Physical and mechanical properties of a poly-3-hydroxybutyrate-coated nanocrystalline hydroxyapatite scaffold for bone tissueengineering

Mohammad Reza Foroughi • Saeed Karbasi •

Reza Ebrahimi-Kahrizsangi

� Springer Science+Business Media, LLC 2011

Abstract A major challenge for tissue engineers is the

design of scaffolds with appropriate physical and

mechanical properties. The present research discusses the

formation of ceramic scaffolding in tissue engineering.

Hydroxyapatite (HAp) powder was made from bovine bone

by thermal treatment at 900 �C; 40, 50 and 60%wt porous

HAp was then produced using the polyurethane sponge

replication method. Scaffolds were coated with poly-3-

hydroxybutyrate (P3HB) for 30 s and 1 min in order to

increase the scaffold’s mechanical properties. XRD, SEM

and FT-IR were used to study phase structure, morphology

and agent groups, respectively. In XRD and FT-IR data,

established hydrogen bands between polymer and ceramic

matrix confirm that the scaffold is formed as a composite.

The scaffold obtained with 50%wt HAp and a 30 s coating

was 90% porous, with an average diameter of 100–400 lm,

and demonstrated a compressive strength and modulus of

1.46 and 21.27 MPa, respectively. Based on these results,

this scaffold is optimised for the aforementioned properties

and can be utilised in bone tissue engineering.

Keywords Hydroxyapatite (HAp) �Poly-3-hydroxybutyrate (P3HB) � Scaffold �Composite � Nanocrystal � Tissue engineering

1 Introduction

Tissue engineers study the design and production of new

tissues to regenerate damaged organs and replace lost

bones [1]. Bone repair and regeneration is a common and

complicated clinical problem in orthopedic surgery. In

huge fractures, the normal healing process fails to work

properly, and bone grafting surgery is therefore required

[2]. Recently, a great deal of interest has been directed

towards creating bioactive ceramic/polymer composites to

be used as bone grafting materials. In the past, scientists

cultured the necessary human cells ex vivo, but complex,

3-dimensional cell-network technology to replace damaged

tissue is ever expanding. A scaffold with an appropriate

physical framework is required in order to form tissue

based on such engineered methods. This scaffold must

allow cells to attach, migrate, duplicate, correspond inter-

cellularly and grow and replace tissues. Both polymer

matrix and ceramic matrix scaffolds have high porosity,

bioactivity, biodegradability and other physical properties

required for bone tissue engineering. Ceramic matrix

scaffolds have higher than polymer matrix scaffolds bio-

activity and are therefore recommended [3].

Natural bone is a 70%/30% inorganic/organic composite

material. Hydroxyapatite (HAp) is the main inorganic

component of bone and is a popular implant material in

bone surgery [4]. Due to its similar chemical composition,

mechanical resistance and stiffness to natural and tooth

bone cores and minerals, HAp has been widely used in

orthopedic and dental implants [5]. HAp minerals can

either be synthesised [6–12] or extracted from natural

sources [13–22].

Based on previous studies, high porosity, excellent

compressive strength and the possibility of cell migration

are important scaffold criteria [23–30]. Polymer replication

M. R. Foroughi � R. Ebrahimi-Kahrizsangi

Department of Materials Engineering, Najafabad Branch,

Islamic Azad University, Isfahan, Iran

S. Karbasi (&)

Medical Physics and Biomedical Engineering Group, School of

Medicine, Isfahan University of Medical Sciences, Isfahan, Iran

e-mail: karbasi@med.mui.ac.ir

123

J Porous Mater

DOI 10.1007/s10934-011-9518-1

methods allow for the formation of uniform scaffolds with

adjustable porosity [31–33]. The potential for improving

the mechanical properties of bioceramics/polymer com-

posite scaffolds by this approach has been demonstrated in

several systems, which have achieved mechanical proper-

ties, in particular compression strength, in the range of

values for cancellous bone [34]. Polymer coatings can be

used to reinforce these types of scaffolds. Miao et al.

coated poly lactic-co-glycolic acid (PLGA) onto ceramic

scaffolds of HAp tricalcium phosphate (TCP), giving the

coated ceramic scaffold a compressive strength and com-

pressive module of 0.66 and 16.85 MPa, respectively [35].

Tan et al. coated PLGA-Bioglass onto ceramic scaffolds of

HAp and reached compressive strength and compressive

module values of 1.36 and 24.58 MPa, respectively [36].

Jun et al. coated porous HAp scaffold with Apatite-Wol-

lastonite glass and 45S5 ceramic glass to increase com-

pressive strength [37]. TiO2 foam-like scaffolds with pore

size *300 lm and [95% porosity were fabricated by the

foam replication method by Novak et al. [38]. In order to

improve the structural integrity of the as-sintered foams,

which exhibited extremely low compression strength

(\0.045 MPa), PDLLA or PDLLA/Bioglass coatings were

developed. The PDLLA coating of a few microns in

thickness was shown to improve the mechanical properties

of the scaffold: the compressive strength was increased by

a factor of *7 (0.3 MPa). Moreover the composite coating

involving Bioglass particles was shown to impart the rutile

TiO2 scaffold with the necessary bioactivity for the inten-

ded applications in bone tissue engineering.

Many other components of biodegradable polymers and

bioactive ceramics have been utilised as biocompatible

scaffolds in tissue engineering. Polyhydroxyalkanoate

(PHA) is a biodegradable polymer used in tissue regener-

ation, drug delivery and patches, either alone or in

composite form [39]. One member of the PHA family is

poly-3-hydroxybutyrate (P3HB), which has a longer deg-

radation time than poly a-hydroxy acids (For example,

PLA and PLGA) and can be produced by various organ-

isms [40]. P3HB is biocompatibile with many different

types of cells. P3HB does not degrade into acidic

by-products, while degraded PLGA acidifies the immediate

environment around the cell [41]. P3HB also has piezo-

electric properties, which can play a critical role in stim-

ulating bone growth and regeneration [42]. Different

composite varieties have been made using P3HB matrix

and bioactive mineral particles like HAp, Wollastonite and

bioactive glasses, in order to improve the strength and

bioactivity level of composite [43]. However, due to a

lack of surface particles, bioactivity is only slightly

increased. Therefore, we recommend the use of P3HB to

reinforce ceramic matrix scaffolds to increase bioactivity

and strength, simultaneously. Bretcanu et al. [44] used

bacteria-derived P3HB to infiltrate 45S5 Bioglass scaffolds

intended for use in cancellous bone substitution after

trauma incidents. Pore morphology and macrostructure

before and after coating with P3HB, as well as coating

homogeneity, were investigated. Polymer coating consid-

erably increased the compressive strength of the scaffolds

(*1.5 MPa at 85% porosity).

To our knowledge, this is the first report on the forma-

tion of scaffolds from natural HAp coated with P3HB. HAp

nanocrystal powder was prepared from bovine bone by

thermal deposition, and porous scaffolds were constructed

using a polymer replication method. In order to enhance

mechanical properties, scaffolds were dip-coated P3HB.

Due to its high stability, mechanical amelioration and true

porosity, P3HB will be an important tool in bone tissue

engineering.

2 Materials and methods

2.1 Procurement of ceramic HAp powder

Bovine bones were boiled for 2 h to remove flesh and fat.

The bones were heated at 60 �C for 24 h to remove

moisture. To prevent blackening with soot during heating,

the bones are cut into small pieces (10 9 10 9 10 mm)

and heated at 400 �C for 3 h in air to allow for analysis of

their organic compositions. The resulting black bone ash

was heated for 2 h at 600, 700, 900 or 1,100 �C. The

resulting white nanocrystal HAp powder was used directly

to construct the ceramic scaffold.

2.2 Fabrication of ceramic slurry

The preparation of slurry stabilised with the proper addi-

tives is critical to scaffold formation; a variety of additives

with a range of biological properties have been studied

[45]. In this research, natural HAp powder with a grain size

of 70–80 nm has been utilised as the scaffold matrix. HAp

powder (40, 50 and 60%wt) was slowly dissolved in dou-

bly-distilled water to prevent agglomeration. The slurry

was stirred at 300 rpm for 30 min to give a homogenous

solution. To maximise mechanical strength, large amounts

of solid material should precipitate from the homogenised

solution onto the polyurethane sponge. We used ammo-

nium poly-methacrylate (DARVAN�C–N, R.T. Vanderbilt

Company, Norwalk, USA) to increase the weight per-

centage of solid substance above 1%wt. Carboxymethyl

cellulose (CMC, Hangzhou Hongbo Chemical Co. Ltd,

China) powder (1%wt) was gradually added to the solution

slowly in order to increase slurry flow. The solution was

stirred at 60 �C until it was fully homogeneous.

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2.3 Preparation of porous HAp scaffold

A commercial polyurethane sponge (MEAY Co., Ltd.

China, average pore size 300–700 lm) was used in this

study. The sponge was immersed in ceramic slurry and

squeezed to remove air. Over time, the slurry was absorbed

into the sponge, and the shape was restored by elasticity.

When the sponge was completely filled it was removed

from the slurry and squeezed to remove excess slurry. The

resulting composite porous body was dried for 24 h in air.

Subsequently, the scaffolds were placed in a heat

treatment furnace. The HAp scaffold was formed in four

stages: (1) treatment at 600 �C, with a heating rate of 3 �C/

min for 1 h, to completely burn the sponge, (2) an increase

in temperature from 600 to 1,200 �C at a rate of 5 �C/min,

(3) treatment at 1,200 �C for 4 h to sinter the ceramic

scaffold and (4) cooling to room temperature at a cooling

rate of 5 �C/min. Cooled samples were removed from the

furnace, measured and then placed in desiccators. Figure 1

shows the sponge and scaffold that was formed at

1,200 �C.

2.4 HAp scaffold coated with P3HB

In this study, 0.6 g P3HB powder (Sigma-Aldrich, St

Louis, MO, USA) was dissolved and heated in 10 mL

chloroform (Sigma-Aldrich, USA) for 6 h in an oil bath

with reflux condenser at 60 �C to give the polymer solution

for coating ceramic scaffold. Next, HAp scaffolds were

immersed in the polymer solution for 30 s and 1 min. To

achieve a uniformly coated surface and remove excess

polymer solution, samples were wrapped in aluminium foil

and centrifuged at 500 rpm for 30 s. Finally, the samples

were placed in a vacuum oven at room temperature for

24 h.

2.5 Physical characterisation

2.5.1 X-ray diffraction analysis

X-ray diffraction methods (XRD, Philips X‘Pert) were

applied to gain information about the structural changes

and phases of HAp nanocrystals and scaffolds. A CuKa ray

was used for analysis. The scan rate was set to 1 �/min, the

imposed voltage and current were 40 kV and 30 mA,

respectively, and the diffraction angle (2h) was varied from

10 to 90� at a rate of 0.4 �/min.

2.5.2 FT-IR spectroscopy

IR was used to characterise HAp nanocrystals and scaffolds

after sintering. A 2-mg dried sample was carefully mixed

with 300 mg dry KBr and pressed into a pellet using a

macro KBr die kit. The solid pellet was placed in a magnetic

holder. The system was purged with dry air for 1 h to

remove water vapour from the sample compartment. Fou-

rier transform infrared spectroscopy (FT-IR: 6,300, JASCO,

Japan) has been used for studying functional groups and

specifically, the degree of 2-hydroxylation of HAp. Spectral

analyses were performed using standard Nicolet and Mic-

rocal Origin software. FT-IR spectra were taken of both as-

received and sintered Hap nanocrystals and scaffolds.

2.5.3 Scanning electron microscopy (SEM)

Scanning electron microscopy (SEM, Philips XL-30,

Netherlands) was used to study the HAp nanocrystals and

scaffold. Samples were coated with gold under an argon

atmosphere.

2.5.4 Transmission electron microscopy (TEM)

Transmission electron microscopy (TEM, Philips CM200

FEG, Netherlands) was used to elucidate the nanoscale

structure of HAp powders.

2.5.5 Thermo gravity analysis (TGA)

Thermal gravity analysis (TGA) measures thermal stability

and compounding materials. In this research, a thermal

gravity analysis device (TG/DTA, TGA 401, Sanatara.co)

was used to record the percent weight decrease of the poly-

urethane sponge versus temperature. The sample was heated

at a rate of 1 �C/min, up to 600 �C, under a nitrogen flow.

2.5.6 Porosity and density measurements

Liquid displacement was used to calculate the porosity and

density of the scaffolds. Scaffold density gives informationFig. 1 Ceramic porous scaffold after heat treatment in 1,200 �C

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about the size, distribution and permeability of pores and

the presence of structural defects in sintered ceramic

frameworks [46]. Due to the hydrophobic properties of

polymer, 96% Ethanol, which can pass easily through

pores, was used instead of water. The mass of the ceramic

sample (W) was measured, and a volume (V1) of ethanol

was poured into a graduated cylinder and measured. The

sample was immersed in ethanol for 5 min until it became

saturated (V2). The discrepancy between volumes (V1-V2)

represents the volume of the scaffold. The ethanol-soaked

scaffold was removed from the graduated cylinder, and the

remaining volume was recorded as V3. V1-V3 represents

the volume of ethanol absorbed by the scaffold [45].

The following equation calculates density of the scaffold

(q), (Eq. 1):

q ¼ W

V2 � V3

ð1Þ

The following equation calculates the amount of open

porosity of the scaffold (e), (Eq. 2):

e ¼ V1 � V3

V2 � V3

ð2Þ

2.6 Mechanical characterisation

Machining and gripping the specimen is a major problem

in the mechanical characterisation of ceramic porous

scaffolds: conventional methods of mechanical character-

isation, such as tensile, biaxial and impact testing, are

usually inapplicable to porous materials [46]. Compression

impact tests for porous bone and HAp samples are instead

used [45–48]. Compressive strength and compressive

module tests for samples with and without P3HB coating

were performed using a compression impact tester (SAN-

TAM-Eng. Design co. LTD.) with a 10 KN load cell based

upon guidelines set in ASTM-D5024-95a. The dimensions

of each sample were 20 9 10 9 10 mm3 for the com-

pression impact test. As ceramic scaffolds are fragile, the

crosshead speed was set at 0.5 mm/min to prevent damage

to the ceramic structure. The load carried by the sample

was considered to be 30% of scaffold’s original length. The

elastic modulus was calculated as the slope of the initial

linear portion of the stress–strain curve. The yield strength

was determined from the cross point of the two tangents on

the stress–strain curve around the yield point.

3 Results and discussion

3.1 HAp nanocrystal

Figure 2 shows XRD spectra of bone ash treated at 600,

700, 900 and 1,100 �C. The spectra indicate a gradient of

increased resolution and peak sharpness with increased

temperature. In spectrum Fig. 2a, peak width increased, but

resolution decreased, indicating a minimal amount of

crystalline HAp. The rise in temperature is responsible for

the narrower peak width; sharp peaks appeared at high-

resolution. This phenomenon is due to the increase in HAp

crystal number and size as temperature rises. Shipman et al.

[49] observed similar changes in HAp particle size with

temperature. In Fig. 2, bone ash has started to crystallise at

600 �C without being transformed into other calcium

phosphate structures. Peaks associated with the conversion

of HAp into a-tricalcium phosphate (a-TCP), b-tricalcium

phosphate (b-TCP) and tetra-tricalcium calcium phosphate

(TTCP) are not seen in any spectra below 900 �C. Sharp,

narrow, clear peaks in Fig. 2d indicate that the material

produced at this temperature is purified and crystallised.

Figure 2d, shows that the obtained product is a single-

phase HAp with no additional stable phase. Diffraction

peaks for the product and the JCPDS: 9-432 standard were

compared favourably. The main distinguishing detail of

Fig. 2d is the difference in diffraction pattern spread,

which indicates the range of crystal grain sizes.

HAp grain size was estimated from the Scherrer equa-

tion to be 70–80 nm. It is expected that the product has a

low average grain size. The peak spread amount is a

parameter that can be in the structural property determi-

nation of a product. The product’s degree of crystallisation

(Xc) has been calculated by Eq. 3 [50]:

Xc ¼K

B

� �3

ð3Þ

In this equation, K represents a constant (equal to 0.24)

and B represents peak spread (FWHM (�)). We estimate

that the degree of crystallisation of the product is 92.86%.

However, other studies show that the degree of

crystallisation is subject to factors such as crystal grain

Fig. 2 Pattern of XRD analysis related to natural HAp of thermal

analysis at temperatures of: (a) bone’s ash, (b) 600 �C, (c) 700 �C,

(d) 900 �C and (e) 1,100 �C

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size; the higher degree of crystallisation, the higher the

average size of crystal grains. In XRD spectra, the powder

obtained at 1,100 �C (Fig. 2e) shows a peak with poor

resolution at a 2h angle 29.475, related to the formation of

tricalcium phosphate beta (b-TCP) At lower temperatures,

no such peak appeared, indicating no transformation into

b-TCP during conversion of HAp unless heated above

1,100 �C. It is noteworthy that XRD analysis gave an

approximation amount related to the distribution of crystal

size. The variation in peak width illustrated a change in

crystal size distribution. In Fig. 3, the peak related to

b-TCP impurities emerged at 1,100 �C. Therefore, the best

temperature for scaffold construction was 900 �C.

In Fig. 3, the FT-IR of HAp, absorption bands were

visible in the middle of infrared waves. In Fig. 3a, a

powerful band at 1,034 cm-1, another band at 1,462 cm-1,

a poor band at 1,420 cm-1 and a sharp band at 604 cm-1

are evident. Recent FT-IR analysis of bone by Bosky and

Camacho demonstrated that phosphate and carbonate are

the core mineral components that correlate with Hap. A

series of absorption bands related to bone protein amide

groups are in the range of 800 and 1,800 cm-1 [51]. In the

FT-IR results for bone and bone ash powder, all organic

components of bone have been eliminated through pyro-

lysis. A small band associated with amide groups from

carbonate impurities (1,401, 1,450, 1,520 cm-1) were

visible in bone spectrum but not in the spectrum for bone

ash. The C–H absorption bands are also absent in the bone

ash spectrum. This evidence demonstrates that the pyro-

lysis of bone eradicates its organic components [52]; all

absorption bands in Fig. 3 belong to mineral components.

Figures 2e and 3e indicate that the best temperature for

preparing pure HAp was 900 �C.

Figure 4 shows TEM and SEM micrographs of HAp

nanocrystal powder. Particles are primarily pseudo-

spherical with a diameter of 70–80 nm; both of these

characteristics are preferred over larger or more irregular

morphology.

3.2 HAp scaffold with/without P3HB coating

Thermo gravimetric analysis (TGA) of sponge plays a vital

role in synthesising ceramics [53]. To prevent cracks in the

ceramic framework, enough time should be allowed for the

polymer sponge to burn completely. Figure 5 shows a

decrease in sponge weight with temperature. Most polymer

left the system at temperatures between 230 and 600 �C, so

it is necessary to consider slow heating rate in this range to

prevent structural degradation due to the departure of

gases. Polymer sponge analysis was performed in two

stages. The first stage ranges from 230 to 420 �C. A huge

decrease in weight due to gas production from the oxida-

tion of the polymer is observed. Weight loss in the second

stage of analysis from 420 to 600 �C was less significant.

The polymer sponge experiences little weight loss above

600 �C; loss that does occur can be attributed to oxidation

of carbon. The polymer sponge has been burnt completely

by 600 �C. To prevent thermal shock and crack develop-

ment, the temperature is increased at a rate of 3 �C/min to

600 �C and maintained for 1 h to burn the sponge

completely.

Figure 6 shows the XRD pattern of the HAp scaffold

with and without P3HB coating. In Fig. 6a, peaks related to

the P3HB are evident, while the pattern in Fig. 6b indicates

the process was mentioned in Sect. 2-4 on the ceramic

scaffold. Nanocomposite coating and the produced coated

polymer scaffold data are shown in Fig. 6c. According to

comparison with the standard JCPDS:9-432 scaffold after

coating, the angle peaks of 13.375� and 16.842� indicate

that the composite structure is indeed a nanocomposite

scaffold.

FT-IR spectra of pure polymer P3HB, nanocrystals of

HAp and HAp/P3HB nanocrystals were collected and

compared (Fig. 7). Spectra for pure HAp show clear peaks

for the apatite phase. The peaks in the 560–616 cm-1 range

are due to the bending vibration of PO4-3 bonds, and the

absorption at 1,020 cm-1 resulted from the stretching

vibration of PO4-3 bonds. The OH- groups of HAp are

evident in a wide peak in the range of 3,300–3,680 cm-1.

In the P3HB spectrum, 1,051, 1,126 and 1,172 cm-1 peaks

from the C–O bond of P3HB (1,126 and 1,172 cm-1 peaks

belong respectively to asymmetrical and symmetrical

stretching vibration of C–O bond) are evident. CH3 groups

display a sharp peak at 1,374 cm-1. A sharp and clear peak

from the carbonyl group stretching vibration is observed at

wave number 1,720 cm-1. As seen in Fig. 7c, a short peak

at wave number 1,717 cm-1 arises from the carbonyl group

of P3HB.Fig. 3 FT-IR spectrum of HAp at temperatures of, (a) bone’s ash,

(b) 600 �C, (c) 700 �C, (d) 900 �C and (e) 1,100 �C

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P3HB has hydroxyl end-groups; a broad peak for this

group can be observed in the range of 3,100–3,680 cm-1.

In order to precisely study the interaction of HAp and

P3HB, FT-IR spectra of the composite and of untreated

Hap from 550 to 650 cm-1 and FT-IR spectra of the

composite with pure polymer from 1,717 to 1,740 cm-1

are illustrated in Fig. 8. The carbonyl group absorption

peak at 1,724 cm-1 is sharper in the nanocomposite peak.

Fig. 4 a TEM of HAp

nanocrystals and b SEM of HAp

nanocrystals (10,0009)

Fig. 5 Weight loss as a function of temperature (heating

rate = 3 �C/min) for pyrolysis of polyurethane sponge

Fig. 6 XRD pattern of the, (a) P3HB, (b) HA scaffold and (c) HA/

P3HB nanocomposite according to standard of JCPDS: 9-432

Fig. 7 FT-IR absorption spectrum, (a) polymer P3HB, (b) HAp

scaffold and (c) HAp/P3HB scaffold

Fig. 8 Comparing of FTIR absorption spectrum, a HAp and HAp/

P3HB, b P3HB and HAp/P3HB

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This peak in the HAp/P3HB nanocomposite is shifted to a

lower wave number of 1,721 cm-1, possibly indicating

hydrogen bond formation between P3HB carbonyl groups

and HAp particles. Peaks associated with PO4-3 in both

states of stretching and bending vibration showed that the

low-resolution ratio of these peaks in nanocomposite is

against pure HAp. The wave numbers of these peaks for the

bending vibration are shifted from 574, 598 and 628 cm-1

in pure HAp to 566, 604 and 633 cm-1, respectively, in

nanocomposites (Fig. 8). In terms of PO4-3 stretching

peaks, a wavenumber of 1,025 cm-1 in pure HAp can be

compared to a nanocomposite wavenumber of 1,019 cm-1

attributable to the ceramic matrix of the scaffold. This

evidence demonstrates the presence of HAp in the com-

posite; the change in wavenumber may indicate hydrogen

bond formation among hydroxyl groups, HAp PO4-3 groups

and P3HB carbonyl groups.

Microstructural SEM studies were performed in order to

examine the outward profile and scaffold morphology with

and without P3HB coating (Fig. 10). Scaffolds without

polymer coating have greater porosity percentages than

scaffolds with polymer coating.

Figure 9a displays electron micrographs of polyurethane

with open pores in the range of 300–700 lm before

immersion in ceramic slurry. Figure 9b shows ceramic

porous scaffold whose porosity diameter ranged from 100

to 400 lm. Figure 9c illustrates micropores formed from

HAp particles; these micro-pores promote tissue growth.

As shown in Fig. 9d, following immersion of the scaffold

in the P3HB polymer solution, a P3HB layer on HAp

Fig. 9 a Polyurethane sponge’s image captured by SEM (509), b cross section of HAp scaffold without polymer coating (509), c view of

micro-porosities’ details (2,5009), d cross section of HAp with P3HB coating (1009) and e P3HB coating on the scaffold at (5009)

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particles is responsible for the scaffold’s high resistance

level; pore size has been reduced to 100–250 lm in

diameter. Figure 9e indicates homogenous P3HB coating

on the ceramic scaffold. FT-IR analysis (Fig. 9d, e) indi-

cates hydrogen bonds [54] between the polymer coating

and ceramic scaffold.

Table 1 indicates the average scaffold density and

porosity with and without P3HB coating; with increasing of

HAp and immersion time, the porosity percentage is

decreased. In a similar study [55], Chen et al. reported a

2.2% decrease in Bioglass macroporosity after coating with

PDLA polymer. The scaffolds gained approximately 12%

weight after coating with P3HB, which may increase

resistance and toughness.

3.3 Compressive strength of HAp scaffold with/

without P3HB coating

Compression impact tests were administered on two types

of HAp scaffold with and without P3HB coating. Results

are listed in Table 2; the compressive strength level of

ceramic scaffolds without polymer coating was 0.11 MPa,

while the compressive strength level of HAp scaffolds with

polymer coating was 1.55 MPa. Table 2 also shows that

increased immersion time increases the mechanical

properties of the scaffold. Ceramic scaffolds sometimes

develop cracks, contributing to a decrease in the mechan-

ical properties level of the scaffold; when immersed in

P3HB, all cracks became filled, and compressive strength

increased significantly.

In other research, Miao et al. coated PLGA onto TCP/

HAp ceramic scaffolds; the compressive strength level and

compressive module were 0.66 and 16.85 MPa, respec-

tively [33]. Tan et al. have coated PLGA-Bioglass onto

HAp ceramic scaffolds and reached compressive strength

and compressive modules of 1.36 and 24.58 MPa, respec-

tively [34].

As a general trend, the compressive strength of a cera-

mic scaffold depends on pore size and shape. In this study,

an approximate 1.5% reduction in pore size was achieved

through coating. Porosity of 80% and higher is desirable

because it allows for more interconnectivity and cell

migration [1]. According to Fig. 10, the best porosity and

mechanical properties were correlated with an immersion

time and HAp weight percent of 30 s and 50%,

respectively.

4 Conclusions

In this research, ceramic scaffolds with 40, 50 and 60%wt

of HAp were formed using a polyurethane sponge with a

porosity similar to natural bone. Scaffolds were coated with

P3HB to give higher compressive strength. Our findings

are as follows:

Pure HAp nanocrystal powder (grain size of about

70–80 nm) was used to create the scaffold following sin-

tering at 900 �C. XRD results for the coated scaffold

indicate angle peaks at 13.375� and 16.842�. A comparison

to the standard (JCPDS:9-432) indicates a nanocomposite

scaffold. FT-IR results for the HAp/P3HB scaffold indicate

a decrease in carbonyl peak resolution and a transition to a

lower wave number in the nanocomposite scaffold, as well

as a decrease in the PO4-3 absorption peak resolution and a

transition to lower frequencies. Hydrogen bond formation

among carbonyl groups of P3HB and PO4-3 and OH- of

HAp could explain these changes as well as the physical

merging of the scaffold. Mechanical and porosity

Table 1 Density and porosity percentage of 40, 50 and 60%wt HAp

scaffold with/without polymer P3HB

Sample 40%wt HAp 50%wt HAp 60%wt HAp

q (gr/cm3) e (%) q (gr/cm3) e (%) q (gr/cm3) e (%)

S1* 1.72 89 2.0 87 2.24 82

S2** 2.10 85 2.32 80 2.48 76

S3*** 2.31 81 2.48 77 2.80 69

* HAp scaffold** HAp scaffold coated by P3HB in 30 s*** HAp scaffold coated by P3HB in 1 min

Table 2 Impact of P3HB on compressive features of HAp (40, 50

and 60%wt) scaffold

Sample Mean

porosity

(%)

Mean

compressive

strength (MPa)

Mean

compressive

modulus (MPa)

S 1 86 0.11 ± 0.04 4.7 ± 0.01

S 2 80.3 1.46 ± 0.3 21.27 ± 0.5

S 3 75.6 1.55 ± 0.3 32.91 ± 0.8

Fig. 10 Comparison between porosity percentage and compressive

strength of HAp scaffold coated with P3HB in 30 s dip-coating time

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percentage tests showed that the best weight percentage of

HAp and immersion time are 50%wt and 30 s,

respectively.

Acknowledgments I would like to express my deep gratitude to

Prof. Hamdi, a faculty member of the University of Malaya and Dr.

Rabiee, a faculty member of Nooshiravaani university of Baabol for

their service and generous support which enabled me to conduct this

research.

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