1

Study of two new DLC coatings for joint prosthesis: effect of the presence of albumin on the tribological

behavior

A. P. Carapeto a, A. P. Serro a,b, R. Colaço c, B. Saramago a

a Centro de Química Estrutural, Complexo I, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal b Instituto Superior de Ciências da Saúde Egas Moniz, Quinta da Granja, Monte da Caparica, 2829-511 Caparica, Portugal c Departamento de Engenharia de Materiais, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal Introduction

It is well recognized that the most common cause of failure of total hip and knee prostheses is the

formation of debris from the counterfaces of ultra high molecular weight polyethylene (UHMWPE).

These particles can lead to the inflammation of the surrounding tissues and give rise to bone

resorption (osteolyse) and loosening of the prosthesis. In the last years, there have been many

attempts to coat orthopaedic materials with hard coatings to decrease their wear rate. Diamond-like

carbon (DLC) has emerged as a potential material for biomedical applications, due to its excellent

tribological and mechanical properties, corrosion resistance, and biocompatibility.

In this work, the tribological behaviour of two new DLC’s coatings (N3FC and a-C) against

UHMWPE is studied. We proceeded to the characterization of coatings by various techniques and

investigated the adsorption of albumin (the most abundant protein in biological fluids) to these

surfaces. The tribological performance of these coatings against UHMWPE was evaluated in the

presence and absence of this protein, and the results were compared with those obtained with

uncoated stainless steel surfaces (SS).

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Experimental

Materials The following materials have been used: ultra high molecular weight polyethylene (UHMWPE)

semi-spheres, AISI 316L austenitic stainless steel (SS) squares cut from 1 mm sheet and the DLC’s a-

C and N3FC coatings, prepared in Argonne National Laboratory (Illinois, USA) by PECVD, which were

deposited both on SS surfaces and on gold quartz crystals.

Hank’s Balanced Salt Solution (HBSS, Sigma, Ref H8264) and bovine serum albumin (BSA, Serva,

Ref 11930) were used to simulate the biological fluids.

Methods

Surface characterization

Optical microscopy and AFM

The Atomic Force Microscopy images were performed with a Veeco™ DI CP-II AFM in contact

mode. The optical microscope of this equipment was also used to observe the surfaces.

SEM

Surfaces were analyzed by scanning electron microscopy (SEM) with an Hitashi S2400 equipment.

X-Ray Diffraction

The x-ray spectra were obtained with a Bruker AXS-KAPPA APEX II, X-Ray Single Crystal

Diffractometer, Mo radiation, at 40 kV and 30 mA, with 2θ=4-100º and acquisition time 18h40m.

Raman Spectroscopy

Raman spectra were obtained with 413 nm excitation line, 11 mW laser power , 20s accumulation

time. Each spectrum is an average of 4 spectra taken from different spots on the sample. All Raman

measurements were performed with a confocal microscope coupled to a Raman spectrometer (Jobin

Yvon U1000) equipped with 1200 l/mm grating and liquid-nitrogen-cooled back-illuminated CCD

detector.

Ultramicroindentation tests

The ultramicrohardness of the coatings was determined using a Shimadzu duh-211S displacement

sensing ultramicroindentation apparatus, with a Berkovich indenter. Samples were tested with a

normal load of 10mN in cycles with a 10 s loading step to maximum load, followed by a 20 s plateau

step at maximum load and a final 10 s unloading step.

Contact Angle measurements

The measurement of the static contact angles of water and protein solution (BSA+HBSS, 4mg/mL)

was carried out through the sessile drop method. Microdrops were generated with a micrometric

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syringe and deposited on the substrate surface inside a chamber previously saturated with water. A

sequence of images, obtained with a videocamera (JAI CV-A50) mounted on a microscope (Wild

M3Z) and connected to a frame grabber (Data Translation model DT3155), was recorded during 1800

s, starting from the moment of the drop deposition, enabling the monitoring of the evolution of the

angle during this period. Image acquisition and analysis was performed using the ADSA-P software

(Axisymmetric Drop Shape Analysis-Profile).

Protein Adsorption

Quartz Cristal Microbalance with Dissipation (QCM-D)

The QCM-D (KSV Instruments Ltd. Finland, model QCM-Z500) was used to determine BSA

adsorption on the surface of quartz crystals coated with the N3FC DLC. The variation of frequency (Δf)

for the fundamental, third, fifth, seventh and ninth harmonics was monitored as a function of time,

during the sequential addition of HBSS (baseline), the solutions of BSA and HBSS (rinsing), to the

quartz crystals. All the experiments were carried out at 25ºC.

Ellipsometry

Measurements were performed using an Imaging Ellipsometer, model EP,3 from Nanofilm Surface

Analysis. This ellipsometer was operated in a polarizer-compensator-sample-analyzer (PCSA) mode

(null ellipsometry). The light source was a solid-state laser with a wavelength of 532 nm. Experiments

were carried out at 25ºC following the same steps of solution introduction as referred in QCM-D.

Tribological Tests

The friction coefficient of the samples was measured in a CSM Instruments linear nanotribometer.

Experiments were performed in lubricated conditions, using HBSS and BSA+HBSS solutions, with

normal forces 25mN and 1mN, velocity 2.5 cm/s and 0.5 cm/s, in a sliding length of 2 mm, at room

temperature.

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Results and Discussion

Coating of the SS substrates with the DLC’s led to a decrease of the average roughness of the

surface determined by AFM (Table 1). However, optical microscope images allow to observe that the

surfaces with DLC are more heterogeneous, presenting small holes in the coating (Figure 2).

Fig. 1 - AFM images of (a) SS, (b) N3FC and (c) a-C.

Table 1 - Average Roughness for the three samples tested: SS, N3FC and a-C.

Ra (nm)

SS 24±6

N3FC 17±1

a-C 15±5

Fig. 2 - Optical microscope images (150x), (a) SS, (b) N3F, (c) a-C.

Using X-Ray diffraction (Figure 3) and Raman spectroscopy (Figure 4) it was found that both DLC's

are essentially amorphous, containing mainly sp3 bonds, which explain its high ultramicrohardness

(Table 2). However, the N3FC DLC has a higher content of sp2 bonds, which leads to a greater degree

of crystallinity, and consequently to an ultramicrohardness slightly lower, according with what was

determined experimentally. Deconvolution of the Raman spectra allowed to calculate the sp2/sp3 ratio,

which were 0.61 and 0.42, for N3FC and a-C respectively.

(a) (b) (c)

(a) (b) (c)

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Fig. 3 - X-ray diffractogram of N3FC (blue) a-C (yellow).

Fig. 4 – Raman spectra of N3FC (green) and a-C (red)

Table 2 - Ultramicrohardness for the three samples tested: SS, N3FC, a-C. Hardness (GPa)

SS 2 [2]

N3FC 11±3

a-C 15±4

The surfaces coated with DLC N3FC became more hydrophilic than SS (Figure 5 a)), which may

be related with its higher sp2 content. In fact, it was found by several authors that hydrophobicity

decreases with the decrease in sp3-bonded carbon in the films [3,4].

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800 900 1 000 1 100 1 200 1 300 1 400 1 500 1 600 1 700 1 800 (a.u.)

dlc1_2sdlc2_8s

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The decrease of the contact angle of the protein solution with time (Figure 5 b)) was more

pronounced on the DLC surfaces, especially on N3FC, which suggests an higher tendency of the

protein to adsorb on these surfaces.

Fig. 5 - Evolution of the contact angle with time for SS, N3FC and a-C in (a) water

and (b) BSA+HBSS.

The adsorption tests with QCM-D and ellipsometry (Figure 6) confirm that albumin adsorbs on

higher amounts to the surfaces of DLC's than to SS. The ellipsometric results show that the amount of

protein adsorbed to a-C surfaces is the highest. Comparison of results obtained by QCM and

ellipsometry suggest that the protein layers present on SS and N3FC should be strongly hydrated.

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Fig. 7 - Comparison of adsorption results obtained by QCM-D (orange) and ellipsometer (green).

The tribological performance of the DLC’s coatings against UHMWPE was evaluated in the

presence and absence of albumin, and the results were compared with those obtained with SS (Figure

7).

The tribological behaviour of the DLC’s is not better than that observed with SS. The average

friction coefficient is slightly higher with DLC’s than with SS. The dispersion of the results obtained

with the DLC’s is higher, both in HBSS and BSA+HBSS, which can be related with the more

heterogeneous nature of the surfaces. Images of optical microscopy (Figure 8) indicate that the wear

of UHMWPE is not lowered when the counterfaces are the DLC’s.

Protein solution revealed a less efficient lubricant than the saline solution HBSS. Indeed, the

presence of protein led to higher friction coefficients in all the studied systems and to a larger wear of

the polymeric surface, which revealed more severe when DLC's were used as counterfaces. The

observed wear mechanisms were mainly abrasion of the polymer with its transfer to the counterface

(Figure 9). Changes in the conformation of the adsorbed protein and possible denaturation shall be

behind the reduction of its lubrication ability.

Fig. 7 - Friction coefficient vs distance in (a) HBSS and in (b) BSA+HBSS, for SS, N3FC and a-C

with normal force 25 mN.

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Fig. 8 - Optical microscope images (150x) of the worn surface of UHMWPE tested against (a) and

(b) SS, (c) and (d) N3FC, and (d) and (e) a-C, in HBSS and BSA+HBSS, respectively.

Fig. 9 - SEM images of SS surfaces after tribological experiments against UHMWPE in

BSA+HBSS.

(a) (b)

(c) (d)

(e) (f)

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00,10,20,30,40,50,60,70,8

0 10 20 30

Fric

tion

Coe

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Distance (m)

1mN

25mN

Reduction of friction and wear, observed in biotribological pairs involving DLC coated surfaces and

UHMWPE, was attributed by Ali [1] to the build up of a transfer layer of DLC on the softer polymeric

counterpart, when the lubricant was distilled water or NaCl solutions. In the presence of the

biomolecules existent in the periprosthetic fluid, no DLC layer transfer occurs and the UHMWPE wear

cannot be lowered. In our case, DLC coatings did not improve the tribological behaviour of the pair

SS/UHMWPE with any of the lubricants.

The effect of the sliding velocity and of the applied normal force was also investigated.

The results for the change of velocity were not conclusive. In the range of velocities investigated

(0.5 – 2.5 cm/s) this parameter does not seem to play a significant role in the tribological behaviour of

the systems.

Concerning the effect of the force, two distinct forces were tested: 1mN and 25 mN (Figure 10).

The value of the friction coefficient increases with the decrease of the force. This shall be related with

the influence of the adhesion forces, which become important at lower loads. More, in the presence of

protein, the friction coefficient at the lower force increases during the experiments, which shall be

attributed to possible changes in the conformation of the protein that result in a high-shear-strength,

with strong hydrophobic interactions between the albumin and the surface and between adjacent

albumin molecules.

Fig.10 - Friction coefficient vs distance in (a) HBSS and in (b) BSA+HBSS, for N3FC with normal

forces 1 mN and 25 mN.

Conclusions

The results of this work revealed “unexpectable problems” with the tribological performance of pairs

DLC/UHMWPE. As found by other authors [1], the studied DLC’s coatings did not improve the

tribological behaviour of the pair SS/UHMWPE, independently of the presence of albumin in the

lubricant. The reasons for this fact may be related with the quality of the coatings, its higher hardness,

or the absence of transfer of a DLC layer to the polymer, among other factors. Further studies will be

necessary to clarify this question.

However, the use of these DLC coatings may be effective to protect the underlying metal against

scratching, corrosion and ion release.

00,10,20,30,40,50,60,70,8

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25mN

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References [1] C. Donnet, A. Erdemir, Tribology of Diamond-Like Carbon Films, Springer Science + Business Media (2008).

[2] X. H. Chen, J. Lu, L. Lu, K. Lu, Tensile properties of a nanocrystalline 316L austenitic stainless steel, Scripta Materialia 52(10) (2005) 1039-1044.

[3] R. Colaço, A. P. Serro, O. L. Eryilmaz, A. Erdemir, Micro-to-nano triboactivity of hydrogenated DLC films, Jounal of Physics D: Applied Physics 42 (2009) 177-185.

[4] R. Paul, S. N. Das, S. Dalui, R. N. Gayen, R. K. Roy, R. Bhar, Synthesis of DLC films with different sp(2)/sp(3) ratios and their hydrophobic behavior, J Phys D-Appl Phys 41 (5) (2008) 55309-15.