hydroxyapatite biomaterials are materials that are very widely used in several health purposes , including as a source of calcium for the manufacture of toothpaste and as an important material in the formation / bone repair .
the chemical properties of hydroxyapatite are bioactive and compatible with the adjacent bone and teeth .
hydroxyapatite is a calcium phosphate ceramic that is totally biocompatible and nontoxic and becomes an integral part of living bone and teeth tissue [ 18 ] .
the raw material for the production of hydroxyapatite biomaterial is very easily available and abundant in indonesia .
the production process was easy and the cost is also relatively inexpensive if done on a large scale . among
the abundant raw materials are the shells of crabs , which are one of indonesia 's main export commodities .
export of crabs commodity by indonesia amounted to 604.215625.000 tons / year without the shell form , while domestic consumption is expected to be so much more , as in makassar , carrying 292.5 tons exported in the form of crab without shell with the main export destination being singapore .
if the mass of crab shells 2550% of the total mass , it can be estimated that in 2012 produced the shells of crabs around 151,053.75 to 302,107.5 tons in indonesia and 73.125 to 146.25 tons in makassar ( a region of indonesia ) , there was just from crabs production were exported .
this value is of course even more if the crab 's consumptions in the country are also taken into account .
this suggests the existence of crab shells is abundant in indonesia , including in makassar .
as known in indonesia , the shells of crabs have not been used , so it will only be a waste disturbing environment [ 7 , 8 ] .
crabs shells containing calcium carbonate ( caco3 ) are very abundant ; amount 4070% , varies according to the species .
calcium carbonate can be further processed into calcium hydroxyapatite [ ca10(po4)6(oh)2 ] [ 4 , 5 ] . according to strassler ,
hydroxyapatite is one of the active ingredient materials that are widely used in toothpaste products for protection against teeth demineralization [ 1113 ] .
the crystals structure of hydroxyapatite will be better by using cao as a precursor of calcium .
however , the use of these compounds also produces carbonate apatite [ ca10(po4)6co3 ] in a fairly large percentage .
this is because the calcination process can not completely eliminate carbon dioxide ( co2 ) in caco3 so that there can be reaction with the precursor phosphate .
however , the carbonate apatite heating at a temperature of 700900c for 25 hours , followed by washing using distilled water , the carbonate apatite can be hydroxyapatite [ 1113 ] . the instrument were used in this research were glass apparatus , ohaus analytical balance , petri dish , porcelain cup , buchner flask , buchner funnel , vacuum pump sargent - welch co. model 1400 , magnetic stirrer , magnetic bar , hotplate idealife , ph meter , furnace thermolyse 6000-barnstead , dessicator , thermometers , spnisosfd oven , stopwatch , shimadzu x - ray diffractometer ( xrd ) model 6000 , x - ray fluorescence ( xrf ) shimadzu , fourier transform infra - red ( ftir ) prestige-21 shimadzu , scanning electron microscopy combined with the ability to generate localized chemical information ( sem - edxa ) variant , and uv - vis shimadzu model 6105 .
this is because the carbonate ion compounds are able to inhibit the crystallization process ca10(po4)6(oh)2 so that the results will be dominated by an amorphous phase .
if the caco3 burned at temperature calcinations , decomposition reaction of caco3 into cao will occur and co2 emissions are dominant and will be issued as a result of the combustion reaction [ 1315 ] .
waste of crabs shells portunus pelagicus species was taken from an exporter crabs company in industry areal , makassar city , south sulawesi province , indonesia .
wahidin hospital , and ( nh4)2hpo4 was obtained from fluka chemical , ch3cooh glacial 100% , ch3coona3h2o from merck , ( nh4)6mo7o244h2o from aldrich , nh4vo3 from aldrich , hno3 and naf from fluka chem . , ( nh4)2c2o4 , distilled water , aluminum foil , and whatman filter paper numbers 40 and 42 . the instruments were used in this research that glass as commonly used in laboratories , ohaus analytical balance , petri dish , porcelain cup , buchner flask , buchner funnel , vacuum pump sargent - welch co. model 1400 , magnetic stirrer , magnetic bar , hotplate idealife , ph meter , furnace thermolyse 6000 barnstead , dessicator , thermometers , oven spnisosfd , stopwatch , shimadzu x - ray diffractometer ( xrd ) model 6000 , x - ray fluorescence ( xrf ) shimadzu , fourier transform infrared ( ftir ) prestige-21 shimadzu , scanning electron microscopy combined with the ability to generate localized chemical information ( sem - edxa ) variant , and uv - vis shimadzu model 6105 .
crabs shells portunus pelagicus waste was cleaned with distilled water and dried at room temperature . furthermore , to transform crabs shells into caco3 and then into cao , calcinations were performed on the samples at 1000c temperature for 5 hours at a rate of temperature rise 5c / minute .
calcium oxide ( cao ) obtaining dominated as result of calcinations and then made suspensions in 100  ml of distilled water with a calcium concentration of 0.3  m. the suspensions reacted by dropwise with a 100  ml 0.2  m of ( nh4)2hpo4 solution through the coprecipitation method , at temperatures around 40c while the solution was stirred for 25 hours .
the precipitation allowed stand overnight or 24 hours at room temperature , and the precipitate is filtered with a whatman filter paper number 40 and dried at 110c for 5 hours .
the pure hydroxyapatite obtained by sintering to the dried precipitate at various temperatures of 500900c for 4 hours [ 16 , 17 ] .
the characterization of the compounds was performed by using x - ray diffraction ( xrd ) , ftir , and sem - edxa .
the proven in vitro demineralization of teeth was conducted through evaluating the concentration of phosphate in solution by using the uv - vis spectroscopy .
the effectiveness of hydroxyapatite to protection against teeth demineralization was tested in acetate buffer ph 5.0 solutions , with 1 m of acetic acid concentration with the addition of hydroxyapatite in varying concentrations and immersion time .
each of 5 beakers was filled with 300  ml of acetate buffer ph 5.0 with acetic acid concentration of 1  m. an acetate buffer was left without adding anything as a comparison .
an acetate buffer was then added 10  ppm of naf left without addition of ca10(po4)6(oh)2 .
other three pieces beaker of acetate buffer containing 10  ppm of naf are adding of ca10(po4)6(oh)2 with variation concentration of 25  ppm , 50  ppm , and 100  ppm .
the immersed times of tooth samples in solution are 3 , 6 , 9 , 24 , and 48 hours , respectively .
furthermore , the phosphate concentration in each solution was measured by using uv - vis spectrophotometer with the wavelength for phosphate ( maks ) being 432  nm .
the calcinations aim to eliminate the organic component in the shells of crabs and convert caco3 compound which is the dominant compound in the shells of crabs into cao through the elimination of co2 in gas form . characterizing the calcinations results
the results can be seen from the diffraction pattern of the crab shells before and after calcinations at a temperature of 1000c for 5 hours ( figure 1 ) . in this figure
can be observed changes in the diffraction pattern of crab shells , where the change in the diffraction pattern is due to the chemical change from mixture of caco3 with organic matter to pure cao .
the appearance of the sharper peaks in figure 1 after calcinations ( b ) is a result of the crystallinity of the cao .
identification by ft - ir as shown in figure 2 showed there is a reduction process of -co3 groups and some of ir - spectra were missing after calcinations .
the elimination of -co3 groups and organic components can also be seen from the data of mass reduction of sample calcinations .
the determinations of calcium contained in sample was conducted by using x - ray fluorescence , where obtained calcium is 66.62% after calcinations .
these results are then used to calculate the stoichiometry in determining the number of results calcinations which is needed to react with ( nh4)2hpo4 as the precursor phosphate .
the precipitation reactions aiming to produce ca10(po4)6(oh)2 used phosphate , ( nh4)2hpo4 , as the precursor and then reacted with cao as calcinations results .
side results ca10(po4)6co3 also occur as a product of reaction of ( nh4)2hpo4 and caco3 presence in the calcinations results .
dried precipitate further sintered on temperature variations of 4001000c for 2 hours ; it is intended to determine the optimum temperature where the ca10(po4)6(oh)2 is formed , furthermore characterized by using xrd , ft - ir , and sem - edxa .
the diffractograms of each sintering results compounds indicate that the temperature is closely related to the formation of crystals .
this is due to the nature of the vibrating atoms moving faster in higher temperatures .
the optimum temperature formation of hydroxyapatite was determined by calculation of the probability of the sample phase from the xrd results analysis according to jcpds standard data , which , jcpds ; 24 - 0033 is standard data for ca10(po4)6(oh)2 ; 09 - 0169 for -ca3(po4)2 ; 29 - 0359 for -ca3(po4)2 , 35 - 0180 -ca3(po4)2 ; 35 - 0180 for ca10(po4)6co3(oh)2 and 19 - 0272 standard data for ca10(po4)6co3(oh)2 .
figures 3 and 4 showed that the temperature is closely associated with the formation of hydroxyapatite phase . in the both graphs it can be seen that the maximum intensity of the phase formation of hydroxyapatite has been found by sintering at temperature of 800c , it means the optimum temperature of hydroxyapatite formation is 800c , and then this result will be used for another application . the formations of hydroxyapatite phase had been dominated at 800c confirmed by percentage probability sample phase ( figure 5 ) , in which the percentage of hydroxyapatite phase formed around 46.61% , while phases of -ca3(po4)2 and phase -ca3(po4)2 are 17.76% and 19.37% , respectively .
however , also there is still presence of a phase ca10(po4)6co3 and ca10(po4)6co3(oh)2 with a range of 11.84% and 4.43% , respectively , which indicates the presence of carbonates .
the x - ray diffractograms of ca10(po4)6(oh)2 synthesized at a 800c temperature sintering can be seen in figure 6 , where the peaks of ha are symbolized by the peak of the crystal ca10(po4)6(oh)2 , while peak -tkf is symbolized crystalline peaks of -ca3(po4)2 , the symbol -tkf for crystalline -ca3(po4)2 , aka for ca10(po4)6co3 , and akb for ca10(po4)6co3(oh)2 .
the highest intensity peak at 31.2572  deg corresponding to crystalline -ca3(po4)2 is seen , the second highest peak intensity is 31.7783 , and the third highest peak intensity is 28.0565 crystals suitable for ca10(po4)6(oh)2 .
ftir results showed that the sintering temperature variation affects the absorption band shapes which generally all sintering results showed absorption band of -oh , absorption band 1 , 2 , 3 , and 4 of po4 , and co3 groups .
infrared spectra in figure 7 show the -oh groups at 633  cm which are characteristic of hydroxyapatite   appearing on the sintering temperatures of 4001000c .
additionally spectrum also showed higher sintering temperature causing the sharper peaks phosphate group ( po4 ) because the nature of the vibrating atoms moves faster at higher temperatures .
the ft - ir spectra of sample were sintered at a temperature of 800c ( figure 8) showing that hydroxyapatite is the dominant compound formed .
the stretching frequencies of po4 group are indicated by 1120.64  cm , 1091.71  cm , 1043.49  cm ( 3 ) , 993.34  cm , 877.61  cm ( 1 ) , 603.72  cm , 565.14  cm , ( 4 ) , and 370.33  cm ( 2 ) . and
a sharp spectrum in the area of 3570.24  cm indicates the presence of free -oh and 3427.51  cm indicating -oh bounded , and this is indicating that the dominant compound is ca10(po4)6(oh)2 . while area of 1654.92  cm , 1458.18  cm , and 1421.54  cm indicates the presence of carbonate groups ( co3 ) it can be identified as ca10(po4)6co3 and ca10(po4)6co3(oh)2 which has not been transformed into ca10(po4)6(oh)2 during the sintering process .
the results of characterization by sem - edxa shown in figure 9(a ) show that the size of hydroxyapatite formed from the synthesis tends to be small and only a few are large .
while figure 9(b ) shows that the surface of hydroxyapatite is smooth and nonporous , this shows that hydroxyapatite which has synthesis from crab shells can function well as inhibiting tooth demineralization . while in figure 10 the edxa spectrum shows the composition of the synthesis yield
was dominated by oxygen ( o ) up to 59.52% , calcium ( ca ) up to 23.76% , and phosphorus ( p ) up to 13.32% .
demineralization of teeth is a process of decomposition of the crystal of ca10(po4)6(oh)2 due to the acidic conditions by releasing ca and po4 ions .
demineralization of tooth causing increased levels of ca and po4 in saliva in direct contact with the tooth . in vitro
, the rate of tooth demineralization can be observed through the concentrations of ca and po4 ions in solutions where the tooth was soaked each unit of time .
therefore , increase of po4 ions concentrations in solution a soaked gear can be one of indicators to measure the rate of tooth demineralization . 

figure 11 shows the relationship between the soaking time of teeth versus the increase of the ion po4 levels in solution where the tooth was soaked as well ; it appears that with the increasing addition of ca10(po4)6(oh)2 into the acetate buffer equals to the rate of demineralization decrease .
it is can be altered by the amount of po4 ions in solutions ; however the addition of ca10(po4)6(oh)2 ions showed lower amount of po4 ions compared to solutions without the addition of ca10(po4)6(oh)2 .
this proves that the ca10(po4)6(oh)2 were synthesized from the crab shell effective for protection against tooth demineralization .
the decrease in the rate of tooth demineralization with the addition of ca10(po4)6(oh)2 can also be observed through analyzing the tooth mass reduction in the fifth variation of acetate buffer solution as shown in table 1
. the greater concentrations of ca10(po4)6(oh)2 in the acetate buffer solution were teeth immersed exhibit the smaller mass of teeth in the solution . 

based on these results it is concluded as follows.waste of shells crabs ( portunus pelagicus ) proved to be used as raw material for the synthesis of ca10(po4)6(oh)2 due to high calcium levels which amounted to 66.62% in addition to the abundant existence as waste.the optimum temperature formation of ca10(po4)6(oh)2 is at 800c.ca10(po4)6(oh)2 were synthesized from waste of shell crabs ( portunus pelagicus ) in vitro effectively inhibiting the rate of demineralization of the tooth where the greater the addition of ca10(po4)6(oh)2 in the solution , the more the inhibiting demineralization of the tooth or the smaller the rate of tooth demineralization in solution .
waste of shells crabs ( portunus pelagicus ) proved to be used as raw material for the synthesis of ca10(po4)6(oh)2 due to high calcium levels which amounted to 66.62% in addition to the abundant existence as waste .
ca10(po4)6(oh)2 were synthesized from waste of shell crabs ( portunus pelagicus ) in vitro effectively inhibiting the rate of demineralization of the tooth where the greater the addition of ca10(po4)6(oh)2 in the solution , the more the inhibiting demineralization of the tooth or the smaller the rate of tooth demineralization in solution .
 crab 's shells of portunus pelagicus species were used as raw materials for synthesis of hydroxyapatite were used for protection against demineralization of teeth . 
 calcination was conducted to crab 's shells of portunus pelagicus at temperature of 1000c for 5 hours . 
 the results of calcination was reacted with ( nh4)2hpo4 , then dried at 110c for 5 hours . 
 sintering was conducted to results of precipitated dried with temperature variations 4001000c for a hour each variation of temperature then characterized by x - ray diffractometer and ftir in order to obtain the optimum formation temperature of hydroxyapatite is 800c . the hydroxyapatite is then tested its effectiveness in protection against tooth demineralization using acetate buffer ph 5.0 with 1  m acetic acid concentration with the addition of hydroxyapatite and time variation of immersion . 
 the results showed that the rate of tooth demineralization in acetate buffer decreased significantly with the provision of hydroxyapatite into a solution where the addition of the magnitude of hydroxyapatite is greater decrease in the rate of tooth demineralization . 
1. Introduction
2. Experimental
3. Results and Discussion
4. Conclusion