37

CHAPTER - II

REVIEW OF THE LITERATURE

2.1 SYNTHESIS OF HYDROXYAPATITE

Bone is a multiphase composite with main constituents of bone collagen

matrix and assembled hydroxyapatite [Ca10(PO4)6(OH)2, HAP], in which HAP is the

major inorganic constituent [84]. The synthetic HAP is widely used in various

biomedical applications in the form of nano powders, films, scaffolds and coatings

[85]. It has attracted the attention of researchers from the past 30 years as filler and

an implant material because of its excellent biocompatibility and bioactivity with

human bone and teeth [86-88]. The chemical species constituting HAP

(Ca, P, O and H) are accepted as non-toxic elements. Moreover, their physical

properties such as fracture toughness and fracture strength, etc., depend on the

structures, compositions and sizes. The HAP of special shape and size could be used

as a seed to induce the directional growth of the hydration products, so as to

reinforce the strength of the calcium phosphate cement greatly [89].

Nowadays nanomaterials have wide-range of applications in a variety of

areas including chemistry, physics, electronics, optics, materials science and

biomedical sciences [90]. Therefore, the development of the synthesis protocols for

nanomaterials over a range of chemical compositions constitutes a steadily evolving

branch of nanotechnology. Hence, researchers have tried to customize its properties

such as bioactivity, morphology and particle size [91,92]. The chemical, structural

and morphological properties of synthetic hydroxyapatite can be modulated by

38

varying the method and conditions of synthesis. So, the synthesis of HAP for

various applications is being carried out by many researchers.

There are a number of innovative dispensation routes for the synthesis of

hydroxyapatite powders including the precipitation method, ultrasonic irradiation

technique, microwave synthesis, sol-gel method, molten salt method, hydrothermal

technique, freezing method, mechano-chemical synthesis and template method.

2.1.1 Precipitation Method

Tas et al., [93] reported that the synthesis of nano-sized (~ 50 nm),

homogeneous and high-pure HAP ceramic powder from calcium nitrate tetrahydrate

and diammonium hydrogen salts which were dissolved in modified simulated body

fluid (SBF) solutions at 37 °C and pH 7.4 using a novel chemical precipitation

technique. There was no decomposition of HAP into the undesired β-TCP phase

even after heating at 1600 °C in air for 6 h. They observed the superior

high-temperature stability of biomimetic HAP powders.

The research on the synthesis of hydroxyapatite particles using precipitation

method have been reported by Khopade et al., [94]. Although the benefits of low

cost and simplicity of precipitation techniques have provided a direct avenue for

HAP synthesis, the deviation from normal HAP phase can be significant with these

techniques [74,95].

Synthesis of HAP particles using precipitation method have been reported by

Patric et al., [96] has shown that the HAP particle size decreases with increasing

temperature. Liu et al., [97] synthesized HAP nanorods at pH of 4.5 in the presence

of suitable surfactant via wet chemical technique at low temperature. The

39

as-synthesized nanorods are found to be pure with the diameter of 50-80 nm and

length of 0.2-1.2 µm. There were no impurities obtained like carbonated HAP

during the process.

Sung et al., [98] reported the synthesis of HAP nano powders using a

modified chemical precipitation route. They explained that the dried HAP powder

was almost in amorphous state with very low crystallinity, showing fine particle size

of ~50-100 nm. Due to the enhanced surface area of HAP nano powder showed high

sintered density at 1000 °C. The Ca/P ratio in powder preparation was varied and the

amount of each HAP and β-TCP phase was analyzed. The powder with a Ca/P ratio

of 1.70 showed the formation of the lowest β-TCP content, while that of 1.75

showed the formation of a CaO phase as well as β-TCP. By using a modified

chemical route and varying chemical composition and sintering β-TCP phase

formation behavior of HAP powder could be controlled for artificial hard tissue

applications.

A process for the synthesis of HAP nanocrystals by a wet chemical method

at low temperature in an aqueous medium was reported by Murugan et al., [99].

Further its chemical and crystallographic functionalities were quite similar to

biological apatite. The in-vitro bioresorbability of the nano-HAP is higher than

conventional HAP and close to biological apatite, which can be attributed to its

surface area owing to nanostructure processing. The smoothness of the nano-HAP

would not damage or harm any living biological organism upon implantation.

Yun et al., [100] have synthesized nanocrystalline HAP by chemical

precipitation method with the aid of ultrasonic irradiation using calcium nitrate and

ammonium dihydrogen phosphate as source materials and carbamide (NH2CONH2)

40

as precipitator. The crystallinity and morphology of the resulting nanoparticles are

dependent on the Ca/P ratio, concentration of Ca2+

, ultrasonic power and synthetic

temperature. When the concentration of Ca2+

exceeded 0.2mol/L and the ultrasonic

power was lower than 300 W, the monophase of HAP powder could not be obtained.

When the reactive temperature and Ca/P ratio increase from 313 to 353 K and 1.67

to 2.5, respectively the as-prepared crystallites exhibited a preferential growth along

the (002) direction and the nanoparticles showed an acicular morphology. Spherical

nanoparticles could be obtained at high synthetic temperature (353K), ultrasonic

power (300 W) and high Ca/P (2.0-2.5). In addition, the crystallite size of the HAP

nanoparticles decreases with the decrease of [Ca2+

] ions and the increase of synthetic

temperature and ultrasonic power.

Pramanik et al., [101] have studied the synthesis of nanosized HAP by a

precipitation technique at room temperature using an aqueous solution of calcium

nitrate and diammonium hydrogen phosphate as starting materials in presence of

various capping agents like triethanolamine, ethylenediamine tetraacetic acid,

diethanolamine and ethylene glycol separately. The capping agents effectively

restrict the nuclei growth of the HAP. The n-HAP powders synthesized with various

capping agents showed crystallites with small particle size and less agglomeration

compared to the n-HAP synthesized without using any capping agent. This

technique may be used for bulk preparation of nanoHAP powder with high specific

surface area and high aspect ratio.

Wang et al., [102] reported the synthesis of hydroxyapatite nanoparticles by

using chemical precipitation method using various organic modifier such as

polyethylene glycol, tween 20, trisodium citrate and D-sorbitol, they found that

41

polyethylene glycol was beneficial for the formation of HAP nanorods with a larger

aspect ratio (average length/average diameter) at high synthesis temperature and

Tween 20 and trisodium citrate favoured the formation of small-sized HAP

nanorods, and D-sorbitol helped the formation of HAP nanorods with long length at

low synthesis temperatures. In their report they also proved that the crystallinity of

the resultant HAP nanorods increased with increase of the synthesis temperature.

The conversion of hydroxyapatite powders into HAP whiskers by using the

refluxing method through the dissolution–reprecipitation process was reported by

Seo et al., [103] in which the amorphous reprecipitates were initially formed in the

Ca(EDTA)2-PO43-

mixed solution and have continuously grown into long and thin

HAP whiskers. It is obvious that the morphology of the final whiskers is altered by

the concentration of H2O2, pH of the starting solution, and refluxing temperatures.

By increasing the H2O2 concentration, pH value and refluxing temperature,

well-shaped whiskers with clean and smooth surfaces were formed. The higher

value of pH was also responsible for the formation of monodispersed HAP whiskers.

Whiskers with high-aspect ratio were obtained from the starting reaction solution of

pH = 9 at the relatively low temperature of 100 °C.

Cengiz et al., [104] have synthesized hydroxyapatite nanoparticles using CaP

tris solution as a calcium phosphate medium by precipitation method. Their results

fit very well with the reference and the crystallite they synthesized in the presence of

CaP tris solution varied in a range of very fine size from 15.88 to 16.12 nm. In terms

of (d002/d300) ratio, which is a measure of uniformity of crystallites, the ratio was

0.99 for the particle synthesized using CaP tris as a medium. As the powder

(nanoparticles) produced from SBF contains large fraction of needle-shaped

42

particles, nanoparticles synthesized in the presence of CaP tris medium almost

consist of desultory structure. There is an acceptable agreement between the results

for CaP tris. They concluded from the results of HAP particles (CaP tris) the length

and width of the particles were less than 500 and 100 nm, respectively and the

composition of the final product produced from CaP tris (1.58) is closer to the ideal

HAP (1.67). Therefore it showed that CaP tris solution can act as a medium to

synthesize HAP nanoparticles.

Shanthi et al., [105] have obtained nano-crystalline HAP rods by

co-precipitation method using cationic surfactant CTAB as template, at ambient

temperature and pressure. The temperature controls like hydrothermal treatment or

refluxing process, which hurdle the bulk production was eliminated. HAP rods with

diameter by 20 nm and length in the range of 100-200 nm were obtained.

Calcium nitrate and tri ammonium phosphate as starting materials for the

preparation of water-dispersible HAP nanorods was reported by Tan et al., [106].

They prepared HAP nanorods with length of 300-400 nm and width 40-60 nm by

chemical precipitation at 90 °C. The sodium citrate used in the reaction medium

improved the colloidal stability of HAP dispersions by adsorbing citrate ions on

HAP surface.

Catros et al., [107] have synthesized nano-HAP using wet chemical

precipitation method at room temperature. The morphological, physico-chemical

and crystallographic analysis revealed the specific features of HAP. Biological

in-vitro experiments revealed that the high affinity and proliferative ability of MG63

cells cultured on to the material.

43

Wu et al., [108] in their study reported the synthesis of mesoporous

nano-hydroxyapatite (n-HAP) particles by low-temperature co-precipitation method

in the presence of CTAB. The cationic surfactant of CTAB was used as a template

to regulate n-HAP crystal nucleation and growth. The results show that the

synthesized particles have the features of high pure phase, low crystallinity and

mesoporous structure. The ratio of surfactant effectively influences the mesoporous

structure of n-HAP particles, including the surface area, the pore volume and the

pore size. The adsorbed amount of Bovine Serum Albumin (BSA) on n-HAP

increases with the specific surface area and the pore volume, and the release rates of

BSA are different due to the different pore sizes and pore structures. The n-HAP

particles synthesized with 0.5 % CTAB exhibited the highest BSA loading and the

slowest release rate due to its highest specific surface area and the smaller pore size,

indicating that it has optimal mesoporous structure for good loading and well

controlled release of BSA. These mesoporous n-HAP materials demonstrate a

potential application in the field of protein delivery due to their bioactive,

biocompatible and mesoporous properties.

2.1.2 Ultrasonic Irradiation Technique

Ultrasound irradiation can be utilized to synthesize bioceramic materials

such as hydroxyapatite too. It was well demonstrated by synthesizing hydroxyapatite

from the hydrolysis of calcium oxy phosphates by Fang et al., [109]. The reaction

time required to prepare hydroxyapatite from a mixture of Ca4(PO4)2O and

CaHPO4(H2O) (brushite) and 38 ± 0.5 °C was reduced from 3 h without sonication

to 15 min with sonication. The morphologies of these two products were however

quite different. Meissner et al., [110] in their studies have observed that the size and

44

morphology of HAP can be controlled and are dependent on the precipitation and

ultrasonic power. Kim and coworkers have also reported the synthesis of

hydroxyapatite particles from H3PO4 and Ca(OH)2 using sonication has better

thermal behavior than the conventional counterparts [111]. Further, the morphology

of the resulting powders could also be conveniently controlled by this method.

Gopi et al., [112] have reported the synthesis of hydroxyapatite nanoparticles

by ultrasonic coupled sol-gel method. They found that the HAP synthesized by this

method possesses an excellent purity and the particle sizes are nano sized.

Gopi et al., [113] have also reported the synthesis of hydroxyapatite

nanoparticles by a novel ultrasonic assisted mixed template directed method. In this

method glycine–acrylic acid (GLY–AA) hollow spheres were used as an organic

template which could be prepared by mixing of glycine with acrylic acid. The effect

of ultrasonic irradiation time on the crystallinity and size of the HAP nanoparticles

in presence of glycine–acrylic acid hollow spheres template were investigated. From

the inspection of the above results it is confirmed that the crystallinity and size of the

HAP nanoparticles decrease with increasing ultrasonic irradiation time.

2.1.3 Microwave Synthesis

Hydroxyapatite powders prepared by reacting calcium chloride and

diammonium hydrogen phosphate in three types of cyclohexane, surfactant

(NP5+NP9), aqueous phase structure: namely, bicontinuous microemulsion, inverse

microemulsion and emulsion were reported by Lim et al., [114]. Both the

bicontinuous and inverse micro emulsions result in the formation of a nanosized,

highly sinterable HAP powder. They were sintered to a relative density of >95 %

45

theoretical density at 1000°C, which is more than 100 °C lower than those reported

microemulsion-derived HAPs also exhibit a higher sintered density and are more

refined in grain size than for that of the emulsion-derived one when sintered at

1200 °C for 2 h. Only a trace amount of β-tricalcium phosphate was detected in the

sintered HAP prepared from the inverse microemulsion composition.

Carbonated hydroxyapatites were synhthesized by the substitution of calcium

carbonate for calcium hydroxide during the reaction with diammonium phosphate

under microwave irradiation was reported by Macipe et al., [115]. Biphasic calcium

phosphate (BCP) ceramics consisting of a mixture of HAP and tricalcium phosphate

(TCP) having various Ca/P ratios synthesized by means of microwave irradiation

using Ca(OH)2 and (NH4)2HPO4 and determined the thermal stability of the resulting

BCP have been reported by Manjubala et al., [116]. Direct synthesis of HAP

nanoparticles by using microwave with different morphologies have also been

prepared and reported by Liu et al., [117].

Yang et al., [118] reported the thermally stable hydroxyapatite poly crystals

prepared by using H3PO4, glucose and Ca(NO3)2.4H2O, as starting materials with

microwave irradiation and co-precipitation method. The results showed that the

various parameters such as aging time, microwave irradiation power and time have

significant effects on thermal stability of HAP.

Microwave heating used to prepare whisker shaped hydroxyapatite by the

hydrolysis reaction under conditions of 70-90 °C, 4-15 h, pH = 1 of solid state

reacted α-TCP was reported in Yoon et al., [119]. Microwave heating increased the

hydrolysis rate and enhanced the development of whisker shaped morphology in the

HAP. The optimum hydrolysis reaction time at 90 °C was ~10 h and was found to be

46

significantly shorter compared with conventional heating (24 h). The morphology

of the whisker-like phase could be controlled by means of adjustments to the

reaction temperature or time.

Single crystal hydroxyapatite nanorods have been synthesized by

Sun et al., [120] by the reverse microemulsion method. They explained the

feasibility of using quaternary reverse microemulsion (TX-100/CTAB/n-butanol+

n-hexanol/ cyclohexane/water) in the preparation of HAP nanorods with diameter

8-15 nm and length 25-50 nm. The presence of the alcohol in the microemulsion and

the double stabilization function of the mixed surfactant on the interfacial film are

important factors in regulating the particle size, which is acting on the particles

growth by influencing the flexibility of the interfacial film.

The synthesis of stoichiometric single crystal HAP nano rods with mono

dispersion and narrow-size distribution have been obtained by Lin et al., [121] using

hydrothermal microemulsion synthesis. They reported the HAP nano rods with

diameter 24-40 nm and length 55-350 nm. The homogeneity in size and shape of the

HAP nano rods was probably attributed to the nano-reactors and the soft template of

the surfactants, and the high crystallization of the products was attributed to the

hydrothermal treatment, which exhibits high mechanical properties.

Powders of spherical BCP powders with an average size in the range of

50-90 nm have also been synthesized and reported by Lee et al., [122] and fabricated

by microwave heating. This was achieved by heating the aqueous solution

containing of Ca(OH)2 and H3PO4 at 300 °C for about 20 min. Qi et al., [123] have

reported a microwave assisted hydrothermal method for the preparation of HAP

47

nanowires without any surfactant and the effects of temperature and time on the

morphology of the produced was also investigated.

Nanosized calcium phosphate powders have been synthesized by an inverse

micro emulsion system using mixtures of kerosene as oil phase, Aliquot 336 as

cationic surfactant and Tween 20 as non-ionic surfactant and aqueous solutions of

calcium nitrate tetrahydrate and biammonium hydrogen phosphate as the water

phase reported by Singh et al., [124]. They explained that the nature of the

surfactants played an important role to regulate the size and morphologies of the

calcium phosphate nano particles. The cationic surfactant Aliquot 336 has been

found to regulate the nucleation and crystal growth. It is also evidenced that the

nature and concentration of the surfactant and the concentration of the precursors

strongly influenced on the growth of this biomaterial. The used materials used by

them were inexpensive and environment friendly.

Cheng et al., [125] reported the synthesis of calcium phosphate porous

bioceramics by microemulsion method. They found that TCP, CDHA and pure HAP

can be obtained at the beginning of the synthesis by controlling the various

experimental parameters such as pH of the reaction, the Ca/P ratio, the heat-treated

temperature and the addition of some glycerol. They concluded that the pH values

and Ca/P ratios are seems to be critical for the formation of apatite from their

experimental investigation. The solid-to-liquid ratio could largely influence the size,

morphology and porosity of the beads made.

Saha et al., [126] have synthesized hydroxyapatite nanopowder by reverse

microemulsion technique using calcium nitrate and phosphoric acid as starting

materials in aqueous phase. Cyclohexane, hexane, and isooctane were used

48

as organic solvents, and dioctyl sulfosuccinate sodium salt (AOT), dodecyl

phosphate (DP), NP5 (poly (oxyethylene)5 nonylphenol ether), and NP12

(poly(oxyethylene)12 nonylphenol ether) as surfactants to make the emulsion. Effect

of synthesis parameters, such as type of surfactant, aqueous to organic ratio, pH and

temperature on powder characteristics were studied. It was found that the surfactant

templates played a significant role in regulating the morphology of the nanoparticle.

Hydroxyapatite nanoparticle of different morphologies such as spherical, needle

shape or rod-like were obtained by adjusting the conditions of the emulsion system.

Arami et al., [127] in their studies have observed the rapid formation of

hydroxyapatite nano strips using CTAB as a nucleation and growth controlling agent

by microwave irradiation method, without the occurrence of common

crystallographic transformations including dissolution and slow recrystallization.

The obtained nano strips have an average width of about 10 and 55 nm. During the

mixing of the CTAB with phosphate precursor, it formed rod-like micelles with

PO43-

groups resulting in the formation of CTAB-HAP complexes and 1D nano

structures during the microwave treatment. In this method the formation of highly

pure and well-crystallized HAP can be obtained during the short-time microwave

irradiation.

García et al., [128] in their work reported hydroxyapatite nanoparticles using

a hydrothermal microemulsion technique with calcium nitrate tetrahydrate and

diammonium hydrogen phosphate as precursors is a new microemulsion with

(CTAB)/toluene/n-butanol/water as a nanoreactor hydroxyapatite particles with an

average size close to 60 nm were obtained at relatively low values of the

microemulsion parameters (W0 = 10 and P0 = 5) while particles with an average size

49

close to 100 nm could be obtained at relatively high values of the microemulsion

parameters (W0 = 45 and P0 = 5).

2.1.4 Sol-gel Method

Sol-gel method was attempted by Layrolle et al., [129]. This process is also

becoming a common technique for the production of ultra fine and pure ceramic

powders, fibers, coatings, thin films and porous membranes. More recently, this

method has been extensively developed and used in biotechnology applications also.

Takahashi et al., [130] developed a sol-gel route for the synthesis of HAP using

Ca(NO3)2.4H2O and phosphonoacetic acid [HOOCCH2PO(OH)2] in an aqueous

solution at 700 °C. The Crystallinity was improved with the increasing of

temperature up to 1100 °C.

The sol-gel approach is an effective method for the preparation of HAP

[131-135] and it offers the control over the formation of particular phases and the

purity of the formed phase, and allows processing at low temperature. This process

involves a low-temperature using chemical precursors that can produce bioceramic

materials with better purity and homogeneity.

The synthesis of HAP using a novel sol-gel technique was reported by

Bezzi et al., [74]. By this method pure HAP powder can be synthesized. The

process included a thermal treatment at 800 °C, which reduces the potentiality

arising from nanometric powder. The compositional, microstructural, morphological

and mechanical characterizations were carried out on the powder as well as on dense

and porous sintered bodies.

50

Han et al., [136] have synthesized nano crystalline HAP powders using

calcium nitrate, diammonium hydrogen phosphate and citric acid at a low

temperature of 750 °C by citric acid sol-gel combustion method. The grain size of

HAP powder is between 80 and 150 nm and the effective diameter is 494.6 nm. The

sinterability was affected by agglomeration of HAP powder and decomposition. The

open porosity of resulting HAP ceramic is 19 % and flexural strength is 37 MPa.

The formation of pure stable, stoichiometric nano crystalline HAP at low

temperature using ethanol-based sol-gel technique was reported by Kuriakose et al.,

[137]. Equimolar solutions of calcium nitrate and diammonium hydrogen phosphate

dissolved in ethanol at 85 °C. The presence of alcohol seems to provide a thermally

stable HAP. The pores in the crystal planes will help the material to attain more

biocompatibility and enables the circulation of physiological fluids. This method

also produced nanocrystalline HAP in the range of 1.3 nm in radius for application

in ideal bone replacement material.

Bigi et al., [138] reported the synthesis of hydroxyapatite gels and

nanocrystals prepared through a sol-gel process. Here, they investigated the effect of

the Ca/P molar ratio on the structural and morphological properties of HAP gels and

nanocrystals. The process was carried out in aqueous and in alcoholic medium

(50% water – 50% ethanol), at 37 °C. Gel samples were obtained by drying the sols

at 37 °C or at 80 °C, whereas powder samples were obtained by filtering the sols.

Heat treatments at temperature as low as 300 °C was enough to obtain the pure form

of HAP from the gels with a Ca/P molar ratio of 1.00 and 1.67. At variance, heat

treatment of the gels with a Ca/P of 2.55 always produces secondary phases. The

degree of crystallinity of HAP increases with the Ca/P molar ratio of the sols, and it

51

is slightly affected by the presence of ethanol in the precipitation medium. Filtering

of the sols provides powders constituted with nanocrystalline HAP that exhibit

degree of crystallinity, crystal morphology and thermal stability closely related to

the sols composition.

The synthesis of HAP nanopowders has been reported by Feng et al., [139]

using sol-gel method. The phosphorus pentoxide and calcium nitrate tetrahydrate

were used a Ca and PO43-

precursors. The crystalline degree and morphology of the

resulting nanopowders are dependent on the sintering temperature and aging time.

The crystalline degree of the HAP nano powders increased with the increase of

firing temperature, and HAP can decompose at 800 °C and above. The crystal size

of the HAP nano powders increases with the increase of aging time.

Bogdanoviciene et al., [140] have developed the calcium HAP samples with

different morphological properties by using aqueous sol-gel route based on

ammonium-hydrogen phosphate as the phosphate precursor and calcium acetate

monohydrate as source of calcium ions. In this process, an aqueous solutions of

(EDTA) or tartaric acid (TA) as complexing agents were added to the reaction

mixture. In both the cases, the polycrystalline single-phase HAP can be produced at

1000 °C. The proper selection of complexing agents in the sol-gel processing allow

to control the grain size and other morphological features of the resulting HAP

powders.

Hosseini et al., [141] have developed HAP powders using calcium nitrate

and triethyl phosphate precursors by an alkoxide based sol-gel technique. The sol of

phosphorus was first hydrolyzed for 24 h with distilled water. The sol temperature,

aging time and heat treatment temperature on apatite formation were studied by

52

increasing the aging time which affected CaO reduction. Also increasing the mixed

sol solution temperature upto 80 °C, this had a positive effect on the disappearance

of impurity phases. The calcium phosphate impurity phases disappeared with the

increase of the calcination temperature < 600 °C. The results indicated that the mean

crystallite size increased and micro-strain decreased significantly with the rise in

firing temperature.

Natarajan et al., [142] have synthesized nanosized HAP particles by sol-gel

method using the water-based solution of calcium and phosphorus precursors. Here

two calcium precursors such as calcium nitrate tetrahydrate and calcium acetate

were chosen as calcium precursors. The influence of aging period, pH, viscosity and

sintering temperature on crystallinity and morphology of the HAP particles were

investigated for the two calcium precursors with triethyl phosphate precursor. The

morphology of nano-HAP towards phosphorus precursor was dependent on the type

of calcium precursor used. The HAP prepared from calcium nitrate and triethyl

phosphate was spherically shaped where as the one from calcium acetate was found

to be fibrous in structure. Both HAP’s were stable up to 1200 °C and their

crystallinity increased with respect to the sintering temperature.

Sanosh et al., [143] have prepared rod like hydroxyapatite nanostructures by

a simple sol–gel precipitation method. The nanoHAP rods produced at 600 °C by

this technique simulated the morphology, size and phase of HAP crystals in human

teeth. From their results, they evaluated the average HAP crystal sizes and showed

proximate values (-30 nm) for both synthesized and human teeth powders.

Mechanisms of nano rod formation was proposed which was attributed to relative

surface energies of facets of HAP crystal and high pH = 11 employed in this study.

53

Their studies showed all the characteristic bands of HAP and prominent presence of

CO23-

bands, which seem to disappear as CO2 in synthesized HAP powders during

calcination at 600 °C.

Padmanabhan et al., [144] have obtained HAP nano-hexagonal rods with

70-90 nm in diameter and 400-500 nm in length by a simple sol-gel route using

calcium nitrate and potassium dihydrogen phosphate as calcium and phosphorus

precursors, respectively. Deionized water was used as a diluting media and ammonia

was used to adjust the pH. The aspect ratio of HAP nanorods was found to be

between 6 and 7. The crystallite size of the HAP nano particles increased with

decreasing the temperature and showed an anisotropic crystal elongation resulting in

nanorods at 700 °C.

Kumar et al., [145] have synthesized plate-like crystals of HAP by

ethanol-based sol-gel method. Calcium nitrate tetra hydrate and diammonium

hydrogen phosphate were used as starting materials and polyethylene glycol was

added as organic modifier at low synthesis temperature of 85 °C. This method

provides the synthesis of pure, porous and stoichiometric HAP at alkaline pH via an

ethanol based sol-gel process. Flexura chain state and numerous ether bonds

presented in polyethylene glycol induce the axis orientation growth of HAP via

interaction between the ether bonds of polyethylene glycol and HAP nano

crystallites resulting in the formation of HAP nano particles. polyethylene glycol

molecules induced the orientation growth which leads to the formation of HAP

platelets with an average size of 50-70 nm.

54

2.1.5 Molten Salt Method

Tas et al., [146] have studied the synthesis of HAP by the molten salt method

using K2SO4 in the temperature range of 1080-1200 °C. They have concluded that

the molten salt synthesis with a K2SO4 flux was found to be a simple and study

technique for manufacturing short (≤ 60µm) HAP whiskers.

The effects of synthesis temperature and reaction time on morphology of the

synthesized HAP nanoparticles were investigated. From the results they have

concluded that molten salt synthesis was found to be a simple technique for

manufacturing (≤ 60 µm) HAP nano particles. Synthesis of fluoride substituted

hydroxyapatite by a molten salt synthesis route using K2SO4 and Na2SO4 as the flux

was reported by Zhang et al., [147].

Viswanath et al., [148] have also studied the mechanical properties of

as-synthesized tricalcium phosphate single crystal which is grown by molten salt

method using K2SO4 as the flux at 1350 °C. Gopi et al., [112] reported the synthesis

of nano HAP by molten salt method. They found that the crystallinity, size and

morphology of HAP nano powders were affected by the calcining time. This method

also forms pure phase of HAP without any impurities and showed rod-like

morphology without detectable decomposition up to 1100 °C.

2.1.6 Hydrothermal Technique

The conventional hydrothermal method is one of the earliest synthesized

methods that were used to produce HAP and by this technology various ceramic

materials including HAP could be synthesized [149]. Though this method offers

advantages to synthesize pure HAP powder difficulties such as long reaction times

55

and agglomeration prevent this method from being the most effective method for the

synthesis of HAP [150-152].

A successive process of hydrothermal treatment using surfactants

(SDS & CTAB) as regulators was reported by Yan et al., [153] for the better

nucleation and crystal growth of HAP. The HAPs obtained at room temperature

were fibrous poly crystals. After hydrothermal treatment, the as formed HAP

nanorods (150 nm x 10 nm) were displayed with uniform morphology. The added

surfactant was then supposed to bind to certain ions, so that the ions could be

incorporated to the existing nuclei at a steady rate and the final shape and size could

be well controlled. The anisotropy of CTAB probably induced the axis orientation

growth of HAP.

Riman et al., [154] reported the synthesis of hydroxyapatite designer

particulates by low temperature hydrothermal and mechanochemical-hydrothermal

method. They explained that the thermodynamic calculations facilitate the process

engineering for hydrothermal and mechanochemical-hydrothermal production of

HAP designer particulates. Due to kinetic factors, the hydrothermal technique is

appropriate for validation of phase diagrams at elevated temperatures, while the

mechanochemical-hydrothermal technique is well suited for validation of phase

equilibria at room temperature. The results emphasize that the hydrothermal

technique is particularly well suited to control HAP size and morphology through

variation of both thermodynamic and non-thermodynamic processing variables.

Conversely, the room-temperature mechanochemical-hydrothermal technique allows

precise control of HAP composition while having a lower degree of control over

particle morphology and aggregation.

56

Wang et al., [155] reported the HAP nano particles with uniform morphologies

and controllable size by low temperature hydrothermal method in the presence of

cationic surfactant. The CTAB is used as template to regulate the nucleation and

crystal growth. The CTAB can bind with phosphate anion of reaction system by the

charge and stereochemistry complementary, so that phosphate anion can be

incorporated to the existing nuclei at a steady rate and the final shape and size of HAP

particles can be well controlled. Compared with the reaction time, the temperature of

reaction is more significant variable in altering the HAP morphology and size.

Li et al., [156] have reported the large-sized hydroxyapatite whiskers with

length of 50–100 nm and width of 0.5 nm synthesized by a facile hydrothermal

method. Dilute reaction solutions containing glutamic acid were used in order to

achieve low degree of super saturation with respective to HAP precipitation. The

super saturation values at different hydrothermal temperatures were

calculated theoretically from ion concentrations with considerations of all association–

dissociation balances between various ions. Experimentally, it was found that

HAP whiskers with smaller sizes were obtained when raising the additive amount of

NaOH or glutamic acid.

Plate-like nanocrystals of HAP synthesized by the hydrothermal method with

the aid (sodium tri polyphosphate, STPP) was reported by Zhang et al., [157]. The

diameters of the nanoparticles become larger with the increasing concentration of

STPP. In the hydrothermal solution, the change of crystal morphology can be

interpreted by the strong preferential adsorption of the HAP crystal, which inhibits

the growth site of (100) plane of HAP crystal and lead to the modification of crystal

structure.

57

Nanocrystalline calcium phosphate crystals synthesized using the simple

hydrothermal method with a Ca-EDTA/PO4 solution were reported in

Xin et al., [158]. The results showed that pure DCPD, DCPA and HAP nano crystals

were obtained from the Ca-EDTA/PO4 solutions at 120 °C, 180 °C and 210 °C,

respectively. Thermal analysis of the precipitated DCPD powders revealed that

DCPD to DCPA and DCPA to HAP transformations occurred at 139 °C and 195 °C.

Revealing these transformation temperatures enable them to design routes of

synthesizing Ca-P crystals with hydrothermal methods.

Hernándeza et al., [159] reported the synthesis of hydroxyapatite

nanoparticles with a hexagonal phase and high crystallinity by using the

hydrothermal method. The hexagonal phase was also corroborated by the

TEM micrographs. Crystal sizes varying from 20 to 48 nm were estimated.

From EDS the Ca/P and Ca/O ratios were estimated. The results show that the better

conditions to synthesize HAP nanoparticles are: t1 = 12, 24 h, T1 = 120 °C, t2 = 24 h,

and T2 = 70 °C.

Shojai et al., [160] reported the synthesis of HAP nanorods using a simple

hydrothermal procedure. The HPO42-

containing solution was added drop-wise into

the Ca2+

solution while the molar ratio of Ca/P was adjusted at 1.67. The results

confirmed the high purity, high crystallinity and high aspect ratio of synthesized

HAP nanorods. They showed high dispersion stability in the dilute dental

experimental adhesive. So it can be regarded as an alternative to other fillers such as

silicates for using in dental adhesives.

Zhua et al., [161] have synthesized rod-like hydroxyapatite nanoparticles

with various aspect ratios by means of low-temperature hydrothermal method in the

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presence of the N-[(2-hydroxy-3-trimethylammonium) propyl] chitosan chloride

(HTCC) template. The results revealed that the size and morphology of HAP

crystals can be tuned by varying pH, hydrothermal temperature and the ratio of

quaternary ammonium in HTCC to PO43−

. HTCC can be incorporated to the

phosphate anions by charge and stereo-chemical complementarily.

Zhao et al., [162] have reported the mesoporous hydroxyapatite

nanoparticles by the hydrothermal method using pluronic block co-polymer F127

and CTAB as templates. The obtained mesoporous HAP was employed as a drug

delivery carrier to investigate the drug storage/release properties using carvedilol

(CAR) as a model drug. The results demonstrated that CAR was successfully

incorporated into the mesoporous HAP host. In vitro drug, consequently,

mesoporous HAP is a good candidate as a drug carrier for the oral delivery of poorly

water-soluble release studies and showed that mesoporous HAP had a high drug

load efficiency and provided immediate release of CAR compared with micronized

raw drug in simulated gastric fluid (pH 1.2) and intestinal fluid (pH 6.8) drugs.

Rena et al., [163] presented a facile and are effective isotropic growth of 1-D

hydroxyapatite (1-DHAP) nanorods via a hydrothermal route in weak acid

environment in the presence of sodium bicarbonate, without using any

template/surfactant reagents and organic solvents. The single crystalline

hydroxyapatite nanorods are with several hundred nanometers in length and tens of

nanometers in width. One dimensional (1-D) growth and aspect ratio could be

controlled by hydrothermal reaction time and temperature. The obtained

nanostructures are with high homogeneity and high purity without crystalline

defects. Single crystals of HAP nanorods preferentially grow along the c-axis. These

59

nanorods are expected to have potential applications in dentistry, medicine and

biocompostites.

2.1.7 Freezing Method

Deville et al., [164] have reported that the freeze casting can be applied to

attain porous scaffolds exhibiting high compressive strength. They have investigated

that parameters like initial slurry concentration, freezing rate and sintering

conditions influenced the porosity and compressive strength of the synthesized

HAP. Thus by using this method they modified porosity of HAP scaffolds which can

be designed for load-bearing applications.

Lee et al., [165] reported the camphene-based freeze casting technique to

produce highly porous HAP bioceramics with 3-dimensionally interconnected pore

channels. The porosity and mechanical properties of the sample were controlled by

adjusting the initial HAP contents used in the preparation of HAP/camphene slurry.

As the initial HAP content was increased from 10 to 20 vol. %, the porosity linearly

decreased from 75 to 56 %, still preserving highly interconnected pore structures.

A reductionin the porosity resulted in the remarkable improvement of the

compressive strength from 0.94 to 17 mpa.

Landi et al., [166] have suggested a cryogenic process, including

freeze-casting and drying method to obtain hydroxyapatite scaffolds having 10 mm

diameter, 20 mm height with completely lamellar morphology. They studied that the

HAP scaffolds had referentially aligned channel-like pores. By changing the process

parameters of HAP slurry, lamellar ice crystals with different thickness grew

throughout the samples. They have proved the interconnection of pores and the

ability of the scaffolds were rapidly penetrated by synthetic body fluid.

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Zhang et al., [167] reported the synthesis of porous hydroxyapatite ceramics

by freeze casting method and gelatin was used to adjust the pore morphology and

microstructure of the porous HAP ceramics. In the addition of gelatin the large and

non-interconnected lamellar pores were changed into small and interconnected

cellular pores. Gelatin addition has greater effect not only on the pore size and pore

morphology of the HAP ceramics but also the viscosity of slurry, the porosity and

shrinkage of HAP ceramics.

Macchetta et al., [168] have studied a room temperature camphene-based

freeze casting method to fabricate hydroxyapatite/tricalcium phosphate (HAP/TCP)

ceramic scaffolds. By varying the solid loading of the mixture and the freezing

temperature, a range of structures with different pore sizes and strength

characteristics were achieved by them. The porosity decreased from 72.5 to 31.4 vol.

% when the solid loading was increased from 10 to 30 vol. %. This resulted in an

increase in the compressive strength from 2.3 to 36.4 MPa.

Gopi et al., [169] have reported a novel method to synthesize nanoporous

hydroxyapatite powders by freezing organic–inorganic soft solutions. The formation

of porous and crystalline HAP nanopowder was achieved via calcining the samples

at 600 °C followed by sintering at temperatures ranging from 900 °C to 1100 °C.

The results showed the formation of a carbon free nanoporous HAP powders due to

the decomposition of organic template enclosing the precipitated HAP. They also

observed that the rapid grain growth with retainment of pores while the crystallinity

of the HAP nanopowder increased with the increase in sintering temperature which

is substantiated from their results. Such organized porous materials can act as a

biomaterial for bone tissue engineering.

61

Farhangdoust et al., [170] have fabricated macroporous hydroxyapatite

scaffolds, which could overcome the current bone tissue engineering limitations. In

this study they investigated, controlled unidirectional freeze-casting at different

cooling rates (2 to 14 °C/min), sintering temperature (1350 °C) and slurry

concentration (7-37.5 vol. %). The mechanical strength of the scaffolds increased as

a function of initial concentration, cooling rate and sintering temperature with

regards to mechanical strength and pore size, the samples with the initial

concentration and the cooling rate of 15 vol. % and 5 °C/min, respectively showed

better results.

2.1.8 Mechano-chemical Synthesis

Tabrizi et al., [171] also proposed the synthesis of nanosize single-crystal

HAP by a mechano-chemical process. Here they explained the feasibility of using

polymeric milling media to prepare HAP nano particle and it exhibits an average

size of about 20 to 23 nm. This method also provides a facile pathway to obtain

single-crystal HAP.

Gergely et al., [172] have reported the synthesis of HAP from recycled egg

shell. The observed phases of the synthesized materials were dependent on the

mechano-chemical activation method (ball milling and attrition milling). They

showed that the ball milling process resulted in micrometer sized coagulated coarse

grains with smooth surface, whereas attrition milled samples were characterized by

the nanometer size grains. This characteristic morphology being preserved even after

firing at high temperature (900 °C) contrary to ball milling attrition resulted in

nanosized and homogeneous HAP even after milling.

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Yeong et al., [173] have synthesized nanocrystalline HAP phase by

high-energy mechanical activation in a dry powder mixture of calcium oxide and

anhydrous calcium hydrogen phosphate. The initial stage of mechanical activation

resulted in a significant refinement in crystallite and particle sizes, together with a

degree of amorphization in the starting powder mixture. This is followed by steady

formation and subsequent growth of HAP crystallites with increasing degree of

mechanical activation. A single phase HAP of high crystallinity was attained by

>20 h of mechanical activation. The resulting HAP powders exhibit an average

particle size of ~ 25 nm. It was sintered to a density of 98.20 % theoretical density at

1200 °C for 22 h.

2.1.9 Template Method

Zhang et al., [174] in their work reported the large-scale hydroxyapatite

single crystal nano-wires synthesized via template technology. The results

confirmed that crystalline order of the HAP precursors was retained in the

electrodeposited nano-wires. The HAP single crystal nano-wires grew in c-axis

co-orientation along with the direction of the template, which beard structural

similarity to the HAP found in the natural bone. It is desired to be studied widely

and thoroughly to interact with and replace natural biological materials.

McQuire et al., [175] reported a new template-directed method for the

fabrication of HAP sponges by using amino-acid-coated HAP nano particles

dispersed within a viscous polysaccharide (dextran sulfate) matrix, and described the

use of these materials for the viability and proliferation of human bone marrow

stromal cells. In presence of excess amounts of aspartic acid, alanine or arginine the

nano particles subsequently organized into macroporous frame works with typical

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pore sizes of 100-200 µm during thermal degradation of the dextran matrix.

The sponge macrostructure was influenced by changes in the heating rate and

sintering time, as well as the use of different amino acids or variations in dextran

functional groups.

Madhavi et al., [176] have reported the macroporous hydroxyapatite

synthesized using polystyrene sphere templates that were impregnated with a

calcium phosphate precursor solution which was allowed to solidify followed by

sintering from 500 to1000 °C in flowing oxygen to remove the polymer and

crystallize the phosphates.

Mollazadeh et al., [177] in their studies reported the synthesis of HAP

crystals via an in situ biomimetic process using poly (vinylalcohol) (PVA) as a

templating agent. The results indicate that the molecular weight of the templating

polymer is an important factor in determining the particle size in the fixed HAP

product. The in situ synthesized particles were less agglomerated which is believed

to be the result of the nucleation of HAP crystallites on the regularly arranged side

groups of polymer chains. In addition to the size, the crystallite shape was also

influenced by the presence of polymer.

He et al., [178] in their work reported the porous hydroxyapatite spheres with

controlled purity of phase and governed aperture were grown by the

template-directed method. The results indicated that the lower concentration of Ca-P

was prone to pure HAP phase and the aperture decreased gradually with the increase

of the concentration of template. Correspondingly, the crystallization

thermodynamics and template-directed growth kinetics were discussed in details.

64

4

Banerjee et al., [179] have synthesized hydroxyapatite nanopowders with

different aspect ratios using reverse micelle template system and it was observed

that increase in aqueous to organic ratio and pH decreased the aspect ratio of the

nanopowders. HAP nanopowders with the highest aspect ratio (rod-shaped) of

7.2±3.2 and the lowest aspect ratio (spherical) of 1.3±0.3 were synthesized for

processing dense compacts. Effect of powder morphology on densification at

1250 °C was studied with different amount of rod-shaped and spherical

nanopowders. It was observed that an increase in high aspect ratio powder content in

the compacts decreased sintered density under pressure less sintering condition.

Li et al., [180] have reported the nanoporous hydroxyapatite with pore size

ranging from about 1–5 nm synthesized by utilizing CTAB as template. The effects

of reaction temperature and CTAB on the phase and morphology of HAP were

investigated by them and they found that the pore structure of HAP was thermally

stable temperature greater than 700 °C and the pore size was independent of

processing temperature and CTAB:PO 3-

at ratio.

He et al., [181] have also reported the synthesis of nanoflake hydroxyapatite

by a biomimetic method according to the biomineralization theory using

Ca(NO3)2.4H2O and (NH4)3PO4.3H2O as reagents and chondroitin sulfate (ChS) as a

template. The results indicated that ChS concentration significantly affected HAP

growth behavior and morphology. Staple fiber-like HAP crystals could be obtained

in the presence of low concentration of ChS and flake HAP crystals synthesized in a

high concentration of ChS (0.5 wt.%). HAP crystallinity increases continuously with

decreasing ChS content.

65

Guo et al., [182] Lamellar hydroxyapatite with worm-like mesopores was

successfully synthesized from the mixed ethanol solutions of Ca(NO3)2·4H2O and

P2O5 in the presence of polyoxyethylene (20) sorbitan monostearate (Tween-60).

From the results it was found that the content of Tween-60 influenced the

crystallinity and the size of lamellar hydroxyapatite, with the volume fraction of

Tween-60 increasing the size of HAP lamellasomes increased. The mesopores,

around 4 nm in diameter, have a high heat resistance. The mesopores forming

templates were from the reactive products of P2O5 and ethanol.

Salarian et al., [183] reported the synthesis of dandelion-like HAP by a

template-directed synthetic method, using CTAB as a template and co-surfactant

polyethylene glycol (PEG600) as a co-template under hydrothermal conditions. The

results showed in the presence of CTAB and a certain concentration of PEG 600

(30%) HAP crystals have a uniform dandelion-like morphology with a diameter of

about 80–150 nm and aspect ratio of about 20 for each tooth. Dandelion-like HAP

crystals have a high surface area of 88 m2g

-1 showing potential applications.

Xiao et al., [184] have reported the spherical micrometer hydroxyapatite

crystals by using Ca(NO3)2·4H2O and (NH4)3PO4·3H2O as reagents and

β-cyclodextrin (β-CD) as template. The concentration of β-CD has an effect on the

morphology of HAP crystals. Spherical HAP with the diameter of 1.0–3.0 µm can

be obtained in the presence of 1.5 % β-CD. The as-prepared samples are the

composites of HAP/β-CD. This is a result of the interaction between HAP and

β-CD. The –OH of β-CD is the major interaction site between Ca2+

and β-CD.

Ye et al., [185] in their study reported the synthesis of hydroxyapatite (HAP)

hollow nanoparticles (HNPs). The HNPs produced through a novel polymeric

66

micelle-templating method. The micelles were structured with completely insoluble

Pluronic P123 molecules at cloud point as the core and Tween-60 molecules as the

shell by the hydrophobic interaction of the alkyl chains with the insoluble P123 core.

The morphology of the HAP HNPs could be transformed from nanospheres to

nanotubes by adding citric acid as a co-surfactant.

Tari et al., [186] have reported the synthesis of hydroxyapatite nanopowders

using the micelle as a template system. The mixtures of CTAB and SDS with

different ratios were used as the template in order to investigate the effect of mixed

surfactant on the morphology of synthesized particles. The results revealed that the

overall morphology of the obtained powders at anionic-rich region

(SDS: CTAB, 99:1) is rod like, but in the presence of cationic rich region

(SDS:CTAB, 1:99) the resulted particles was sheet like which because of the

interaction of surfactants with opposite charges. The resulted HAP nano particles in

the presence of SDS were rod like but their morphology was less oriented than

anionic-rich region. Our experience showed that the type of surfactant has great

effect on the morphology of synthesized particles.

Son et al., [187] presented a novel and simple reaction for the preparation of

HAP nanoparticles by the use of β-CDs template. The β-CD content slightly affected

the particle size of products. In addition, the synthesis procedure (mixing sequence

of ion precursors) also influenced the particles size and morphological structure of

nanoparticles. The HAP nanoparticles synthesized in the presence of β-CD exhibited

as an aggregation. The HAP nanoparticles had very narrow size distribution, and the

composition and crystalline phase were similar to those of the stoichiometric HAP.

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However, the crystallinity of synthesized HAP nanoparticles was low compared with

that of commercial HAP.

2.2 LIMITATIONS OF CHEMICAL METHODS

All the above work has been done in the presence of various toxic chemicals.

Chemical synthesis methods lead to presence of some toxic chemicals which is

absorbed on the surface that may have adverse effects in the medical applications.

2.3 ADVANTAGES OF GREEN TEMPLATE METHOD

The synthesis protocols for HAP particles involving environmentally

mediated materials like plant extract offers numerous benefits of eco-friendliness

and compatibility for pharmaceutical as well as biomedical applications. Further,

green template synthesis proves to be the best method rather than the chemical and

physical method, as it is economical, environment friendly and easily scaled up for

large scale synthesis and in this method there is no need to use high pressure,

energy, temperature and toxic chemicals.

We aim to synthesize the HAP particles using a green chelating agent as

template under atmospheric pressure, and to investigate their morphology,

crystallinity and particle size.

Nayar et al., [188] in their work used biomolecules in waste and medicinally

important materials for the synthesis of hydroxyapatite nanoparticles. Orange and

potato peel, eggshell, papaya leaf and calendula flower extracts have varied

biomolecules, which exert a significant, control on the in situ synthesis of nanosized

HAP particles. This study proved that only small quantities of biomolecules exert

significant effect on the microstructure that may lead to novel properties.

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Klinkaewnarong et al., [189] have synthesized nano crystalline HAP

powders by a simple method using aloe Vera plant extract solution. The particle

sizes of the powder they obtained were 40-171 nm. Their results also showed that

the synthesized HAP powders were in hexagonal structure and was fully formed

after calcination at 500 °C. The crystal size of HAP samples increased with

increasing the calcination temperature and the aloe Vera plant extracted solution can

act as a new template to produce nano crystalline HAP powders.

A novel and effective method for the preparation of water-dispersible

nano-hydroxyapatite particles was reported by Zhou et al., [190]. nHAP was

prepared in the presence of grape seed polyphenol (GSP) solution with different

concentrations. Chemical precipitation method was adopted to produce pure nHAP

and modified nHAP (nHAP-GSP) at 60 ◦C for 2 h. The results indicated that the

spherical nHAP particles with a diameter of 20–50 nm could be synthesized at 60

°C. The zeta potential values of pure nHAP and nHAP-GSP are −0.36 mV and

−26.1 mV, respectively. According to the sedimentary time, the colloidal stability of

nHAP-GSP in water could be improved dramatically with the increase of GSP

content and the particles tended to exist as dispersive nanoparticles without

aggregation. All the results indicated that GSP exhibited strong binding to nHAP

and enhanced the colloidal stability of nHAP particles.

The role of the addition of a chelating agent from various sources on the

purity, crystallinity and morphology of the HAP particles obtained by the green

template synthesis method was investigated by Gopi et al., [191]. The sucrose

extracted from the various natural sources was found to be pure and coincide with

the commercially available one as evident from the results of FT-IR and 13

C NMR.

69

The LC–MS results revealed the variation in the concentration of the hydrolyzed

products of sucrose. The FT-IR and 13

C NMR results emphasized that the HAP

particles synthesized by this method were found to be pure and free from any

organic moieties. The uniform morphology and less agglomerated HAP particles

were obtained from the natural sources of sucrose than the commercially available

one. The crystallinity, particle size and the morphology of HAP are strongly

dependent on the natural sources of chelating agent used, especially the hydrolyzed

products of sucrose which was apparent from the LC-MS, XRD and SEM results.

The HAP particles of fairly well-defined dimensions with reduced size can be

achieved by using only the stem sugarcane extract powder as the natural source of

chelating agent. This green template approach towards the synthesis of HAP

particles has many advantages such as ease with which the process can be easily

scaled up, biodegradability, biocompatibility, economic viability and its uses in

biomedical applications.

Hydroxyapatite nanorods with excellent antibacterial properties have been

successfully synthesized using the green template method and was reported by

Gopi et al., [192]. Tartaric acid is used as green template for the overall synthesis of

HAP which has been obtained from the extracts of fresh fruits such as banana,

grapes and tamarind, respectively. The synthesis of HAP using commercially

available tartaric acid was also carried out for comparison purpose. The phase pure

HAP nanorods without any impurities have been found from the FT-IR and XRD

results. When changing the sources of tartaric acid from commercial to natural, the

crystallinity and crystallite size were decreased. Among the three different natural

fruit sources, the HAP nanorods obtained from the extracts of tamarind possesses

70

reduced crystallinity and crystallite size when compared to the HAP nanorods from

the extracts of grapes and banana. SEM images showed that even though the

nanorods were formed in all the cases, the uniform size distribution was obtained

only by using the tamarind extract as the green template when compared to other

fruit sources. The antibacterial results reveal that the as-synthesized HAP nanorods

exhibited a strong antibacterial activity against both the Gram-negative bacteria of

E.coli and Klebsiella species. The HAP nanorods synthesized by this green template

method can be used as an impending material for various biomedical applications.

Gopi et al., [193] also used an inexpensive, non-toxic, ecofriendly,

abundantly available green waste for the consistent and raid synthesis of HAP

nanoparticles. The HAP nanoparticles were green synthesized using pectin extracted

from the peels of banana as template. The pectin extracted from the cell wall

materials of banana peel was found to be pure. They obtained mixed phases of Ca-P

when the concentration of pectin was lower and phase pure HAP when the

concentration of pectin increases. Also the crystallinity and crystallite size of HAP

decreased as the concentration of pectin increases. They found the concentration of

pectin plays a major role in controlling as well as reducing the size of nanoparticle

aand found the optimum concentration for obtaining the uniformly distributed

discrete spherical HAP nanoparticles was found to be 0.15 wt.%. The as-synthesized

HAP nanoparticles derived from banana peel pectin showed strong antibacterial

activity against both the gram positive and negative bacteria like S. aureus and

E. coli, respectively. The HAP nanoparticles synthesized by this green pectin

mediated method can be used as good biomaterials for various biomedical

applications.

71

. Hence, the present work deals with the green chelating agent as template for

the synthesis of HAP powders. Here in my work I have used using malic acid, oxalic

acid, sucrose and tartaric acid as green chelating agent obtained from various plant

natural sources such as apple fruit, tomato vegetable, pineapple ripe fruit, carrot

root, sugarcane stem, banana fruit, grapes and tamarind. For comparison, HAP

powders have also been synthesized using commercially available malic acid, oxalic

acid, sucrose and tartaric acid. As far as the literature is concerned, there are no such

reports claiming the synthesis of HAP by this green chelating agent as template.

Hence, we propose this method as a simple and an effective route to produce bulk

bioceramic material.