3rd International Conferences and Workshops on Basic and Applied Sciences 2010

ISBN: 978-979-19096-1-7

The Effect Of Composition Co:Cr Variation to Mechanical

Properties And Cytotoxicity Co-Cr-HA Composite
Aminatun, Siswanto, Fitri Wijayanti*
*Departement of Physics
Airlangga University, Surabaya, Indonesia
e-mail: aminatun_unair@yahoo.co.id
low level of density, corrosion resistance, and
acceptable to the body or does not cause
hypersensitivity. However, based on its mechanical
strength, the Ti still cannot be used in several
medical applications (Shamsul, et.al, 2007).
Besides the presence of Al and 4% V ions in
titanium alloys provide long-term health problems.
Low resistance of titanium alloys which can
accelerate ions presence is harmful to the body
(Goenharto, et al, 2008).

Abstract
Sintesis of composite Co-Cr-HA (Cobalt
Composite Implants / CIC) created by varying the
composition of Co: Cr respectively (82%: 18%),
(79%: 21%), (76%: 24%), (73%: 27%), (70%:
30%) of added 5% HA from the Co-Cr alloy.
Manufacturing process involves mixing, pressing
and sintering. The process of mixing ingredients
produced using milling machines with a speed of
600 rpm for 30 minutes and then pressed at a
pressure of 370 MPa and sintering carried out at a
temperature of 1000C for 3 hours. XRD test
results showed that the diffraction pattern of crystal
form and the emergence of phase oxide layer CoO,
Co3O4 and CrO2 on the surface of the CIC result of
sintering processes that do not vacuum. Sample
hardness value of 351.096 13.339 kgf/mm2 to
490.130 76.540 kgf/mm2. Compressive streght
increases with the addition of the composition of
Cr, the highest value indicated the composition of
CIC in the ratio of Co: Cr by 70%: 30% of 102.979
0.153 N/mm2. Test on fibroblast cells by MTT
assay method obtained results that the percentage of
living cells with the highest composition of
95.837%. Overall characterization of CIC showed
good results, it is shown by the percentage of living
cells is still above the tolerance limit of 60%.

The development of new biomaterials-based alloys

or composites that can be applied in medical safety
is required. Found Co-Cr alloy and has been used as
base material for the first time in the medical dental
implants due to the planting of Co is very good
resistance in the oral environment (Samsul, et al.,
2007). Furthermore, studies done about the addition
of hydroxyapatite to the Co-Cr alloy.
Shamsul et al., 2007, utilized Cobalt Composite
Implants (CIC) which is a Co-Cr alloy with a
ceramic hydroxyapatite (HA). The study varying
the composition of the HA between 0% to 20%
added to the Co-Cr alloy. On the variation of 5%
HA, otherwise have better properties when
compared with the other composition variations. In
that study said that the composite Co-Cr-HA with
the addition of 5% HA can be applied in the
biomedical as bone implants, but not exposed to the
metal alloy composition of Co-Cr in the composite.
It is therefore necessary to modify the composition
of Co-Cr in order to know the potential of CIC with
the mechanical properties in accordance with
human bones.

KeyWords: Cobalt Implant Composite (CIC),

Pressing, Sintering, Hardness, Compressive Streght
1

Introduction

Types of biomaterials used in medical applications

and dentistry are a metal (alloy), ceramics,
polymers, and composites. Biomaterials are used to
replace or repair a tissue or organ function which
damaged. Material from the type of metal's mostly
used as bone implant biomaterials are stainless
steels (316L-steel), Co-Cr alloy, pure Ti (cpTi), and
Ti alloy (Ti-6Al-4V) (Shamsul, et al. 2007).

Methodology

2.1
Process Samples
Sample preparation in this process involves mixing
powders Co, Cr and HA. Material is weighed
according to the composition in Table (1) and then
mixed using a milling machine equipped with speed
control device that is dimmer. The addition of
alcohol is very helpful in smoothing the mixture,

Titanium has been widely used in medical and

dentistry because of its good like having a relatively
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Aminatun, The Effect of Composition Co:Cr Variations to Mechanical Properties and Cytotoxicity Co-Cr-HA
Composite

due to the materials used (cobalt, chromium and

HA) is insoluble in water. And then, the sample
milled using 600 rpm of speed for 30 minutes
(Shamsul, 2007). Afterwards, the samples are
removed from the tube milling and then dried,
because the sample in wet state after being mixed
with alcohol. Samples in the form of powder is
weighed according to the needs of characterization
then pressed with a pressure of about 370 MPa.
Samples sintered at 1000oC for 3 hours with the rate
of increase in 20 C / min so as to achieve 1000oC,
and held for 2 hours (Shamsul, 2007), then
performed the characterization. Furthermore, on the
whole sample were characterized by XRD test,
Vickers hardness and compressive strength.

the incubation for 4 hours and then add in DMSO

(dimethyl Sulfoxida). After 15 minutes, the cell
cultures in a shaker and then read using Elisa
Reader Spectrophotometer.
The percentage number of live cells for MTT test
can be calculated by the following equation:
% livecell =

Co (%)
82
79
76
73
70

Cr (%)
18
21
24
27
30

x 100 %

With, % Live cells = percentage of cells after

treatment, treatment OD = optical density values of
samples after treatment, media control OD = optical
density value of media control, cell control OD =
optical density value of control cells.

Table 1: CIC constituent raw material

composition in % of weight
Sample
I
II
III
IV
V

OD treatment + OD mediacontr ol
OD cellcontro l + OD mediacontr ol

HA (%)
5
5
5
5
5

Result

3.1
The test of X-ray Diffractometer (XRD)
XRD test was conducted to identify synthesized
compounds through a search match using the
program Crystal Impact analyst. Results of search
match XRD data shown in Figure (1), (2), (3), (4)
and (5)

2.2
Cytotoxicity testing
Cell BHK 21 (Baby Hamster Kidney 21) clone 13
is saved at temperature of 85C in Nunc vials with
eagle medium, carried out thawing stored is placed
in an incubator at 37C to melt . Then centrifuged
at 3000 rpm for 10 minutes, the precipitate is a cell
culture; storage media are located on the top and
discarded. 20 ml of cell culture is inserted into a
small roux bottle added by grower medium of eagle
and also added by 10% of calf serum, and then
inserted into a CO2 incubator at 37C for 2 x 24
hours, cells will be full.
Cell cultures were divided into micro plate 96 well
each 1.5 ml plus eagle medium and 10% of bovine
serum, then put in a CO2 incubator with a
temperature of 37C for 2x24 hours. Once
confluent the media was removed and washed 2
times with PBS solution, monolayer culture which
is clean is then added back with the eagle media
and the EMS 5% which was mixed with a sterile
CIC sample. Another one that is used as a control is
cell culture medium without CIC compound, and
then all of a micro plate is stored again in incubator
CO2 for 24 hours.

Figure 1: Search match curve CIC-I

Monolayer cell cultures that have been removed

from the CO2 incubator, then the media was
removed and washed with PBS 5 times to clean
waste products of metabolism of cells. Then,
ditrypsinasi cell culture with versence trypsin
0.25% to deteriorate the cell attached to the wall of
the bottle. Once added with MTT, cell cultures in

Figure 2: Search match curve CIC-II

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3rd International Conferences and Workshops on Basic and Applied Sciences 2010

3.2

ISBN: 978-979-19096-1-7

Mechanical properties

Table 2: The test results of mechanical properties

of CIC

Figure 3: Search match curve CIC-III

Sample

Vickers Hardness/
VHN (kgf/mm2)

371,463 12,404

79,589 0,122

II

351,096 23,105

94,627 0,140

III

426,077 85,500

99,152 0,152

IV

490,130 132,572

102,668 0,157

392,888 19,341

102,979 0,153

Compressive
strength
(N/mm)

3.3
Microvickers Hardness
Measurement of hardness value of CIC is using a
hardness tester with a suppressor on the sample
surface with a 136 angle of diamond pyramidshaped. The amount of loads should be greater than
the yield strength of metals to plastic deformation
occurs. VHN values taken on 3 (three) points of the
sample surface so that CIC hardness values
obtained as in Table (2) and Figure 6.
Figure 4: Search match curve CIC-IV

G
rafik Kekerasan
CIC
adap
Vickers
Hardness
Vsterh
Composition
Variasi komposisi Co-Cr
VHN
(kgf/mm)

Co-Cr Variation

600
500

IV
III

400

II

300
200
100

CIC

0
0

Figure 6: Vickers Hardness CIC

Increasing Cr composition should increase the
value of CIC hardness because of the nature of
individual Cr is harder than the Co. But apparently
at the highest composition of Cr (30%), hardness
value is decreased. As shown in Figure (6). The
presence of oxide compounds CoO, Co3O4, CrO2,
believed to be the cause of the hardness decreases.
Besides, the samples do not appear homogeneous,
as seen from the standard deviation of hardness
value is still quite large.

Figure 5: Search match curve CIC-V

Based on Figure (1), (2), (3), (4) and (5), search
match results show that in each sample appears
compounds CoO (cobalt oxide), Co3O4 (cobalt
oxide), CrO2 (chromium oxide) and HA (CA10
(PO4)6 (OH)2). The presence of oxides in each
constituent compound of CIC as a result of the
sintering process during sample preparation is not
done in a vacuum furnace (air vacuum), so that the
oxygen in the furnace will bind to the elements Co
and Cr to form CoO, Co3O4, CrO2.

3.4
Compressive strength
Compressive strength is the maximum stress that
can be accepted by a material in the form of
compression or tension without failure. The tools
used are autograph equipped with a Control /
Measure Unit. This tool provides a force from
above or from below the surface of the sample so
that the magnitude of P (force) when the specimen

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Aminatun, The Effect of Composition Co:Cr Variations to Mechanical Properties and Cytotoxicity Co-Cr-HA
Composite

is damaged will be read on the instrument so that

the compressive strength of CIC can be calculated,
as shown in Table (2).

Citotoxicity Vs Composition
Co-Cr Variation
%Live cell

G
rafik Kekuatan
TekanVs
CIC
Terhadap
Compressive
strength
Composition
Variasi komposisi Co-Cr

Co-Cr Variation

(N/mm)
120
100

IV

III

II

80

60

40

Figure 8: Grafic % Live Cell of CIC

20
0

CIC
0

Figure 7: Compressive Strength of CIC

Figure (7) show a good correlation between the
compressive strength of the composition of Co-Cr
variation. The images provide information that the
compressive strength value of the CIC will continue
to rise if the composition of Cr increase (decrease
compositions Co.), this is in line with the density of
the CIC. Value of compressive strength of human
cortical bone by 30-160 N/mm2 then from the
result above shows that the value of compressive
strength CIC 79.589 N/mm2 - 102.979 N/mm2 have
compatibility with these networks.

However, chromium and alloys CoCr are well

tolerated without significant toxicity by cell lines
with concentrations below 50%. In this study, the
composition of chromium is added at 18-30%, with
the hope of Co-Cr composition variations on the
CIC does not cause any toxic properties. If
associated with XRD results, which make up the
CIC of CrO2 that have properties not toxic because
they do not contain Cr6+ ions that are dangerous to
human tissue. Most living cells are shown by the
CIC V with OD value of 0.266 0.043 where the
value of its OD close to the OD in the control cells,
namely the composition with the highest Cr
(composition Co lowest).

3.5
The results of cytotoxicity test
Elisa Reader readings indicate that the CIC is not
toxic to fibroblast cells (cell lines), this is indicated
by the percentage of living cells is still above 60
percent ( 60%), the optical density of the treatment
is still close to the optical density of control.

Table 3: The percentage of fibroblasts with

MTT test
Sampel
I
II
III
IV
V

Optical
Density
(OD)
0,177 0 033
0,191 0 036
0,209 0 034
0,221 0 040
0,266 0 043

Figure (8) shows the cells can live in the CIC

environment, so that CIC can be applied in
biomaterials. According to in vitro studies have
been conducted, stating that the cobalt is toxic to
humans as in cell lines in culture medium.

Conclusions

1. Variation in the composition of Co-Cr affects

the mechanical properties (hardness and
compressive strength) as well as cytotoxicity
CIC.
2. Value of hardness and compressive strength
increases as an increasing of Cr composition
(decrease in Co) and the value of both in
accordance with the cortical bone.
3. Toxicity value decreases as an increasing of Cr
composition (decrease in Co). The entire
samples in this study are not toxic indicated by
the percentage of cells that lived more than
60%.

% Living Cells
(OD)
78,606
81,317
84,947
87,294
95,837

Table (3) is the result of calculating the value of the

optical density of each sample CIC invested on the
eagle and the EMS media containing BHK-21
fibroblast cells. Optical density can be interpreted
as the ability of a material to absorb the light. OD
value equivalent to the number of live cells within
the CIC. The higher the OD value then the more
and more of the living cell, so that the table shows
that most living cells is occur in cells with a CIC V
environment .

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ISBN: 978-979-19096-1-7