it less attractive for the formation of micro-organisms. For the last 50 years biomaterial called Diamond like carbon (DLC) sparking scientist's interest due to its great properties as biomaterial. The thermal conductivity of DLC is similar to metal. It is very unyielding, durable and non-corrodible. It also gets very low friction and smooth surface [3].What's more the amorphous nature of DLC opens the possibility of introducing additional elements, such as Si, F, N, O and their combinations into the coating whilst still maintaining the amorphous phase of the coating [1].All its properties make it become a very important and popular biomaterial for artificial implants into human body.

Scientists always try to plate Ion-implanted DLC on the surface of stainless steel, and then test the antibacterial capability. The ion implanted included N+, O+, F+, Si+, Si and so on. As the molecular surface energy can reflect the antibacterial property of biomaterial, when the molecular surface energy is high, it is easier to absorb bacterial .So we could get antibacterial property through calculating molecular surface energy.

4. Theoretical consideration

4.1 The definition of biomaterial:

Biomaterial is a kind of biocompatible material which is used for diagnosis, repair, function improvement of human tissue and organs. It is always used to construct artificial organs or prostheses and replace natural body tissues such as valves、stents、joints 、soft tissues and so on.

4.2 Biomaterial including:

There are varieties species of biomaterials, and they are widely used. Biomaterials include three categories, metal materials (such as alkali metals and their alloys, etc.), inorganic materials (bioactive ceramic, hydroxyapatite, etc.) and organic material.

Organic materials are mainly a collection of high molecular polymer materials, high polymer materials usually divided into synthetic polymer material and natural polymer materials .Synthetic polymer material contains polyurethane, polyester, polylactic acid, polyglycolic acid, lactic acid glycolic acid and other synthetic plastics and rubber which is for medical use).

Natural polymer materials is about collagen, silk protein, cellulose, chitosan and so on;

According to the use of materials, these materials can be divided into bioinert materials, bioactive materials and biodegradable materials.

Depending on whether the degradation products of the polymers can be metabolized and absorbed by the body, bio-degradable polymer can be divided into non-absorbable and bio-absorption. According to the affection of material to blood components and performance ,when exposure to the blood, material can be divided into blood compatibility of polymer material and blood incompatibility material. According to the compatibility and reflect of material to the body, the material can be divided into biocompatible and bio-compatible polymer and so on.

The Diamond-like carbon is a kind of synthetic polymer material and bioinert material which belongs to organic materials. It is well biocompatible to the human body [4].

4.3 The application of biomaterial:

Biomaterials are mainly used in four aspects.

1. The function of afford or pass the load. Such as artificial bones, joints teeth, ligaments and tendons, etc.

2. The function of control the flow of blood or body fluids. Such as artificial valves, blood vessels, etc.

3.The function for electricity, light and sound conduction. Such as cardiac pacemakers, intraocular lens Cochlear replacements, etc.

4.The function of filling or beautify the human body. Such as Skin repair devices (artificial tissue), Breast implants, etc [5]

4.4 Features of biomaterial:

As biomaterials are mainly used in the human body, four characteristics are required .

(1) Biological functions, varies according to different biomaterials.

(2) Biocompatibility, the relationship between Materials and the human body, including blood compatibility and tissue compatibility (non-toxic, non carcinogenic, non-pyrogenic reaction, no immune rejection, etc.).

(3) Chemical stability, resistance to biological aging (particularly stable) or biodegradable (controllable degradation).

(4) Processibility.It could be molded, disinfected (UV sterilization, high-pressure boiling, ethylene oxide gas sterilization,, and other alcohol disinfection) [6].

4.5Biocompatibility

4.51 The response of organism to biological materials - host reaction

It contains biological response and changes in response to biological organisms

biological response

Blood reaction

1, platelet thrombosis;

2, the coagulation system activation;

3, fibrinolytic system activation;

4, hemolytic reaction;

5, leukocyte reaction;

6, cytokine response;

7, protein adhesion;

immune response

1, the complement activation;

2, the humoral immune response (antigen -ibody reaction);

3, the cellular immune response.

tissue reaction

1, the inflammatory response;

2, cell adhesion

3, cell proliferation (abnormal differentiation)

4, the formation of film Rang

5, changes in the cytoplasm

changes in response to biological organism

Acute systemic reaction

Allergy, toxicity, hemolysis, fever, nerve palsy

Chronic systemic reaction

Toxicity, teratogenicity, immune, dysfunction, etc

Acute local reactions

Inflammation, thrombosis, necrosis, rejection, etc

Chronic local response

Cancer, calcification, inflammation, ulcers, etc.

4.52. Materials response in human body

Materials response in human body could lead to the damage in the nature structural of material and the lost of its function. It is mainly divided into the following three aspects: Metal corrosion, Polymer degradation and Attrition [7].

4.521 Metal corrosion

Corrosive environment in vivo:

(1) saline solution is an excellent electrolyte for the electrochemical corrosion and hydrolysis;

(2) the existence of molecules and cells which is of the ability to catalyze or rapid destruct allothigenous constituents will produce bio-corrosion of metallic materials.

In terms of biological materials, it is mostly localized corrosion, specifically including stress corrosion cracking, pitting corrosion, intergranular corrosion, corrosion fatigue and corrosion cracks, resulting in overall damage to biological materials.

Although metallic materials remain inert in vivo, but there may still be some substance dissolved in biological tissues, and produce toxic reactions in organizations, resulting in tissue damage. Such as stainless steel when dissolved of Cr +6 in the biological tissue may cause toxicity [8].

4.522 Degradation of polymer

Polymers when used in long-term, are affected by oxygen, heat, ultraviolet light, mechanical, water vapor, acid-base and microbial factors, and are gradually loses its elasticity. Cracks, harden appear, it become brittle or soft, sticky, discoloration, etc. So that physical and mechanical properties is getting worse. Polymer fragments, low molecular weight monomer material are very easy to form when polymer is aging, so people need to be very careful when using it. For durable device, a certain strength and other mechanical properties must be maintained. Aging products can have toxic effects on the surrounding tissue [9].

For example, surgical suture degradation will produce acidic substances when it is of degradation, if there is a little catabolite, it is easy to be neutralized by chemical substances in the body and at the same time too many aging products will cause harm to surrounding tissue.

4.523 Wearing

Ti6Al4V is commonly used in artificial joint materials, due to it is easily to oxidized TiO2 at the surface, its poor wear resistance. When implanted in the body, dark brown viscous material formed surrounding the joints which is caused by the wear and tear of the joint, causing pain. Life expectancy of Titanium artificial hip joint is generally less than 10 years.

Currently, a large number of artificial hip joint is formed by a hard metal or ceramic femoral head and ultra high molecular polyethylene acetabular cup. But it's life is no more than 25 years. Long-term data showed that the main reason for implant failure is osteolysis caused by high molecular polyethylene wear particle the interface, leading to prosthesis loosening. The wear and tear caused by foreign particles - giant cell reaction, also known as particle disease, is the main reason for late failure [10].

4.6Parameters used to measure the property of the biomaterial

Parameters as follow are used to measure the property of the biomaterial [11]

Biocompetibility

Inflammatory reaction

Compressive stress

Thermal stability

Mechanical properties(Low friction coefficient)

Antibacterial

Durable

Surface chemical composition

9. Surface topography

10. Surface roughness

11. Surface free energy

5.Result

5.1 bacterial culture

5.11 Choice of bacteria and the Germiculture

Bacteria used in the bacteria adhesions experiment always are Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa and Gram-nagative rod as they are the most common bacteria forming biofilms on the surface of medical device and frequently causing device-associated infections[12].

S. epidermidis cause polyer-associated infection (prosthetic or valve endocarditis) However, Gram-nagative rod is also be involved in the experiment as it always cause the contact lenses-associated keratitis. These bacteria survive not only in normal atmospheres, but also in hypoxic atmospheres. So the destructibility to inner-implanted biomaterials in man-made environments is immeasurable. So the use of these bacteria in study of antibacterial properties is necessary.

S. epidermidis (F1661); S. aureus (F1557) and P. aeruginosa(F1692) were obtained from Institute of Infection and Immunity, Nottingham University, UK.. The bacteria were subcultured and preserved in 15% glycerol in TSB (Tryptone Soya Broth, Oxoid1, UK) as frozen stock at -80。C. For all adhesion tests, TSA (Tryptone Soya Agar) plates were streaked out with a loop from the frozen stock and grown overnight at 37。C. A single colony was inoculated in 20 ml TSB and grown statically overnight at 37 。C. Five hundred microlitres from this culture were further inoculated into 100 ml TSB in a conical flask and grown in a shaker-incubator at 37 。C and 250 rpm.

The culture was grown to mid-exponential phase since cell harvested in this state showed a better adhesion to solid surface[9]. The bacteria were harvested by centrifugation at 4500 rpm for 5 min at -4 。C, washed once in sterile distilled water and resuspended in the media (pre-warmed to 37 。C) at a CFU/ml concentration[13,14,15].

5.12 The preparation of Ion implantation experiment

Ion implantation leads atoms into the layer of the surface of a solid substrate by bombarding the solid with ions which energy ranging from the keV to MeV. In most cases scientists choose the stainless steel 316 as the substrate. During ion implantation a beam of dopant ions with fixed energy is swept across the target surface. The ions have a sufficiently high velocity, about cm/s, as a result they penetrate the surface and rest at a depth of 10-1000 nm, however, the accurate depth is depending on their energy and mass, and on the mass of the atoms of the substrate material. When an energetic ion penetrates the substrate material, it will endure a series of collisions with the target nuclei and

Electron, lose its energy and come to rest at last [16].

According to the experiment of Q. Zhao, the ion-implanted diamond-like carbon was prepared on stainless steel 316 discs with diameter 10 mm . The stainless steel 316 discs were cleaned in an ultrasonic bath containing acetone for 10 min, rinsed with distilled water and dried before being put into coating chamber. The substrates were further cleaned, prior to deposition, by Ar+ bombardment.

And ion contents in the DLC films were altered by changing flow rate.

5.13Static adhesion method

According to Zhao et al .The samples were put into a aseptic beaker with 100 ml bacterial suspension ( CFU/ml) and incubated at 37.C for 1 hour under a soft stirring at 20 rpm in a shaker incubator (Stuart Scientific,UK). Each sample was taken out from the suspension using aseptic forceps and the sample was moved down-up vertically in a glass tank with aseptic distilled water at 37 .C under a constant shear stress of 0.014 N to remove loosely bound bacteria and then transferred to a aseptic glass beaker with pre-warmed aseptic distilled water. The bacteria adhered to the surface was removed completely from the surface by sonication for 10 min. Hundred microlitres of the sonication suspension and , and dilutions by sterile distilled water were plated out on

TSA plates (two plates for each concentration) and incubated overnight at 37.C. The colonies were counted on the following day. The colonies number was converted to CFU on the sample surface. In order to investigate the influence of the contact time on the amount of adherent bacteria, bacterial adhesion was allowed to occur for 1 h, 5 h, 18 h and 24 h in phosphate buffered saline (PBS) solution. In order to investigate the influence of media on the amount of adherent bacteria, three other types of media (sterile distilled water, 10% tryptic soy broth and 30% tryptic soy broth) were also used. Then the relationship of the amount of adherent bacteria on modified surfaces using different media was obtained. The experiments were carried out in triplicate (i.e. three samples of each type) and repeated three times in order to confirm reproducibility [17].

5.14 Flow chamber method

Bacterial adhesion under dynamic condition was studied using a flow chamber. The cover is made of glass plate and the flow chamber is made of PTFE plate. The width of the flow chamber is 50 mm, the depth 8 mm and the length 133 mm. Bacterial suspension flowed through the system at 37.C. In this study, a constant flow rate 0.12 ml 1 was used, which yields a laminar flow (Reynold number of

6.3). The suspension was recirculated using a peristaltic roller pump. Effects of contact time (1 h, 5 h, 18 h and 24 h), bacterial types (S. epidermidis, S. aureus and P. aeruginosa) and ionimplanted elements (N+, O+ and SiF3+) on bacterial retentionwere investigated using this device [17].

5.2 experiment of bacterial adhesion to the ion-implant DLC for this decade

For all these experiment, Atomic force microscopy is used to determine the surface roughness and topography. However X-ray photoelectron spectroscopy is used to determine the surface chemical composition of the films [18].

The thickness and the compositions of the coatings were measured using a digital micrometer and an energy dispersive X-ray microanalysis (EDX), respectively.

The incorporation of PTFE particles into Ni-P or Ni-Cu-P matrixes by gradually increasing the PTFE content from the substrate to the top surface improved the corrosion-resistant properties of the coatings significantly [17,18].

Implant

Ion

Bacteria used for the adhesion test

Result

parameter

Method to get ion-implanted DLC

and the ion contents in the DLC films

Si

Staphylococcus aureus

bacterial adhesion decreased with the increasing of silicon content or with increasing sp3/sp2 ratio

Thickness of diamond-like carbon films is 1 μm

magnetron sputtering technique

Tramethylsilane changing

tetramethylsilane flow rate.

Si+,

N+

O+

Staphylococcus epidermidis and

Staphylococcus aureus

SiF3+ ion-implanted stainless steel surface performed much better than N+ ion-implanted steel, O+ ion-implanted steel and stainless steel 316L in inhibiting

bacterial attachment under static conditions or laminar flow conditions.

energy of ion implantation for N+, O+ and Si+ were ions and 30 keV

An ion implantation technique

Ag-PTFE nanocom-posite coating

Ag-PTFE nanocomposite coating

Ag-DLC films were prepared via PECVD technique. These films

demonstrated good quality (ID/IG ratio) when compared to the pure DLC films, with just a slightly reduction in hardness. Silver do not chemically bind with carbon, but enhance its properties (stress reduction and antibacterial activity). Ag-DLC performed antibacterial activity against E. coli

DC-pulsed PECVD supply

F-DLC

Pseudomonas fluorescens

The experimental results showed that the incorporation of fluorine into the DLC coatings reduced bacterial attachment and increased bacterial removal. The F-DLC coatings with higher F content (39.2 at.%) reduced bacterial attachment by 48.8% and increased removal by 90.2%,compared with a standard DLC coating.

RF PECVD with a mixture of argon(Ar), methane

(CH4) and tetrafluor-omethane

(CF4) gases and a graphite target

Change the content of CH4 and CF4

Ni-P, Ni-P-PTFE, Ni-Cu-P Ni-Cu-P-PTFE

thermophilic streptococci

The attachment of

Thermophilic streptococci on graded electroless Ni-P-PTFE coatings could be reduced by 82-97%.

The incorporation of

PTFE particles into Ni-P or Ni-Cu-P matrixes did not improve the corrosion-resistant

Properties. The incorporation of copper into Ni-P or Ni-P-PTFE matrixes also improved the corrosion-resistant properties of the coatings.

Figure 1 The comparison of parameter result of the experiment [19, 20, 21, 22, 23, 24,25]

As for the bacterial, S. epidermidis and S. aureus were more prone to attach to a surface, compared with P. aeruginosa.

6. Discussion and future prospects

In this review, the antibacterial capability of Ion-implanted DLC on the surface of stainless steel is obviously better than the none-implanted DLC. Through calculating the free surface energy of all the materials and the experiment, in all ions-implanted DLC, who gets the lowest free surface energy owns the best antibacterial capability .

In this review, the antibacterial of ion-implanted DLC is mainly discussed. However, the antibacterial is mainly focus on the artificial bones, joint prosthesis, teeth and so on, which is very hard and not easy to be deformed.

In the future, maybe more attentions should be focus on artificial valves, blood vessels ligaments and tendons and so on, which is soft and hard to control its movement.As for artificial valves or blood vessels which must contact with the blood, a lot of new parameters should be added to the experiment. Thus, the research into effects to adhesion properties of Platelet must be included.

7. Conclusion

Ion-implanted DLC is a kind of new developed biomaterial which is very promising. The research into the property of every ion-implant DLC is very necessary. We can not only find out the antibacterial properties of every different ion-implant DLC and choose the best one through comparison, but also found out many different kinds of ion-implant DLC for different use. In the future we should focus attention on ion-implant DLC used for the replacement of soft tissue, such as artificial valves, blood vessels.

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