Materials Technology

Advanced Performance Materials

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Effect of Zn/ZnO integration with hydroxyapatite:

a review

Varun Saxena, Abshar Hasan & Lalit M. Pandey

To cite this article: Varun Saxena, Abshar Hasan & Lalit M. Pandey (2017): Effect
of Zn/ZnO integration with hydroxyapatite: a review, Materials Technology, DOI:
10.1080/10667857.2017.1377972

To link to this article: http://dx.doi.org/10.1080/10667857.2017.1377972

Published online: 26 Sep 2017.

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 Materials Technology, 2017
https://doi.org/10.1080/10667857.2017.1377972

Effect of Zn/ZnO integration with hydroxyapatite: a review

Varun Saxena, Abshar Hasan and Lalit M. Pandey
Bio-Interface & Environmental Engineering Lab, Department of Biosciences and Bioengineering, Indian Institute of Technology, Guwahati,
India

ABSTRACT ARTICLE HISTORY

Hydroxyapatite (HAp) is one of the most studied ceramic for various biomedical applications Received 1 June 2017
such as bone tissue engineering, biologicals delivery systems and bioactive coatings. It Accepted 6 September 2017
owns extensive biocompatibility. However, lower mechanical properties for load bearing KEYWORDS
applications, lower antimicrobial activity and lower biological interaction rates are the major Zinc/Zinc oxide;
lags for biomedical applications of HAp. Various researchers have tried to integrate HAp with hydroxyapatite; mechanical
various metals and metal ions to overcome these holdups. In this review, we have described properties; antimicrobial
the crystal structure of both HAp and zinc oxide (ZnO) and have majorly focused on the zinc activities
and ZnO integration with HAp. Zn/ZnO offer several physical and biological properties. We have
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evaluated the effect of zinc integration over physical state and mechanical properties of HAp
with latest research examples. We have appraised the additional biological properties provided
by the incorporation zinc into the HAp through recent examples of researchers with varying
degree of successes.

Introduction
Substantial advances are under investigation in the
orthopedics. Researchers are more interested in the
development of new biomaterials, including metals
and metal oxides [1–4], that can substitute the dam-
aged bone/enamel. Depending upon the injury, various
metals and metal oxides are being utilised as either scaf-
fold or implant for the treatment. That either guides the
regeneration of tissues for the formation of new bone
or replaces/supports the completely damaged bone. The
nano materials are employed for various biomedical
applications such as bone tissue engineering [5], drug
delivery [6], antibacterial activity [7] and environmental
cleanup [8,9]. The nanosized materials possess unique
surface properties, which helps them in enhancing the
interfacial interactions at material-tissue interface.
Various materials are under investigation for ortho-
pedic applications [10–13]. Among various materials for
orthopedic applications, hydroxyapatite (HAp) remains
the most chosen material because of its chemical similar-
ity with bone. However, various properties such as medi-
ocre mechanical properties (Especially high brittleness
and subordinate fracture toughness), low dissolution
rate, lower antimicrobial activity and lower biological
interaction rates limits HAp for its orthopedic applica-
tion up to a distinct level. Hence, to widen the orthope-
dic applications of HAp chemical doping and composite
formation are the best techniques. The integration of
metal ions in the HAp alters the lattice parameters which
subsequently effects the crystallinity, chemical stability
and functionality [14]. The integration with various ions
i.e. Mg, Si, Fe, Zn, Se [15–17] has been proven successor
for the synthesis of biomaterials for orthopedic applica-
tions. The ease of synthesis remains the add-on-benefit
for the researchers. HAp is well-known to be synthesised
through various chemical routes. The substitution of
ions can easily be achieved through additional or altered
salts, while chemical synthesis [18].
These integrations not only alter the crystal struc-
ture, but also alter the in vivo and in vitro behaviours.
Various researchers have tried to dope and composite
the HAp with various cationic (Li, Mg, Zn, Ag, Cu, Al)
and anionic (carbonate, selenite, chlorine and fluorine)
ions with diverse level of successes for biomedical appli-
cations [17,19–24]. Various metal ions such as Zn, Mg
and Sr are found to enhance the osseointegration [25].
Among all these metal ions, doping of Zn has shown
better osseointegration along with the effective antimi-
crobial activity [26,27]. Integration of zinc with HAp
offers vast properties such as load bearing and corrosion
resistance [28], antibacterial and antifungal activities
[29–32] and applications including a better cell adhe-
sion [17], differentiation of mesenchymal stem cells [19].
In this review, we have focused and compared zinc
integrated HAp with enhanced physical and biological
properties than nave HAp. We will discuss about the
physical and biological effect of zinc integration to the

CONTACT  Lalit M. Pandey  lalitpandey@iitg.ernet.in

© 2017 Informa UK Limited, trading as Taylor & Francis Group
 2   V. SAXENA ET AL.
HAp, citing various recent examples with varying degree
of success.
Hydroxyapatite
The term apatite was coined from Greek word ‘apate’
which is defined as deceit because of the various colour
represented by different crystals. The variability in colour
was owned by the crystals because of different elemen-
tal configuration with the basic formula M10(ZO4)6X2,
where M represents variety of mineral such as Ca, Mg,
Fe and Cd, Z represents P, As, V, S, C, S and X represents
F, Cl, Br and OH [33]. Among all the apatite, calcium
hydroxyapatite is the most common apatite; which is a
phase of calcium phosphate. Calcium phosphate pos-
sesses variable phases depending on the Ca:P ratio.
These phases are listed in Table 1.
Calcium phosphate is the major constituent of hard
tissues such as bone, tooth enamel, dentine and calci-
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fied parts of tendons. Among all the phases of calcium
phosphate listed in Table 1, hydroxyapatite retains the
maximum similarity with the mineral component of
these tissues [34–36]. In addition, it owns substantial
bioactivity, biocompatibility, osteoconductivity and
facilitates direct bonding to the natural bone [35]. Pure
HAp (Ca/P molar ratio of 1·67) is thermally stable at pH
4–12 [36]. HAp triggers the biomineralisation phenom-
enon, by its chemical composition [37] and by altering
the surface potential [38]. This makes HAp the most
important material to understand the biomineralisation
phenomenon [39]. The HAp has been utilised for various
biomedical applications, such as scaffolds for orthopedic
applications [40–42], drug delivery [43–46], gene deliv-
ery [47–49] and environmental remediation [50–52].
Crystal structure of hydroxyapatite
Crystal shape, size, crystallinity, thermal stability, mor-
phology and solubility are the basic properties of a
material, which makes it efficient for a biomedical appli-
cation. HAp possesses two basic forms: monoclinic or/
and hexagonal crystal structure. The basic crystal of HAp
is represented in Figure 1.
The orientation of OH group decides the final crystal
structure. If adjacent OH groups are pointed in oppo-
site direction, HAp carries hexagonal crystal structure,
whereas in monoclinic orientation adjacent OH groups
are in the same direction and the direction converses
in adjacent column [53]. The basic crystal properties
Table 1. Variable phases of HAp.
S. No. Ca/P ratio General name
1 0·5 Monocalcium phosphate
2 1·0 Dicalcium phosphate
3 1·33 Octacalcium phosphate
4 1·5 Tricalcium phosphate
5 1·67 Hydroxyapatite
of both hexagonal and monoclinic forms are listed in
Table 2.
The material science researchers are continuously
endeavouring to mimic natural bone composition by
supplementing/doping the HAp with various elements.
These substitutions change the lattice parameters
depending upon the site of substitution. The substitu-
tion can occur at either calcium or phosphate point. For
example, substitution of Ca ions with Fe ions causes
distortion in the lattice parameters a and c and conse-
quently grounds the crystal properties [54]. The most
common substitution is carbonate substitution, which
improves bioactivity and solubility [55]. Depending
upon the substitution, carbonated apatites are of two
types: A (CO32− ions substitute for OH- ion) and B
(CO32− ions substitute for PO43− ions). The imbalanced
charge distribution, due to the change in the stoichiome-
try of the compound, causes the impurities in the crystal
lattice [56]. In general, cations substitute the Ca2+ ion,
SiO4 replaces the PO4 tetrahedra and anions replace the
OH- group in the basic crystal lattice [57]. As discussed
earlier the carbonate ion can replace both the hydroxyl
and the phosphate groups.
The lattice parameters governs various properties of
the hydroxyapatite and thus changes in these parame-
ters often alters various physical (crystallinity, thermal
stability, morphology), chemical (solubility), and other
physicochemical and biological properties [58]. Hence,
the routes of the synthesis, type of ion substitution, con-
centration of the ion substitution are the basic param-
eters, which usually govern the application of HAp by
tuning the lattice parameters.
Organic/inorganic hybrid network structures of
nanocomposite scaffolds have been widely explored for
bone tissue engineering. Various compositions have been
traversed to mimic natural bone. W.W. Thein-Han et al.
2009 studied structure–property–process relationships
between the pure chitosan and chitosan-nano HAp. It
was reported that 1 wt% HAp concentration enhanced
the compression modulus from 6·0 to 9·2 kPa as required
for bone tissue engineering [59,60]. Similar approach
was progressed by Nagini Maganti et al. 2011. The com-
pression modulus of 95% deacetylation chitosan-5wt%
nano-HAp scaffold was enhanced from 6 to 9·2 kPa [61].
Chitosan and ethylene glycol functionalised nano-HAp
was also approached to from organic/inorganic network
structure. These three-dimensional scaffolds exhibited
high modulus, controlled swelling behaviour, reduced
Formula Applications
Ca(H2PO4)2 Fertilizers, leavening agent
CaHPO4 Food additive
Ca8(HPO4)2(PO4)4 Precursor of HAp
Ca3(PO4)2 Anticaking agent
Ca10(PO4)6(OH)2 Orthopedic and other
biomedical
 MATERIALS TECHNOLOGY   3
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Figure 1. Crystal structure of Hap.

Table 2. Hexagonal and monoclinic crystal structure of Hy- possess three inherent mechanisms for antimicrobial
droxyapatite. action predominantly by Reactive Oxygen Species (ROS)
Properties Hexagonal Monoclinic generation, by attack to nucleus and protein and by the
Space group symmetry P63/m P21 /b disruption of cell wall. These modes of actions differ
Lattice parameter a= b=9·432 Å, c= a=9·4214 Å, b=2a,
6·881Å, γ=120° c=6·8814 Å, γ=120°
from the mechanism of the resistance generation by the
Sintering temperature Lower temperature, Elevated temperature, microbes i.e. efflux, drug modification, target modifi-
ambient (25°C) to 850°C cation and enzyme deactivation. Hence, ZnO NPs are
100°C
Details Axial Ca ion holds Adjacent OH groups considered as the best suitable nanoantibiotic for anti-
the tetrahedral of are in the same biotic resistant bacteria.
PO4. Adjacent OH direction
groups in opposite
direction.
Crystal structure of ZnO

water uptake and better osteogenic activities [62]. These
studies suggest the potential use of chitosan and nano-
HAp based composite/structure for bone tissue engi-
neering. However, elemental dissimilarity remains the
major hunt.
Zinc oxide
Zinc is another most common mineral element in hard
tissues. Zinc has various physiological roles in immune
system and serves in cell division and cell growth. In
a study, the deficiency of zinc was found involved in
Alzheimer’s and Parkinson’s disease [63]. In addition,
zinc also act as an antidepressant [64]. Zinc oxide (ZnO)
is a wide band-gap II–VI binary compound semicon-
ductor (3·36 eV). It possesses unique optoelectrical and
chemical properties. ZnO is a superlative material for
nanoscale optoelectronics and piezoelectric nanogen-
erators [65]. ZnO helps in bone regeneration as well as
it possesses antimicrobial activity [66,67] with enhanced
mechanical properties [68]. ZnO nanoparticles (NPs)
The crystal structures of ZnO mainly possess three ori-
entations: hexagonal wurtzite, cubic zinc blende and
rocksalt. Rocksalt is also named as rochelle salt. The most
common and thermodynamically stable orientation is
wurtzite. In this crystal structure, sp3 hybridisation gov-
erns the whole structure with each O2− remains fenced
by four Zn2+ at the junctions of a tetrahedron and vice
versa. Cubic substrate nurtures the zinc blende structure
at consecutive growth. Synthesis of ZnO at higher pres-
sure provides the Rochelle salt symmetry [69].
The lattice parameters a  =  0·32495  nm and
c = 0·52069 nm remain in the ratio of c/a = 1·602 with
P63/m space group in hexagonal wurtzite crystal [69].
Zn2+ remains at the corner of tetrahedron and covers
the O2−. This arrangement of ions provides a typical sp3
covalent hybridisation. This sp3 hybridisation subse-
quently provide the valance band and the antibonding
counterpart provides the conduction band [70]. This ori-
entation is achieved by interconnecting two sub frames
of Zn2+ and O2−. ZnO carries polarity along hexagonal
axis. This polarity is administered by the hexagonal
 4   V. SAXENA ET AL.
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Figure 2. Hexagonal wurtzite structure of ZnO.
symmetry. The two major properties retained by ZnO i.e.
piezoelectricity and impulsive polarisation are due to the
hexagonal wurtzite crystal structure [70]. As represented
in Figure 2, the overall crystal structure of ZnO is the
result of alternating tetrahedral planes of O2− and Zn2+
ions, which remains slanted along the c axis.
ZnO lattice carries a dipole moment originated due to
the polar surfaces. These polar surfaces are produced due
to the charge carriers i.e. oppositely charged ions. The
Zn ion produces the positively charged surface (0001)
and the oxygen ion produces the negatively charged sur-
face (0001−). This dissimilarity in charge produces both
the dipole moment and polarisation along c axis, which
consecutively diverges the surface energy [71]. These
polar faces due to electrostatic forces are more stable
than the nonpolar faces. ZnO crystal also carries various
point defects in the lattice. These defects may be either a
vacancy or interstitial defect. In some cases, these defects
may extend in form of threading and planer disruption.
This results in a non-stoichiometric crystal with Zn1+dO
configuration, where d refers excess zinc atoms. These
defects are the main factors to produce N type semi-
conductor ZnO, with excess zinc, as these excess zinc
act as a donor site. In ionic form, the interstitial sites
are occupied by these Zn2+ ions with Miller index (⅓,
⅔, 0·875) [72]. Wurtzite is the most common form of
ZnO, which is thermally stable. Other forms such as
zinc blende and Rochelle salt symmetries are formed
in special conditions. Atomic terminations provide zinc
oxide, which is a wide range of novel structures. A num-
ber of profound factors or kinetic parameters including
temperature and pH can tune the morphology and the
growth rate of the lattice. These parameters govern the
surface activities of the growth facets and consequently
alter the overall crystal structure. Thus, a well-defined
crystalline growth takes place after a proper nucleation.
These nucleation points define the overall morphology
of the synthesised ZnO.
Antimicrobial mechanism of ZnO
ZnO nanoparticles show broad spectrum of antibac-
terial activities. A bacterial cell protects itself from the
action of drugs with the help of membrane followed
by cell wall. This cell wall remains the outer most part
of the bacterial cell and is typically made up of pepti-
doglycans (amino acids and sugars). The bacterial cells
protect themselves by regulating the permeability of the
cell membrane towards drugs and other organic and
inorganic molecules. In this way, Gram negative bacte-
ria are more resistant than the Gram positive bacteria,
as they possess an outer membrane followed by the
plasma membrane [73,74]. Thickness of the cell mem-
brane remains 20–80 nm. The cytoplasm of the bacte-
rial cell majorly encompasses biomolecules including
certain ions and water, which offers conductivity to
the cellular structure and generates an overall nega-
tive charge over bacterial cell [27]. An agent that kills
the bacterial cell is known as bactericidal agent, in the
similar way, if it inhibits the cellular growth; the term
is given as bacteriostatic. The respective concentrations
are known as Minimum Bactericidal Concentration
(MBC) and Minimum Inhibitory Concentration
(MIC).
The nanoparticles of small size can pass through the
cell wall and consequently can damage the microorgan-
ism [74]. Several studies have confirmed the antibacte-
rial activity of the ZnO NPs [26,75–79]. In general, ZnO
NPs breakdowns the bacterial cell wall by disrupting
membrane protein and lipids and causes the outflow of
the cellular components. Although, the exact mecha-
nism of antimicrobial activity of ZnO is under investi-
gation by many researchers, three major mechanisms,
membrane damage, cellular intake of ZnO NPs or Zn2+
ions and reactive ion species (ROS) production are the
possible modes of action. Figure 3 explains the antimi-
crobial activity of the ZnO NPs.
Membrane damage
The membrane damage is caused by the electrostatic
attraction between ZnO and bacterial cell. This results
in the specificity of ZnO to kill only microbial cells. The
cellular permeability for ZnO is enhanced by the mem-
brane blebbing produced due to the crystal defects of
ZnO [80].

Figure 3. Various mechanisms of antibacterial activity of ZnO nanoparticles.
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Release of zinc ions
The released Zn2+ ions inhibit the bacterial cell growth
by disrupting the cellular lipids and proteins. The dis-
ruption in the protein and lipid structure punctures the
cell wall resulting in bacterial cell death. Several studies
have focused on the ZnO toxicity present in the growth
media. This mode of action is generally size depend-
ent and occurs in ZnO NPs only and not in micro or
macro molecular structure of the ZnO. Ananth et al.
2017 proved that discharge of Zn2+ ions is one of the
major aspects of the bactericidal activity of ZnO against
E. coli, S. iniae, S. parauberis and E. tarda bacteria [81].
This cytotoxicity mode of Zn2+ ions also depends on the
morphology of the ZnO crystal.
Generation of ROS
This is one of the most observed antibacterial mecha-
nism of ZnO. Reactive Oxygen Species (ROS) such as
OH-, H2O2 and O2− are produced under both UV and
visible light [80,82], which damage the cell membrane.
The antibacterial activity depends on high reactivity
and oxidising property of ROS. Sarwar et al. studied the
antibacterial activity of ZnO against V. cholera and con-
cluded that reactive species disintegrate the components
of cell membrane such as proteins and lipids, damage to
DNA. The disintegration of cell membrane causes the
outflow of cellular components [83].
Factors affecting antimicrobial mechanism of ZnO
Various factors affect the antimicrobial effect of the
ZnO. These factors comprise the UV illumination effect,
morphology, particle size, concentration and the crystal
defects of the ZnO. These parameters affect the antimi-
crobial activity of the ZnO by various mechanisms. An
exposure of UV light has been proved to enhance the
antimicrobial effect by activating crystal defects [65].
The effect takes place via excitation of electrons, which
MATERIALS TECHNOLOGY   5
enhances the charge distribution between the conduc-
tion and the valance band resulting in the generation
of more ROS. In the similar way, various morphologies
affect the antimicrobial activity of the ZnO based on the
contact points between the cell membrane and the ZnO.
The antimicrobial activity of the ZnO depends on
the particle size, which in turn regulates the Zn2+ inter-
nalisation [79,84–86]. Table 3 summarises various mor-
phologies, synthesis procedures and the antimicrobial
effect of ZnO.
Bacterial cells have various defence mechanisms
to overcome the external environmental factors. For
example, some bacteria produce defensive proteins
under stress conditions. Campylobacter species pro-
duce a number of genes/proteins, which protect cells
from diverse stresses. Majorly this defence mechanism
is observed against oxidative stress. ROS are removed
by the utilisation of antioxidant enzymes. Molecular
oxygen produces superoxide ions O2− upon reduction.
These superoxide ions are converted into hydrogen per-
oxide and diatomic oxygen using superoxide dismutase
[85,87]. This hydrogen peroxide is removed by utilising
Catalases enzyme which degrades H2O2 into water and
oxygen [88]. In Catalase mutants, such as mutants of
E.coli, H2O2 is removed by alkyl hydroperoxide reduc-
tase (AhpC) [89]. This AhpC also repairs the oxidative
mutilation to molecules [90,91]. Besides these mecha-
nisms, various essential molecular chaperons prevent
protein aggregation and refolding [92].
Various experiments prove that the ZnO nanopar-
ticles cause oxidative stress in bacterial cells to kill the
bacteria. Xie et al. examined the effect of ZnO over C.
jejuni, and concluded that ZnO down-regulated various
virulence factors, which governs the defence mechanism
of bacteria such as toxin production, mobility and host
cell adhesion. Hence, the antibacterial mechanism of
the ZnO is found to be effective over bacteria’s defence
mechanism [93,94].
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6 

Table 3. Morphologies, synthesis procedure and the antimicrobial effect of ZnO.

S.N Crystallinity Morphology Synthesis method Reaction temperature Particle size (nm) Microbes MIC Reference
1 Hexagonal wurtzite ZnO-1 Purchased NA 14·7 nm MIC in wt%/MBC in % w/w [111]
Platelet E. coli 0·25/1·44 wt%
S. aureus 0·88/1·72 wt%
P. aeruginosa 1·40/1·81 wt%
A. brasiliensis >8/>8 wt%
 V. SAXENA ET AL.

C. albicans Visual
ZnO- 2 Purchased NA 17·5 nm E. coli 0·38/1·56 wt%
Platelet S. aureus 1·68/3·18wt%
P. aeruginosa 5·50/7·52wt%
A. brasiliensis >8/>8wt%
C. albicans Visual
ZnO-3 Purchased NA 76·2 nm E. coli 4·00/6·40 wt%
Rod like S. aureus 4·50/6·81 wt%
P. aeruginosa 6·46/> 8 wt%
A. brasiliensis >8/>8 wt%
C. albicans Visual
2 NA Nanoparticles Purchased NA 10–50 nm P. gingivalis MIC- 437·5 μg/mL, MBC- [112]
F. nucleatum 500 μg/mL
P. intermedia
A. actinomycetemcomitans
3 Hexagonal wurtzite Nanoparticles Surfactant assisted precip- Room temperature 12·84 Bacteria, reduction % [113]
itation E. coli 91% reduction
S. aureus 89% reduction
Coating of ZnO by ultra- Fungus, reduction %
sonication C. albicans 91% reduction
M. canis. 91% reduction
4 NA Spherical Precipitation method Room temperature 10 Vibrio cholerae At 25 μg/mL growth [83]
inhibition 80% in 6 h
5 Hexagonal Wurtzite Nanoparticles Non-hydrolytic solution 65°C 20–30 S. aureus 15 μg/mL [114]
process E. coli 15 μg/mL
S. typhimurium 15 μg/mL
K. pneumoniae 5 μg/mL
6 Hexagonal wurtzite Nanoparticles sol–gel combustion sol–gel- 100°C 19·89 ± 1·43 nm MBC (μg/mL) [115]
E. coli 18 ± 0·5
Combustion- 200°C P. aeruginosa 14 ± 0·6
S. aureus 16 ± 0·2
B. subtilis 12 ± 0·5
7 Hexagonal Wurtzite Nanoparticles Precipitation method Room temperature 12 nm E.coli 1 mM (80 μg/mL) MIC [65]
5 mM (400 μg/mL) MBC
8 Hexagonal Wurtzite 1D nanowire Hydrothermal 90°C 30 nm Dia S. aureus 87·5 μg/mL [116]
350 nm length P. aeruginosa 137·5 μg/mL
E. coli 125 μg/mL
9 NA 1D nanowire vapour transport process ZnO (A)- 430°C 150 nm Dia E. coli 83·3 ± 8·5% survival [117]
S. aureus
ZnO (B)- 560°C 150 nm Dia E. coli 31·8 ± 16·7% survival
S. aureus
ZnO-(C)- 560°C 150 nm Dia E. coli 17·4 ± 5·8% survival
S. aureus
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10 Hexagonal Wurtzite nanorod arrays Hydrothermal NA 30–70 nm Dia MBC (μg/mL) [118]
E. coli 1221 μg/mL
∼250 nm length B. atrophaeus 407 μg/mL
11 Hexagonal wurtzite nanorod Ultrasound assisted pre- NA 100 nm (Zone of inhibition at 80 μg/mL concentration) [119]
cipitation method E. coli 12·1 mm
P. aeruginosa 12·1 mm
S. aureus 9·7 mm
B. subtilis 21·8 mm
C. albican 11·4 mm
12 Hexagonal wurtzite nanorods Precipitation method Room temperature 90 nm E. coli 512 μg/mL [120]
S. aureus 256 μg/mL
13 Hexagonal wurtzite Precursor dependent Soft jet plasma generated 27°C ZnO RO- 100 nm Dia E. coli 3·1 μg/mL [121]
S. iniae 1·6 μg/mL
S. parauberis 1·6 μg/mL
Nanorods (ZnO RO) E. tarda 6·3 μg/mL
ZnO FL- >1000 nm Dia E. coli 3·1 μg/mL
Micro flower-like (ZnO FL) S. iniae 0·8 μg/mL
S. parauberis 1·6 μg/mL
E. tarda 6·3 μg/mL
Spherical (ZnO-SP) ZnO-SP- 150 nm Dia E. coli 6·3 μg/mL
S. iniae 12·5 μg/mL
S. parauberis 12·5 μg/mL
E. tarda 12·5 μg/mL
Layer structures (ZnO SH) ZnO SH- 500 nm Dia E. coli 12·5 μg/mL
S. iniae 12·5 μg/mL
S. parauberis 12·5 μg/mL
E. tarda 25·0 μg/mL
14 Hexagonal wurtzite Nanorods Solvothermal 150°C 10~20 nm Dia S. mutans MIC 512 μg/mL [122]
300–500 nm length MBC >1024 μg/mL
15 Hexagonal wurtzite Flower like Hydrothermal 90°C 400 nm Dia Wt % of Zn for zero survival ratio (in %) of bacteria [123]
E. coli 4 wt%
3 μm length S. aureus 4 wt%
16 Hexagonal wurtzite Mulberry like Sonication followed by 80°C 150 nm Inhibition Rate [124]
Sheet like, Hydrothermal 100 nm C. albicans Mulberry like 95%
Flower like C. albicans Sheet like 85%
C. albicans Flower like 50%
17 Hexagonal wurtzite Flower Cooling followed by pre- Ice cooled and heated 800 nm length E. coli MIC/MBC [125]
cipitation and heating at 75°C S. aureus 25 μg/mL
18 Hexagonal Wurtzite micro-flowers Precipitation 90°C 150–200 nm Dia S. aureus 5 μg/mL [126]
E coli
S. typhimurium
2–3 μm length K. pneumoniae
19 Hexagonal wurtzite flower-shaped Surfactant assisted sol-gel 55°C ZnO NPs 69·6 nm Zone of inhibition at 500 μg/mL [127]
E. coli 22 nm
S. aureus 19 mm
C. albicans 20 mm
MATERIALS TECHNOLOGY 
 7
 8   V. SAXENA ET AL.

Table 4. Mechanical properties of Hap and Zn incorporated HAp/HAp composites.

S.No. Mechanical properties HAp Zn/HAp Zinc percentage and composite References
1 Relative density ρr 97·2 ± 0·4% 96·1 ± 0·5% 5 mol % (sintered at 950°C) [98]
Elastic modulus (E) (GPa) 173 ± 17 58 ± 2
Reduced modulus (Er) (GPa) 162 ± 14 60 ± 2
Hardness (HV) (GPa) 13·6 ± 2·2 2·05 ± 0·2
Fracture toughness (K1c) (MPa m1/2) 0·5 ± 0·1 1·5 ± 0·5
Brittleness index B (μm−1/2) 25·6 ± 4·1 1·33 ± 0·4
2 Densification (%) 91·2 95·3 2·5 wt% (selective laser Sintering, 12 W) [128]
Compressive strength (MPa) 3·09 17·89
Stiffness (MPa) 112·86 313·48
Fracture toughness (MPa m1/2) 1·09 1·40
Microhardness (GPa) 3·51 4·20
3 Bulk modulus (Gpa) 79·488 79·874 2·5 wt% (calculated data) [129]
Shear modulus (GPa) 41·734 42·019
Young’s modulus (GPa) 106·554 107·249
4 Vickers microhardness (Gpa) 5·83 ± 0·06 5·76 ± 0·17 2 mol % [130]
Fracture toughness (MPa m1/2) 1·12 ± 0·16 1·05 ± 0·10 Ca/P- 1·63 (sintered at 1100°C)
5 Vickers Hardness (GPa) 4·00 4·76 0·5 wt% (sintered at 1200°C) [131]
Fracture toughness (MPa m1/2) 0·98 1·34

Effect of zinc integration to HAp over its mechanical strength. Higher mechanical strength and
mechanical properties and microstructure the reduction in the lattice parameter c of HAp doped
Downloaded by [La Trobe University] at 06:26 30 September 2017

with Zn ion are also reported recently (2017) by other

As mentioned earlier, HAp possesses high brittleness
groups [101,102]. Table 4 summarises various mechan-
and lower fracture toughness. The bioapatites are having
ical properties of HAp and Zn incorporated HAp.
better biological and indentation properties than HAp
Various studies are ongoing to reduce the formation
under physiological conditions. This prompts various
of β-TCP in the HAp doped with zinc. In this contrast,
researchers to focus on the substitution of HAp with
Huang et al. (2016) co-doped Zn2+ ions with F- ions and
various metal ions, which subsequently can improve
co-doping inhibited the formation of β-TCP phases at
these lags for biomedical applications. Incorporation of
200°C. It was concluded that the mechanical and chemi-
Zn enhanced the mechanical properties of HAp [95].
cal stability of co-doped HAp is higher than that of pure
These substitutions revised the lattice parameters, which
HAp at physiological conditions [103].
altered the crystallinity as well as the physical properties
Summarily, we can conclude that the doping of zinc
of HAp [96]. The hexagonal crystal structure of HAp
alone do not play a role in the generation of other phases
allows various metal ions either to replace the calcium
such as β-TCP. Zn doped HAp when sintered at higher
ion from the HAp lattice or to sit in the void area. The
temperature such as 900°C, showed the other phases.
chemical charge imbalance due to the monovalent and
Iconaru et al. 2017 sintered the Zn doped HAp at 500°C
trivalent doped ions causes the changes in the chemi-
and Ran et al. 2017 did not sinter the composite. The
cal properties. Although, doping of divalent ion do not
major phase as HAp along with the dopant phase was
cause the charge imbalance, yet the size of the metal ion
observed in both the studies, with a reduction in c lat-
plays an important role. Ionic radius of zinc (0·074 nm)
tice parameter, indicating that only zinc doping does not
are smaller than calcium (0·099 nm). Hence, incorpo-
disperse the HAp into other phases. However, sintering
ration of zinc into the HAp lattice declined the c lattice
temperature also plays an important role in the HAp
parameter and enhanced the lattice parameter, which
phase generation during synthesis.
decreased the crystallinity of HAp with increase in the
zinc concentration [14,97]. Zn doped HAp shows dif-
ferent mechanical behaviour due to two reasons: first Effect of zinc integration to HAp over
the difference in the size of Zn and Ca and second the biological properties
different melting temperatures of Ca and Zn i.e. 842°C Integration of zinc not only alters the physical and
and 419·5°C, respectively. mechanical properties of HAp, but also adds various
In a recent study, for 10 h milled sample of 5 mol% biological activities to the HAp matrix. These biologi-
doped Zn transformed 65% of HAp into the amorphous cal functionalities open a vast area towards the biolog-
phase [98]. This process reduced the elastic modulus ical applications of HAp. The detailed discussions are
(E), reduced modulus (Er), hardness (Hv) and brittle- as follows.
ness index (B), and in enhanced the fracture toughness
(k1c). β-TCP (β-tricalcium phosphate) reflections were
Biological responses
also seen in the XRD pattern along with HAp reflections
after sintering around 900°C [99,100]. The reductions in Incorporation of zinc enhances the biological responses
a and c lattice parameters were also observed. This con- of HAp. Ibrahim et al. 2017 studied the corrosion behav-
firms that the mechanical alloying transforms a portion iour of heat-treated Mg-1·2Zn-0·5Ca alloy under physio-
of crystalline HAp into amorphous HAp and enhances logical conditions. A lower corrosion rate (8·9 mm/year)

for the heat-treated alloy was observed than that of pure
metal. The immersion test revealed the heat induction
can control the resorption time. The heat treated alloy
for 3 h was reduced 44·1% after 28 days of immersion
in modified seminal body fluid (SBF); while untreated
alloy were completely degraded [95].
Begam et al. 2017 studied the MG63 osteoblast cell
response to the zinc integrated HAp [17]. The study
revealed that in one hour around 42% MG63 cells
attached over the zinc integrated sample and around 35%
MG63 cells attached over the pure HAp when sintered
at 1100°C. Similarly, an increase in the cells attachment
(53%) was observed in one hour for the doped sample
sintered at 1250°C. It was also concluded that the cell
viability and cell proliferation remained higher for the
samples sintered at higher temperature. Various surface
properties such as grain size, surface area and porosity
depends on the sintering temperature, which in turn
regulate the cellular interactions [17]. The same group
Downloaded by [La Trobe University] at 06:26 30 September 2017
studied the bone morphogenetic protein interaction
with zinc doped HAp. The proliferating osteocytic mass
with higher cellular reaction and without any inflam-
matory response was reported. The woven bony matrix
with a proper differentiated haversian system was also
observed. These findings indicate the zinc doped HAp
as a promising biomaterial for bone regeneration [104].
Bostancioglu et al. 2017 studied the mesenchymal
stem cell adhesion and differentiation over zinc doped
hydroxyapatite nano-coated surfaces. The study revealed
the higher osteogenic activity and alkaline phosphatase
activity in doped sample along with the higher expres-
sion of osteopontin gene [105]. Ahmadzadeh et al. 2016
showed the similar results in vivo studies in a rabbit tibial
osteotomy model. The biocompatibility and osteocon-
ductivity were significantly higher for zinc integrated
HAp graft [106]. All these studies favour that incorpo-
ration zinc in HAp matrix enhances the osseointegra-
tion and biological responses of HAp, thus opens the
door for various orthopedic as well as other biomedical
applications.
Antibacterial activity
As discussed in section 3·2, zinc and ZnO possesses
excellent antimicrobial activity against various patho-
gens. Among all these pathogens, zinc shows highest
lethal activity against both Gram-positive and Gram-
negative bacteria. In this section, we will discuss few
of the recent studies, which prove that the incorpora-
tion of zinc into HAp enhances its antibacterial activity.
Predoi et al. 2017 explained the antibacterial activity of
zinc doped HAp against both Gram-positive and Gram-
negative bacteria (S. aureus and E. coli) at lower concen-
tration of zinc, (Ca + Zn)/p = 1·67. The concentration
of zinc was 0·01<xzn  <0·05. It was concluded that the
antimicrobial activity of the synthesised material also
MATERIALS TECHNOLOGY   9
depends on colloidal dispersions of zinc ions apart from
its concentration [96].
As discussed in section 4, higher concentration of
zinc may introduce some unwanted phases such as
β-TCP and also reduces the crystallinity of the doped
HAp. Hence, the lower concentration of zinc is preferred.
Barros et al. 2017 studied the adhesion of S. aureus and
E.coli in presence of zinc doped HAp, and the adhesion
was higher for the former. Interestingly the reduction
in the adherent cells was found 90% in both cases. This
concludes the zinc doped HAp as a promising alternative
to antibiotics, which subsequently reduces the chances
of antibiotic resistance [107].
Huang et al. 2017 described the antibacterial activity
of Cu–Zn co-doped HAp. It was reported that Cu alone
showed significant antibacterial activity, but the higher
concentrations were found lethal to the human cells.
Hence, the Zn ion was co-substituted along with Cu,
which was found highly toxic to the bacterial cells but
was biocompatible to the human cells [108]. Anwar et al.
2016 studied the antibacterial activity of zinc incorpo-
rated HAp against B. Subtilis, S. aureus, E. aerogenes and
E coli. It was found that 2 wt% doping of Zn was more
lethal to all the bacterial strains than 0 wt% and 4 wt%
of Zn doping [109]. Cai et al. 2015 studied the antibac-
terial activity of mesoporous hydroxylapatite/zinc-oxide
nanofibers. It was found that 1:10 (w/w) ratio of ZnO/
HAp was lethal for S. aureus and 1:5 (w/w) ratio of ZnO/
HAp was lethal for E.coli [110]. Hence, from these stud-
ies it is concluded that even a lower concentration of zinc
is antibacterial when incorporated into the HAp matrix.
Antifungal activity
In addition to the antibacterial activity, Zn incorpo-
rated HAp has also shown excellent antifungal activity.
Such vast antimicrobial activity attracts the biomaterial
researchers to explore the all possible biomedical appli-
cations of Zn incorporated HAp. Iconaru et al. 2017
explored the antifungal activity of zinc doped HAp
against Candida albicans. It was found that neither under
dark conditions nor in day/UV light HAp coating over
Ti or Si could show antifungal activity. However, a sig-
nificant decrease in the fungal cells was observed under
day/UV light condition when zinc was incorporated into
the HAp matrix. But under dark conditions even zinc
incorporation could not show the antifungal activity
[102].
In the similar way, Groza et al. 2016 studied the anti-
fungal activity of zinc doped HAp. The antifungal activ-
ity of both Ag and Zn doped HAp was explored. The
fungicidal effects of Ag doped HAp and the potential
antifungal effect of Zn doped HAp against C. albicans
biofilm were observed. The biofilm was diminished over
time in the presence of Zn doped HAp and a 72 h of
incubation time was sufficient to decrease the fungal
 10   V. SAXENA ET AL.
cells significantly [30]. These indicate the potential use
of zinc doped HAp as an antifungal agent.
 
Conclusion
The nano-sized material contains higher surface to vol-
ume ratio, which makes them ideal for various biomedi-  
cal applications such as tissue engineering, drug delivery
and bio-imaging. The antimicrobial activity of the zinc
nanoparticles is size and morphology dependent. The
incorporation of Zn/ZnO into HAp matrix alters various
lattice parameters and subsequently alters the physical  
and biological behaviour of HAp. Incorporation of zinc
not only ameliorates the mechanical strength of the
HAp, but also enhances the biological responses, anti-  
bacterial activity and anti-fungal activity. In addition to
these properties, integration of zinc controls the bio-
degradation rate of HAp. Although zinc incorporated  
Downloaded by [La Trobe University] at 06:26 30 September 2017
HAp has been studied widely for various biomedical
applications; still many of its applications are uncharted.
These unexplored applications include: the antifungal  
activity in dark, anti-cancerous activity, opto-electrical
properties and application, coating material with lower
corrosion rate, etc. Hence, these properties open a new  
field to the researchers to discover the more biomedical
applications of zinc integrated HAp.
 
Acknowledgements
The authors would like to thank the Department of Science  
and Technology, Government of India for financial sup-
ports (Sanction nos: DST/INSPIRE/04/2014/002020,
ECR/2016/001027).
 
Disclosure statement
No potential conflict of interest was reported by the authors.
 
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