MODIFICATION HYDROXYAPATITE WITH ZnO AS

ANTIBACTERIAL
Dwi Aprillia Wulandaria, Diana Vanda Welliaa , Novesar Jamaruna
a

Materials Chemistry Laboratory Department of Chemistry, Science Faculty, University of

Andalas
e-mail: dwiaprillia99@yahoo.com
Department of Chemistry, Faculty Unand, Limau Manis, 25163

Abstract
Modification with zinc oxide (ZnO) via sol-gel method has been successfully carried
out. The analysis of X-Ray Diffraction (XRD) showed hexagonal crystal system and
the addition of zinc can increase the intensity and crystallinity of the sample. The
spectrum of Fourier Transform Infrared (FTIR) showed the presence of functional
groups suitable for the identification of HA in wave numbers 3000-3600 cm-1, 10001045 cm-1 and 1300-1490 cm-1 indicating the presence of the O-H, P-O and C-O,
respectively. Antibacterial test has been done using disc diffusion method and
resulted that the capability of HA as antibacterial increasing with the increase of ZnO
concentrations. HA/ZnO 1:1 with a concentration of 1000 ppm has inhibitory activity
with a maximum diameter of 8 millimeters.
keywords: Hydroxyapatite, ZnO, Sol gel, Antibacterial.
I. Introduction
Hydroxyapatite (HA), Ca 10 (PO 4) 6 (OH)
2, is the main mineral constituent of
bone and tissue sources of calcium in
living organisms. Synthetic HA is an
inorganic material that shows the
versatile nature of biocompatibility,
bioactivity and osteoconductivity. HA
also has the ability as an antibacterial
that can inhibit biofilm formation and
colonization of planktonic cells.1
HA is often modified with other
metals to obtain superior biological
properties again. Some form of
modification in the form of the HA
surface coating with another metal or
metals deposited there as well with the
the matrix HA.1 Several studies have
been done conducted on several HA is
coating with silver (HA/Ag) for the
purpose of obtaining the optimum
capability HA and Ag as antibacterial,
against gram-positive bacteria and
negatif2,
as
antifungal
Candida
albicans1, with Zirconia (HA/Zr) to
improve the mechanical properties so
more robust.3

In addition to Ag metal, other

metals that can be used to modify HA
to be able to enhance the biological
capability as antibacterial, among
others, can be used with heavy metals
oligodinamic high power, one of which
Zn. The use of Zn metal as a carrier of
metal ions Zn2+ has high antibacterial
properties, good chemical stability,
and relatively low prices. Zn is known
as an essential heavy metals, toxic
metal though, it is needed by the body
for growth in very small amounts.
Toxicity owned Zn metal will effect
when it enters the body in the sheer
number of large organisms or exceeds
the threshold toleransinya.4 Research
on HA modified with Zn is expected to
improve the ability of HA as new in the
field of inorganic antibacterial.
Tests conducted on the bacteria
Escherichia coli (E.colli) by disc
diffusion method to look at the
diameter of the clear zone is not
overgrown
with
bacteria.
This
bacterium is a reasonable amount of
beneficial bacteria for digestion, but if
excessive
breeding
can
produce
enterotoxin of E. coli as a cause of

some
cases
of
diarrhea
and
meningitis.5
In this study, the method used is
the sol-gel method, because it has a
better purity and can be used at low
temperatures, one of which is in the
process of synthesis HA.6
II. Research Methodology
2.1.
Chemicals,
equipment
and
instrumentation
For the materials used in the study,
among
others,
Synthetic
hydroxyapatite obtained from the
Materials
Laboratory
(Andalas
University),
Media
Nutrient
Agar
(Merck), Nutrient Broth Media (Merck),
cultures of E. coli, 65% HNO3 (Brataco),
distilled water, (NH4)2HPO4 (Merck),
NH4OH technical (Brataco), Zn (NO3) 2
(Merck).
While equipment used in the study,
including glass cup, erlenmeyer, test
tubes, measuring cup, ose, spatulas,
hot plate stirrer, magnetic bars,
analytical balance, buchner spiritus,
sterile cotton, cotton buds, calipers,
oven, incubator, autoclave, furnace,
pulvurizer.
Tools of instrumentation used X-Ray
Diffraction
(XRD;
Shimadzu
diffractometer 610 40 kV - 30 mA, Cu
K), Fourier Transform Infrared (FTIR ;
Jasco FTIR-410), Scanning Electron
Microscopy-Energy Dispersive X-Ray
(SEM - EDX; JEOL SEM).
2.2. Research procedures
2.2.1 Modification of HA/ZnO
Into a 500 mL HA gel was added Zn
(NO3)2 with various concentrations of
HA/ZnO
1: 0.25, 1: 0.5, 1: 0.75 and
1: 1. Stirring during the process of
adding a constant speed. After the
addition of Zn(NO3)2 is completed
stirring process continued for 10 min
and aging for 24 hours to form a gel.
Obtained gel was heated in a 70 C
oven for 24 hours, powder in furnace
at a temperature of 600 C. Powder is
obtained HA/ZnO with a variation of 1:
0.25, 1: 0.5, 1: 0.75 and 1: 1.
2.2.2 Antibacterial Activity Test
The process begins with a media NB,
then carried to the culture of E. coli,
after 24 hours, the bacteria grow with
marked turbidity in NB media. Testing

is done by setting the inhibition of 2

pieces
that
have
petridish
NA
containing solid media. Each sample is
tested against HA and HA / ZnO with a
variation of 1: 0.75 and 1: 1 with a
concentration of 500 and 1000 ppm
diluted with distilled water.
Petridis inserted into each each paper
disc that has been soaked in
antibacterial solution (HA and HA /
ZnO). At the center of the petri paper
disc placed as a negative control. After
24 hours of observations were made of
the clear zone are not overgrown
microbes. Measured against clear zone
diameter and compared with a
standard diameter of the inhibition of
HA and Zn on the growth of bacteria.
III. Hasil dan Pembahasan
3.1. Analysis
of
the
results
of
characterization
The addition of Zn (NO3)2 as a source
of
Zn2+ to form ZnO. Increasing the
intensity of the color is directly
proportional to the increase in the
concentration of Zn(NO3)2 is added to
the HA matrix, the composite ratio of
HA/ZnO 1: 1 is the intensity of the
green color is more concentrated.
Powder HA/ZnO obtained at various
concentrations is characterized by a
wide
range
of
instrumentation
equipment under the same conditions.
Characterization by XRD, conducted
on
3
samples
at
calcination
temperature of 600 oC with a ratio
HA/ZnO 1: 1, 1: 0.75 and HA synthesis.
XRD data of test results compared to
the standards of the International
Centre for Diffraction Data (ICDD) for
ZnO with refference code 00-019-1463
and 01-075-9526 for refference code
hydroxyapatite.

on the surface of the sample. Additions

of Zn on the HA does not affect the
results of its FTIR spectrum. So it can
be concluded that, no new chemical
bonds are formed between HA with
ZnO.

Figure 1. diffractogram XRD results of

HA/ZnO concentration ratio 1:1 after
calcination at 600 oC for 3 hours.
The results of XRD analysis in Figure
1 in the form of HA/ZnO variation in
concentration of 1:1 shows the best
crystallinity with the highest intensity
at 2 angles 31,74o, while the peak for
ZnO is found at an angle 2 32,16o. HA
crystals obtained system is hexagonal
with
space
group
P63/m.
The
difference of the three samples
HA/ZnO 1:1, 1:0.75 and HA synthesis
seen in crystallinity and increase the
intensity of Zn(NO3)2. From the XRD
data sample results, the more sources
of Zn were added to the HA will
increase the intensity of the peaks and
minimize the width of the peak of HA.
Thus modification of HA with ZnO will
increase the peak intensity and narrow
peak width thus increasing the
crystallinity of HA.
The analysis of Fourier Transform
Infrared (FTIR) is very important to
know the functional groups found in
the sample, whether it is in accordance
with the existing functional group on
HA, Ca10(PO4)6 (OH)2. Some functional
groups that can be identified by FTIR,
among others, the hydroxyl group on
the O-H bond P-O and C-O bond in the
fingerprint region. Of the three FTIR
spectra as found in figure 2 seen a
wide valley that marks the OH
stretching specific to HA over wave
number 3000 cm-1 to 3600 cm-1. While
the PO group stretching at 1000-1045
cm-1 wave number. CO group is found
at 1300-1400 cm-1. At the peak of 1632
cm-1 seen the OH bending vibrations

a.
Figure 2. FTIR spectra of the samples
HA/ZnO concentration ratio 1: 1 after
calcination at 600 C for 3 hours,

3.2 Application Antibacterial Activity of

HA and HA/ZnO
Antibacterial test
activities as an
antibacterial agent against E. coli is
done by using the disc diffusion
method. Where the inhibition of
antibacterial can be seen in the
diameter of the clear zone is not
overgrown with bacteria and compared
with the negative control.
Figure 5 showed antibacterial test
results on samples of HA, HA/ZnO 1:
0.75 and HA/ZnO 1:1. The images
show the diameter of the clear zone on
the concentration of HA/ZnO at a
concentration of 1000 ppm of 1:1 by 8
mm,
for HA/ZnO 1:0.75 at a
concentration of 1000 ppm obtained
clear zone size of 6 mm, while for the
HA concentration of 1000 ppm only
obtained a diameter of 5.7 mm. The
results obtained show HA/ZnO 1:1
coined the inhibitory effect on bacterial
growth greater than HA/ ZnO 1: 0.75
and synthesis HA.

b.

c.

References
1. Zamperini,C.A.,Andre,R.S.,Longo,V.M
., 2013, Antifungal Applications of
Ag-Decorated HA Nanoparticles,
Hindawi Journal, No. ID 174398, Hal
9.
Figure 5. Clear zone around the growth of
E. coli that has been given a antibacterial
HA and HA/ZnO (at a concentration of 500
ppm and 1000 ppm).

The ability of increasing inhibition,

comparable with the addition of Zn to
the HA. The positive charge of the Zn2+
ion and a negative charge on the
bacterial cell membrane will cause
attraction between
Zn2+ ions and
negatively charged areas on the
bacterial cell wall to form an
electrostatic bond. This will form an
electrostatic bond stress leads to
damage the cell wall permeability,
reducing the normal intake of nutrients
to sustain life, causes cells to become
dead.4 Thus, a growing number of Zn2+
in the sample, the greater the ability to
inhibit the growth of E. coli bacteria.
IV. Conclusion
Modified hydroxyapatite with ZnO via
sol gel method has been successfully
carried out. Analysis X-Ray Difraction
(XRD) showed that the addition of zinc
can increase the intensity and
crystallinity of the sample. Spectrum of
Fourier Transform Infrared (FTIR)
shows, the presence of functional
groups suitable for the identification of
HA which indicates the OH group, PO
and CO. Antibacterial test by disc
diffusion method showed the ability of
HA as antibacterial increasing to an
increase in the concentration of ZnO.
HA/ZnO 1: 1 at a concentration of 1000
ppm maximum antibacterial activity
against Escherichia coli
with a diameter of 8 mm.
V. Acknowledgements
Acknowledgements profusely, the two
counselors,
who
have
provided
comments and suggestions in the
smooth conduct of research and in the
preparation of the journal.

2. C.S.Ciobanu,
2012,Antibacterial
activity
of
silver-doped
HA
nanoparticles against gram-positive
and
gram-negative
bacteria,
Nanoscale Research Letters, No.1,
Vol.7, Hal 324.
3. Vasconcelos,M.C.2011,Barreto,
Tailoring the Microstructure of Sol
Gel Derived Hydroxyapatite/Zirconia
Nanocrystallin
Composites,
Nanoscale Research Letters, 2011,
No.6, Hal. 20.
4. Angraini,
D.,2013,Sintesis
Zn(II)/Silika dengan Metode Sol-Gel
sebagai
Material
Antibakteri
terhadap
Escherichia
coli
dan
Staphylococeus
aureus.
Thesis,
FMIPA,program pasca sarjana, UGM,
Yogyakarta.
5. Kusuma,F.A., 2010, Escherichia Coli
,Fakultas Farmasi UNPAD,Bandung.
6. Agrawal,
K.,
Gurbhinder,
S.,
Devendra, P., Satya, P., 2011,
Synthesis and Characterization of
Hydroxyapatite Powder by Sol-Gel
Method for Biomedical Application,
Metallurgical
and
Materials
Engineering Journal, No. 8, Vol.10.