Advances in Health Sciences Research, volume 32

Proceedings of the 1st Aceh International Dental Meeting (AIDEM 2019), Oral Health
International Conference On Art, Nature And Material Science Development 2019

Effect of Hydroxyapatite From Nile Tilapia

(Oreochromisniloticus) Scale on Surface Hardness of
Conventional and Resin Modified Glass Ionomer
Cement (In Vitro Study)
Astrid Yudhit 1, *, Kholidina Harahap 2 and Sabrina Chairunnisa Nasution 3
1
Department Dental Material and Technology, Faculty of Dentistry, Universitas Sumatera Utara, Indonesia
2
Department Dental Material and Technology, Faculty of Dentistry, University of Sumatera Utara,
Indonesia
3
Undergraduate Student, Faculty of Dentistry, Universitas Sumatera Utara, Indonesia
*Corresponding author email : [email protected]
Authors email : [email protected]

ABSTRACT
Surface hardness of restoration materials affected its wear resistance and longevity. The addition of hydroxyapatite
could improve mechanical properties of restoration material. Hydroxyapatite can be synthesized from fish scales for it
contain calcium carbonate. The aim of this study was to evaluate the surface hardness of conventional and resin
modified glass ionomers cement by incorporation of different wt% of hydroxyapatite from Black tilapia
(Oreochromisniloticus) scales. Hydroxyapatite was synthesized from black tilapia scales by calcination method.
Samples with a diameter of 5 mm and a height of 2 mm were made from conventional and resin modified glass
ionomer cement, then divided into 7 groups (n=10), namely control, 2wt% GIC, 5wt% GIC, 8wt% GIC, 2wt%
RMGIC, 5wt% RMGIC, and 8wt% RMGIC. Surface hardness test was measured with Vickers Hardness Tester.
Result showed hardness value from each groups respectively were 49.81 ± 3.55 VHN, 34.49 ± 1.82 VHN, 32.58 ±
1.61 VHN, 36.63 ± 3.09 VHN, 32.05 ± 3.77 VHN, 36.23 ± 4.27 VHN, and 19.27 ± 1.71 VHN. One-way ANOVA test
showed a significant differences in all group (p <0.05). It can be conclude that hydroxyapatite from fish scale had an
effect on surface hardness of conventional and resin modified glass ionomer cement, whereas it decreased the
hardness of conventional GIC, yet it increased the RMGIC hardness.

Keywords: hardness, hydroxyapatite, fish scale, gic, rmgic

Copyright © 2021 The Authors. Published by Atlantis Press B.V.

This is an open access article distributed under the CC BY-NC 4.0 license -http://creativecommons.org/licenses/by-nc/4.0/. 5
 Advances in Health Sciences Research, volume 32

1. INTRODUCTION and source. It was reported that adding synthetic HA

Glass ionomer cement (GIC) was first introduced into GIC and RMGIC can improved the properties of
in 1971, then widely use and still popular until now cement, for example adding 8wt% HA increased
due to its fluoride release characteristics, chemical flexural strength [16], 5wt% HA increased surface
bond with tooth structure and metal, easy to handling, hardness of GIC [17], and 2 and 5wt% nano
and low coefficient of thermal expansion. Main hydroxyapatite increased wear resistance of RMGIC
component of GIC is basic glass powder, water and [18]. Some studies used HA from eggshell and
polyacid [1,2]. Glass ionomer cement is the most reported that 8wt% [19] increased GIC’s flexural
suitable material to use in FDI World Dental strength and 5wt% [20] HA increased its hardness.
Federation new preventive dental treatment concept Fish scale was a waste that can be used as a source
like minimum intervention [3]. Moreover, GIC also of hydroxyapatite [10,11,21]. Fish scale containing
used on World Health Organization (WHO)’s fats, water, protein, and some chemical compound like
atraumatic restorativEtreatment, dental treatment calcium phosphate in high concentration. The calcium
procedure that does not requires complicated phosphate in the fish scale can be process to be a
techniques [4]. Despite advantages, GIC has poor hydroxyapatite [21]. Nile Tilapia
mechanical properties among other dental restoration (Oreochromisniloticus) fish is the top rank production
materials, and it made GIC has limitation as of freshwater fish in North Sumatera province of
restoration on high stress bearing area [2]. Some Indonesia. Base on this fact, we have an interest to
efforts have been done in order to overcome these utilize Nile fish scales as a source of hydroxyapatite
disadvantages, like incorporation GIC with metals, and incorporate it into GIC and RMGIC to improve
glass fiber, hydroxyapatite, silica, and CPP-ACP [5]. the properties.
Resin modified glass ionomer cement (RMGIC) The aim of this study was to evaluate the effect of
was used in dental clinic on 1991 [6]. RMGIC has hydroxyapatite from fish scale (Oreochromisniloticus)
main component as same as conventional GIC with an on surface hardness of conventional and resin
adding of monomer component (typically modified glass ionomer cement. In this study,
hydroxyethyl methacrylate/HEMA) and conjoin hydroxyapatite particles were adding to GIC and
initiator system (Camphorquinone) [1,2,6]. Moreover, RMGIC by 2, 5, and 8 wt% of GIC and RMGIC’s
RMGIC was developed to manage aesthetic, shorten powder.
the setting time, and improved the mechanical strength
of conventional GIC [2,6]. Nevertheless, RMGIC has 2. MATERIALS AND METHODS
high flexural strength but low in compressive strength Materials used in this study were conventional
[2]. glass ionomer cement for restoration (Fuji IX, GC,
Some previous studies was done in order to Japan), Resin modified glass ionomer cement with
improve the mechanical properties of GIC and shade A3 (Fuji II LC, GC, Japan), and Hydroxyapatite
RMGIC by adding material as reinforced, such as from Nile Tilapia fish (Oreochromisniloticus) scales
fibres or particles like silicon carbide (SiC), that produced by using combination methods of
fluoroapatite nano-bioceramics, and forsteritenano- alkaline heat treatment and calcination method
particles, and hydroxyapatite [7,8,9]. Hydroxyapatite [11,12].
is a bio-ceramics material that widely used in the Synthesis of hydroxyapatite. Fish
biomedical fields due to its similar structures to the (Oreochromisniloticus) scale waste was freshly
main minerals construction of bones and teeth [10,11]. collected from local market in Medan, North Sumatera
Hydroxyapatite can produce from various sources, Indonesia. The fish scales soaked and washed
both natural and synthetic. Natural sources of rigorously in distilled water to remove salts and dirty
hydroxyapatite such as mammalian bones [12], matters then let it dried in open air under the sun light
clamshells [13], rocks [14], and eggshells [15]. for 2 hours and grounded sifted using 200 mesh (74
Hydroxyapatite can also be made in the laboratory µm) sieve. The scales splinters were washed with 0.1
through a series of chemical processes. There are M HCl and several times with distilled water to
several methods for making hydroxyapatite crystals, remove the protein. Further, the fish scales splinter
like precipitation methods, biomimetic deposition, sol- were treated with 5% (w/v) NaOH, heated and stirred
gel method, electro deposition method, and at 70ºC for 5 hours to eliminated the remaining protein
calcinations [16]. and result a precipitate. These precipitate powders
Previous studies have been conducted to evaluate were treated with 50% (w/v) NaOH, heated up to 100
the effect of adding hydroxyapatite to glass ionomer ºC and stirred for 1 hour for alkaline heat treatment.
cement and these studies different on HA percentage

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The powder washed rigorously with deionized water was RMGIC+8wt%HA. Post hoc analysis shows all
until neutral and then dried at 60 ºC for 8 h. The group were significant difference compare to control.
calcination of the powder was done in 800 ºC for 1 h
to produce the hydroxyapatite (HA).
Table 1. Mean and standard deviation (SD) of Vickers
GIC and RMGIC Samples. In this study, groups were control GIC and RMGIChardness
without for
HA,each
GIC+2wt%
groups HA, GIC+5wt% HA, GIC+
GIC and RMGIC was mixed manually and placed into and One way ANOVA between groups.
metal master mould with 5 mm in diameters and 2 mm
in depth. For the GIC, celluloid strip and glass object GIC + HA p RMGIC + HA p
wt%
was placed on top and bottom of the samples. Then, Mean SD value Mean SD value
placed the 1 kg load on the top of sample until it set 2.95
control 49.813 3.551 22.362
for 2 minutes and 20 seconds. For the RMGIC, 1
celluloid strip and glass object was placed on top and 3.77
2 34.492 1.825 0.00* 32.050 0.00*
bottom of the samples. Then, placed the 1kg load on 1
the 4.26
5 32.584 1.610 36.227
6
top of sample for 5 minutes. The setting proccess was 1.70
8 36.660 3.122 19.270
done by using LED light cured unit for 20 seconds 8
with tip unit direction was perpendicular on top of *there was a significant differences (one way
samples and no curing distance. Hardness test. ANOVA p<0.05)
Hardness test was conduct using Vickers hardness
tester machine (Future-Tech FM-800, Japan) with load
50gf for 15 second. Indentation was done 3 times in
different areas of samples and then VHN values were
averages. Statistical analysis. One-way ANOVA
(p≤0.05) with post hoc least significant differences
(LSD) was used to compare all groups.

3. RESULT
In this study, means and SD of Vickers hardness
value on GIC groups control, GIC adding 2, 5, and
8wt% HA were 49.813±3.552 VHN, 34.492±1.825
VHN, 32.584±1.610 VHN, and 36.550±3.122 VHN
respectively as shown in table 1. Meanwhile, in
RMGIC groups data showed means and SD of Vickers Fig. 1 Vickers Hardness of GIC and RMGIC after
hardness were 22.362±2.951, 32.050±3.771, adding 2, 5, and 8wt% Hydroxyapatite from Fish
36.227±4.266, and 19.270±1.708 VHN for groups Scale.*significant difference between control-
control, RMGIC adding 2,5, and 8wt% HA, GIC+HA (post hoc LSD p<0.05); **significant
respectively. One way ANOVA analysis present that difference between control-RMGIC (post hoc LSD
there was a significant differences between groups p<0.05)
(p<0.05).
Furthermore, post hoc LSD analysis as shown in 4. DISCUSSION
fig.1, result shows that the surface hardness of GIC
was decreased after adding 2, 5, and 8wt% One of crucial mechanical properties of dental
hydroxyapatite compared to control group. The data materials is surface hardness that determines a
shows that the lowest Vickers hardness was on GIC manifestation of material’s resistance for scratching or
after adding 5wt% hydroxyapatite, and followed by abrasion. Surface hardness test seems fit to assess
2wt% and 8wt% groups. Meanwhile, the Vickers longevity and degradation of dental materials, as to
hardness value of RMGIC was increased after adding remark surface condition after storage in medium, as
2 and 5wt% HA. In other hand, it decreased after indication of wear’s resistance and durability of
adding 8wt% HA. The data shows the highest materials, and to observe setting process of materials
hardness was on RMGIC+5wt% HA, and the lowest [2,22].

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Hydroxyapatite (HA) or calcium hydroxyapatite assumed that it was hard to achieve a maximal
is a type of apatite material with the chemical formula incorporation and reaction among HA and GIC. It was
(Ca10(PO4)6 (OH)2) which is often applied in the assume that P/L ratio and particle size had effect on
medical and dental fields as substitute material due to cement hardness [23,24]. Nevertheless, Moshaverinia
its similarity structure to bone and tooth [10,16,21]. et al concluded that the main problem in HA modified
HA that produce from natural resources are better than GIC is the poor of mechanical properties of HA itself
synthetic for its biocompatibility, using uncomplicated [5].
method on production, and low cost [5,10]. Meanwhile, in RMGIC groups showed that an
Furthermore, HA from natural source can be ponder as increase of Vickers hardness after adding HA.
materials that eco-friendly, sustainable, and RMGIC has two processes on setting reaction, which
economical to process [12]. are acid-base and polymerization. These two reactions
This study used combination, which are alkaline made RMGIC have a complicated structure and to
heat treatment and calcination method to produce the reach an optimum property, manufacture’s suggestion
hydroxyapatite. The calcination method produced on duration of curing step is extremely important [6].
more crystalline HA than alkaline heat treatment. But, The liquid of RMGIC that used in this study
calcination method produced more large particles than containing HEMA (25-50%) and UDMA (1-15%)
alkaline heat treatment and less Ca/P ratio. Therefore, [26]. It was reported that there is a good interaction
we used combination method to overcome the between HEMA and HA as in physicochemical, it was
problem [12]. the Lewis acid-base interaction among HEMA as the
In theory, primary setting reaction of glass electron giver and HA as the receptor. Furthermore,
ionomer cement is in consequence of the acid-base HEMA monomer had strong acid-base interaction
reaction and following with calcium and aluminium with HA surface, so that it could act as coupling agent.
ions from glass powder to cross-linking with Thus, the existing of HEMA and HA in RMGIC can
polyacrylic acid that forms cement matrix [5]. Wilson enhance the hardness of RMGIC [27].
et al theorem that through adsorption, a complex series Moreover, RMGIC showed the highest hardness
of ion exchanges will occur among calcium ions and was on RMGIC+5wt% HA, and the lowest was
phosphate from hydroxyapatite and glass powder with RMGIC+8wt%HA. These result similar to previous
help of polyacrylate acid. This interaction will form a study which had reported that mechanical properties
layer, known as “intermediate layer”, which is highly of GIC/RMGIC would improve when adding less than
acid resistance and hard to break.16Hence, as HA add 5 wt% and will decreased if adding more than 5wt%
into GIC powder, there will be more salt bridges and [9,17,18,24]. It was reported that more than 5wt% HA
cross-linking that form as the powder mixed with exhibit congregate form within the glass ionomer resin
liquid, and these will stronger the cement and enhance matrix provide as defect sites and may reduce the
its biocompatibility. hardness. Meanwhile, over than 5wt% HA could cause
Our results presented that GIC after adding of 2, incompletely polymerization of resin component of
5, and 8wt% HA exhibit lower Vickers hardness than RMGICs and could worst its mechanical properties
control group. We assume that this finding occurred [18].
due to some factors. First, previous study suggests that
particle size has effect on GIC strength. They suggest
to use particle size that not too large or too small to 5. CONCLUSION
achieve a high strength [16,23]. It was reported that a It can be concluded that hydroxyapatite from
bigger size of HA particles has a porous surface and Nile Tilapia fish (Oreochromisniloticus) in 2, 5, and
lower crystallinity than the small ones and had effect 8wt% has effect on surface hardness of GIC and
on strength of cement [16]. Others reported that RMGIC. Adding 2, and 5wt% HA into RMGIC would
nanoparticle HA has better size in occupying void increase the surface hardness, but adding 8wt% HA
among GIC particles and act as reinforce material in decrease the surface hardness. On contrary, adding
GIC [24]. While in this study, the size of the HA into GIC decrease the surface hardness. Within
hydroxyapatite of tilapia scales is estimated only in the limitation of this study, further investigation to
micro size as the sieve only 200 mesh (74 µm). Then, evaluate the effect of hydroxyapatite on mechanical
in consequent of high bulk density of HA, previous properties of GIC and RMGIC are needed.
study recommended that to reach the fusion of HA
into GIC low P/L ratios [25]. In this study, P/L ratio
was 1:1 as manufacture suggestion and the
manipulation was done manually by spatulation, so we

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ACKNOWLEDGMENTS Journal of Ceramic Processing Research.

Financial support of this research work is given by 10(2):129, 2009
University of Sumatera Utara with Talenta USU [12] Pu’ad NASM, Koshy P, Abdullah HZ, Idris
Funded Contract 2019 Number: MI, Lee TC. Syntheses of hydroxyapatite from
4167/UN5.1.R/PPM/2019 date April 1st 2019. natural sources. Heliyon. 8;5(5); e01588, 2019
[13] Huda AN, Subuki I, Ismail MH. Synthesized
hydroxyapatite powder from clamshell via
REFERENCES chemical precipitation method. Advanced
[1] Noort R V 2007Introduction To Dental Materials Research. 911:72, 2014
Materials3rd Ed, Philadelphia Elsevier 217 [14] Asri El S, Laghzizil A, Coradin T, Saoiabi A,
[2] Anusavice KJ 2008Phillips’ Science of Dental Alaoui A, M’hamedi R. Conversion of natural
Materials 11th Ed, India Elsevier 471 phosphate rock into mesoporous
[3] Zanata RL, Navarro MF, Barbosa SH, Lauris hydroxyapatite for heavy metals removal from
JR, Franco EB. Clinical evaluation of three aqueous solution. Colloids and Surfaces A:
restorative materials applied in a minimal Physicochemical and Engineering Aspects.
intervention caries treatment approach. J 362(1-3): 33, 2010
Public Health Dent, 63 (4); 221-6, 2003 [15] Rivera EM, Araiza M, Brostow W, Castano
[4] Frencken JE, Pilot T, Songpaisan Y, VM, Diaz-estrada DR, Hernandez R,
Phantumvanit P. Atraumatic Restorative Rodriguez JR. Synthesis of hydroxyapatite
Treatment (ART): rationale, technique, and from eggshells. Materials Letters. 41(3):128,
development. J Public Health Dent, 56; 135, 1999
1996 [16] Arita K, Yamamoto A, Shinonaya Y,et.al.
[5] Moshaverinia A, Roohpour N, Chee WWL, Hydroxyapatite particle characteristics
Schricker SR. A review of powder influence the enhancement of the mechanical
modifications in conventional glass-ionomer and chemical properties of conventional
dental cements. J Mater Chem, 21;13:19, 2011 restorative glassionomer cement. Dental
[6] Sidhu SK, Nicholson JW. A review of glass Material Journal. 30(5): 672, 2011
ionomer cement for clinical dentistry. J Funct [17] Al-Hamaoy A R, Alobiedy A N, Alhille A H.
Biomater, 28;7(3); E16, 2016 Glass ionomer cement mechanical properties
[7] Bahrami-Abadi M, Khaghani M, MonshiA, enhancement using hydroxyapatite micro and
Doostmohammadi A, Alizadeh S. nano particles. ARPN Journal of Engineering
Reinforcement of glass ionomer cement: and Applied Sciences. 13(6); 1, 2018
incorporating with silk fiber. Journal Of [18] Poorzandpoush K, Omrani LR, Jafarnia SH,
Advanced Material And Processing, 4(3);14, Golkar P, Atai M.Effect of nano
2016 hydroxyapatite particles on wear of resin
[8] Moshaverinia A, Ansari S, MoshaveriniaM, modified glass ionomer by tooth brushing
Roohpor N, Darr J, Rihman I. Effects of simulation. J Clin Exp Dent. 9(3); e372, 2017
incorporation of hydroxyapatite [19] Mozartha M, Praziandithe M, Sulistiawati.
nanobioceramics into conventional glass Pengaruh penambahan hidroksiapatit dari
ionomer cements (GIC). Acta Biomateralia, cangkang telur terhadap kekuatan tekan glass
4:432, 2008 ioomer cement. Jurnal B-Dent. 2(1); 75, 2015
[9] Sayyedan F.S, Fathi M.H, Edris H, [20] Mawadara PA, Mozartha M. Pengaruh
Doostmohammadi A, Mortazari V, Hanifi A. penambahan hidroksiapatit dari cangkang telur
Effect of forsterite nanoparticles on mechanical ayam terhadap kekerasan permukaan GIC.
properties of glass ionomer cement. Ceramics Jurnal Material Kedokteran Gigi.5(2); 8, 2016
International. 40: 10743, 2014 [21] Pon-On W, Suntornsaratoon P,
[10] Akram M, Ahmed R Shakir I, Wan Ibrahim Charoenphandhu N, Thongbunchoo J,
WA, Hussain R. Extracting hydroxyapatite and Krishnamra N, Tang M. Hydroxyapatite from
its percursors from natural resources. Journal fish scale for potential used as bone scaffold or
Material Science. 49 (4);1461, 2014 regenerative material. Materials Science and
[11] Bahrololoom ME, Javidi M, Javadpoura S, Ma Engineering C. 62;183, 2016
J. Characterisation of natural hydroxyapatite [22] Shintome LK, Nagayassu MP, Di Nicolo R,
extracted from bovine cortical bone ash. Myaki SI. Microhardness of glass ionomer
cement indicated for ART technique according

9
 Advances in Health Sciences Research, volume 32

to surface protection treatment and storage

time. Braz Oral Res. 23; 439, 2009
[23] Bala O, Arisu H D, Yikilgan I, Arslan S, Gullu
A. Evaluation of surface roughness and
hardness of different glass ionomer cements.
Eur J Dent. 6(1); 79, 2012
[24] Rahman I A, Masudi S M, Luddin N, Shiekh R
A. One-pot synthesis of hydroxyapatite-silica
nanopowder composite for hardness
enchancement of glass ionomer cement (GIC).
Bulletin Materrials Science. 37; 213, 2014
[25] Nicholson JW, Hawkins SJ, Smith JE. The
incorporation of hydroxyapatite into glass
polyalkenoat (“glass-ionomer”) cements: a
preliminary study. Journal of Material Science:
Materials in Medicine. 4; 418, 1993
[26] Safety data sheet GC Fuji II LC.
http://www.gcamerica.com
[27] Arcis RW, Lopez-Macipe A, Toledano M.
Mechanical properties of visible light-cured
resin reinforced with hydroxyapatite for dental
restoration. Dental Materials. 18; 49, 2002

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