Review Article Substituted Hydroxyapatites with Antibacterial Properties - Hindawi

16 Pages·2015·2.71 MB·English

Hindawi Publishing Corporation
BioMed Research International
Volume 2014, Article ID 178123, 15 pages
http://dx.doi.org/10.1155/2014/178123
Review Article
Substituted Hydroxyapatites with Antibacterial Properties
JoannaKolmas,EwaGroszyk,andDagmaraKwiatkowska-Róhycka
DepartmentofInorganicandAnalyticalChemistry,FacultyofPharmacy,MedicalUniversityofWarsaw,
Ul.Banacha1,02-097Warsaw,Poland
CorrespondenceshouldbeaddressedtoJoannaKolmas;[email protected]
Received28February2014;Accepted14April2014;Published11May2014
AcademicEditor:GiuseppinaNocca
Copyright©2014JoannaKolmasetal.ThisisanopenaccessarticledistributedundertheCreativeCommonsAttributionLicense,
whichpermitsunrestricteduse,distribution,andreproductioninanymedium,providedtheoriginalworkisproperlycited.
Reconstructivesurgeryispresentlystrugglingwiththeproblemofinfectionslocatedwithinimplantationbiomaterials.Ofcourse,
thebestantibacterialprotectionisantibiotictherapy.However,oralantibiotictherapyissometimesineffective,whileadministering
anantibioticatthelocationofinfectionisoftenassociatedwithanunfavourableratioofdosageefficiencyandtoxiceffect.Thus,the
presentstudyaimstofindanewfactorwhichmayimproveantibacterialactivitywhilealsopresentinglowtoxicitytothehuman
cells.Suchfactorsareusuallyimplementedalongwiththeimplantitselfandmaybeanintegralpartofit.Manyrecentstudieshave
focusedoninorganicfactors,suchasmetalnanoparticles,salts,andmetaloxides.Theadvantagesofinorganicfactorsincludethe
easewithwhichtheycanbecombinedwithceramicandpolymericbiomaterials.Thefollowingreviewfocusesonhydroxyapatites
substitutedwithionswithantibacterialproperties.Itconsidersmaterialsthathavealreadybeenappliedinregenerativemedicine
(e.g.,hydroxyapatiteswithsilverions)andthosethatareonlyatthepreliminarystageofresearchandwhichcouldpotentiallybe
usedinimplantologyordentistry.Wepresentmethodsforthesynthesisofmodifiedapatitesandtheantibacterialmechanismsof
variousionsaswellastheirantibacterialefficiency.
1.Introduction Due to its poor mechanical properties (brittleness and
inflexibility), the use of hydroxyapatite as a substitute bone
Calcium phosphates—mainly hydroxyapatites—have for materialisunfortunatelylimitedtoplacesthatarenotsubject
many years played a key role in biomaterial engineering togreattension.Ontheotherhand,ahydroxyapatitematerial
due to their high biocompatibility and their bioactivity in isaperfectcomponentofcompositeimplantswithsynthetic
human mineralized tissues, especially in bones and dental polymers and biopolymers [8–10]. Dense hydroxyapatite
mineralized tissues, such as enamel, dentine, and cement bioceramics, formed into suitable shapes, may be used in
[1, 2]. Biological apatite, being the main component of the the creation of implants for the middle ear and eye (orbital
inorganicfraction,isacarbonatehydroxyapatite,depletedin implant),aswellasPDimplants(PercutaneousDevice),and
hydroxylgroupsandrichinsmallamountsofvariousions, are included in inner dialysis systems [11–13]. Hydroxyap-
2+ + + 2+ 2− 4−
includingMg ,K ,Na ,Mn ,HPO4 ,andSiO4 [1–3]. atite is widely used as a coating of metallic implants for
Synthetic hydroxyapatite (HA, HAp) is used in recon- bones in order to improve and accelerate the process of
struction and repair surgery, conservative dentistry, dental osseointegration [14, 15]. Hydroxyapatite powder plays an
implantology,andpharmacy(Figure1)[1,4–6]. important role in dentistry (e.g., in the treatment of dental
A porous hydroxyapatite material may be used as a pulpanddentinehypersensitivity)andisassociatedwiththe
substitute bone material that fills tooth sockets after tooth exposureofdentinaltubules[16,17].Hydroxyapatite,present
extractionorelseasabiomaterialformingascaffoldfornewly in toothpastes and dental gels, reduces the deposition of
formed bone (see Figure2) [7]. Between such material and accretions on teeth. It can also be used as a component of
thebone,theformationofbiologicalfixationtakesplace:the dentalcementsandfillings[18].Itisalsoworthmentioning
structureofthelivingbonetissuepenetratesthefreespaceof that microporous structures of hydroxyapatite can serve as
thematerial,thuscausingthemorepermanentfixationofthe carriers of drugs supplying medicinal substances directly
implantinthebone[4]. to a destination. The studies on hydroxyapatite systems for
2 BioMedResearchInternational
vacancy, which is compensated for by the simultaneous
2+
releaseofonecationofcalcium(Ca )andonehydroxylion
Bone (OH−)[27].
substitutes
Thiscapacityfortheionexchangeofhydroxyapatiteshas
beenusedrecentlyinbiomaterialengineering.Onealsouses
thefactthattheintroductionofevensmallquantitiesofsome
Composite ionsmaycausechanges/improvementsinbiological,physic-
implant Implant
components coatings ochemical, or mechanical properties [28, 29]. For example,
theintroductionofmagnesiumionshasapositivebiological
Hydroxyapatite effectandimprovestheosseointegrationalpropertiesofthe
material[30].Inturn,theintroductionofasmallamountof
2+
Mn ionstotheapatitefavoursosteoblastsproliferationand
increasesthebiocompatibility[31].
It should be emphasized that infections around bone
Drug implants comprise the key problem for modern recon-
Dental
delivery
materials struction surgery [32, 33]. The defence mechanisms of the
systems
bodymustbesupportedbytheintroductionofantibacterial
factors,particularlyantibiotictherapy.Thistypeoftherapyis
performedusingtheoralrouteandislesseffective:thedoseof
Figure1:Schematicviewofhydroxyapatites’applications.
theantibioticmustbehighinordertoensureitsappropriate
concentration around the inserted implant. In turn, local
antibiotictherapyappliedattheplaceofinfection(e.g.,atthe
locationoftheimplantfilledwithanappropriateantibiotic)
also has its limitations: on the one hand, the concentration
the controlled release of anticancer drugs, antibiotics, and
oftheantibacterialdrugmustbehighinordertoensurethe
growthfactorswerereportedinseveralpapers[19–21].
efficiencyofthetreatment;andontheotherhand,itmustnot
Hydroxyapatite is a crystalline calcium phosphate of
exceedasafeleveloftoxicitytocells.
the general formula: Ca10(PO4)6(OH)2. Stoichiometric and
Therefore, implantation surgery seeks additional factors
perfectlypurehydroxyapatitecrystallizesinthemonoclinic
that increase the antibacterial activity around introduced
system [22]. Broadly, HAp crystallizes most often in the
biomaterials.
hexagonalsystem,aspacegroupofP6/3mc.
Important structural features of the stoichiometric At present, a fairly common tendency is to use the
+
hydroxyapatite are its structural hydroxyl groups, arranged antibacterialpropertiesofcertainions,includingsilver(Ag ),
2+ 2+
at the edges of elementary cells, forming the columns copper(Cu ),andzinc(Zn ).Inaddition,onetriestolook
3+
–OH–OH–OH—(see Figure3). Oxygen atoms of these forthepossibilityofusingotherions,suchasceriumCe ,
3+ 2− 4+
groupsarespacedinsuchawaythattheyareunabletoform galliumGa ,seleniumSeO3 ,titaniumTi ,andstrontium
hydrogen bonds [23]. Hydroxyapatite includes two types of Sr2+,amongothers.
calciumcations,referredtoasCa(I)andCa(II).Theatoms The mechanisms of the antibacterial activity of ions are
of calcium Ca (I) are located at the edges of a hexagonal notfullyexplained.However,theliteratureprovidesinforma-
unitcell,whiletheatomsofcalciumCa(II)formequilateral tion on three hypothetical mechanisms [34–37] (Figure4).
triangles with the column of structural hydroxyl groups in Accordingtothefirstmechanism,theionspenetrateintothe
themiddle.Thephosphateionsarethelargestionsthatbuild bacterialcelland,byaffectingtheproductionofintracellular
unitcells,beingtheonetodetermineitsstructure[24]. ATP,theydisrupttheprocessofDNAreplication.Thesecond
One of the primary features of hydroxyapatite is its mechanism is associated with the accumulation of ions
capacityforionsubstitution(i.e.,ionexchange).Thismeans in the cell membranes of bacteria, and thus with changes
thatthelocationsforhydroxylionsmaybeoccupiedbyions in their permeability (the gradual release of proteins and
− −
of a similar size and charge, such as Cl or F . In turn, the lipopolysaccharides).Thetransportationofprotonsthrough
3−
locationsforphosphateionsareoccupiedbyBO3 ions,and the cell membrane is prevented, and consequently it leads
2+ 2+
thelocationsforcalciumcationsbytheionsMg ,Mn ,or to the destruction of the cell membrane and the death of
Sr2+[1,25,26]. the bacterial cell. The third mechanism is based on the ion
In some cases, we are talking about the total exchange inductionofreactiveoxygenspecies(ROSs).Oxygenradicals
(thisispossibleinthecaseoftheexchangeofOH–ForCa– areabletoreactwiththecomponentsofthemembraneand
Sr) or partial exchange (limitations are mostly due to the cell wall of bacteria, as well as other cell components (e.g.,
difference in the sizes of the ions, charges, and their spatial mitochondria),causingirreversiblechangesintheirstructure
structures). andthusthedeathofthebacterialcell.
Animportantaspectisalsothepossibilityofthesubstitu- The purpose of this paper is to review the available
tionofionswithdifferentcharges.Forexample,itispossible literature and summarize the various achievements in the
to exchange phosphate ions (−3) with carbonate ions (−2). field of hydroxyapatites modified by ions with antibacterial
Suchasituationleadstothecreationofapositivelycharged properties.Wehavedescribedtheachievementsconcerning
BioMedResearchInternational 3
(c)
(a)
3 mm
6 mm
10𝜇m
(b) (d)
Figure2:AhemiporousHApscaffold:(a)overview,(b)cross-section,(c)porouspart,and(d)densepart.Reprintedfrom[7]withpermission.
Protein Membrane
DNA damage
Membrane
disruption Oxidized cellular
Ca I component
+
Ag Release of
metal ions
2+
Zn
PO4
Ca II Ca I
Ca II
OH Ca II R eoasxctpyiegvceeines e−
ROS production e− Interrupted transmembrane
Figure 3: Hydroxyapatite structure—a schematic view (adapted electron transport
from[1]).
Figure4:Antibacterialmechanismsofnanoparticlesandtheirions,
adaptedfrom[38]withpermission.
thesynthesisandtestingofthebiologicalandmicrobiological
+
activity of these compounds. We have tried to present 2.Ag -SubstitutedHydroxyapatite
our own point of view as to the applicatory potential of
these materials. We hope that our research will serve as a Forhundredsofyears,silverhasbeen widelyknownforits
valuableguidetohydroxyapatitebioceramicswithadditional strongantibacterialproperties[39]ofanexceptionallybroad
antibacterialactivity. spectrum, including both gram-positive and gram-negative
4 BioMedResearchInternational
Ag+ Ag+ Increasingattentiononthepartofresearchershasbeen
attractedbyhydroxyapatitedopedwithsilverions(Ag-HAp).
IthasbeenshownthatAg-HApexhibitsstrongantibacterial
+
Ag activity[52–57],andonehasproposedtwomechanismsfor
itsactivity.Thefirstofthemsuggeststhatmicroorganismsare
S-H S-Ag attractedbyelectrostaticforcetothesurfaceofthehydrox-
yapatite, where there is a direct interaction between the
bacterialcellmembraneandsilverions.However,according
S-H S-Ag
to the second mechanism, silver ions are slowly released
from the interior of the hydroxyapatite, which discloses its
bactericidalactivitythroughoutthematerialsurroundingit
[55].
There are various methods for the preparation of Ag-
Figure5:ThemainproposedmechanismofAg+ions’antibacterial HAp:
activity(adaptedfrom[49]).
(i)wetmethodsynthesis,usingthestartingmaterialsin
the form of salt [55, 57–61] or acids as well as bases
andoxides[62];
bacteria, viruses, and fungi [40, 41]. Of all the antibacterial (ii)thesol-geltechnique[63,64];
metals,silverhasthehighestefficiencyatconcentrationsas
(iii)ionexchangebetweenthepurehydroxyapatiteanda
lowas35ppb,atwhichithasnotoxiceffectonmammalian
silversaltsolution[59,65–67];
cells [42, 43]. In addition, microorganisms have a relatively
poor ability to develop immunity to silver. Therefore, it is (iv)ultrasonicspraypyrolysis(USSP)[68];
widely used in many forms in order to limit the growth of (v)themicrowavemethod[69].
microorganisms, both in medicine (e.g., for the treatment
of flash burns, catheters, the coverage of implant materials, The resulting materials may be then subjected to treat-
urinarytractinfections,dentistry[36,44]andinotherfields ment, such as heat-treating by sintering in an oven or
[45,46]). dryingwiththeuseofthesublimationmethod.Depending
Themechanismoftheactionofsilveronmicrobialcells uponthemethodofsynthesisorthepossibletreatment,the
depends upon its form and is complex in each case since it resulting apatites are characterized mainly by their degree
remainsinsufficientlyunderstood in detail[47]. In thecase of crystallinity and their specific surface area. It affects the
ofsilverions,itisbasedprimarilyontheirinteractionwith efficiency of the antibacterial activity because the rate and
groups–SH(thiolic)ofproteins[48](seeFigure5),including extent of the release of silver ions from the apatite increase
theexchangeofahydrogenatomswithsilveratoms,leading alongwithitssurfacearea,andthustechniquesthathelpin
totheformationofS–Agbonding[49]. obtainingfinecrystalswillleadtotheobtainingofamaterial
Thisresultsintheinactivationofbothproteinslocatedin exhibiting stronger antibacterial activity. For example, it
thecellmembraneandthosewithinthecytoplasm.Following was proved that drying the silver-hydroxyapatite using the
the denaturing action of silver ions, a number of changes sublimation method leads to the preparation of a material
occur in the bacteria cell, including dysfunction of the with finer crystals, thereby also enhancing the activity of
respiratorychainandmembranepumps;thecellmembrane antibacterialAg-HAp[57].Inaddition,methodsofsynthesis
+
shrinksandseparatesfromthecellwall,andasaresultthe (suchasUSSP,thedirectincorporationofAg ionsintothe
cell contents leak out and the cell wall is torn apart [47, structure of HAp, as well as increases in the crystallinity of
50]. Silver ions also interact with DNA molecules, causing Ag-HAp by sintering) lead to the preparation of a material
+
their condensation and loss of capacity to replicate [47]. It with a slower release of Ag ions, and thus to prolonged
was also shown that silver ions intensify the production of antibacterialactivity[68].
ROSs [51], which undoubtedly enhances their antibacterial DuringAg-HApsynthesis,differentamountsofsilverare
properties [49]. In turn, silver in its nanoparticle form acts introducedwithintherangeof0.1%to10%ofweight,usually
primarily on the basis of the induction of the formation of inseveralincreasingconcentrations,allowingaseriesoftests
ROSs that interact with the cell wall and cell membrane of for the various contents of silver. The aim of such a study
bacteria, causing their depressurization and an increase in is to determine the optimum concentrations, characterized
permeability. As a result, the cell content flows out and the by a high efficiency of action against a broad range of
celldies[52–54]. microorganisms.
+
Authors studying the mechanism of the action of Ag The broad antibacterial spectrum of Ag-HAp is similar
ions suggest that the thickness of the peptidoglycan layer andhasthesamewidthasthespectrumofsilverions,includ-
of gram-positive bacteria may—at least to a lesser extent— ingbacteria,viruses,andfungi[70].Moreover,likethesilver
+
protect the bacterial cell from the influx of Ag ions to the ions,ithasamoreeffectiveinfluenceongram-negativerather
+
inside,whichwouldexplainthehigheractivityofAg against than gram-positive bacteria, due to the differences in the
G− (e.g., Escherichia coli) as opposed to G+ bacteria (e.g., structuresofcellwalls[70].Evensuchalowcontentofsilver
Staphylococcusaureus)[47,50]. as0.2%ofweightinAg-HApeffectivelyinhibitsthegrowth
BioMedResearchInternational 5
1.4 Ag: HAp (𝜒 =0.01) notonlythetotalcontentofsilverinthehydroxyapatitebut
1.2 Ag also the conditions of its synthesis and the method for its
1 dryingandheattreating.Nevertheless,onecanobserveasig-
m 0.8 nificantandstillgrowinginterestinthispromisingmaterial.
n
20 0.6
6 2+
0.4 3.Cu -SubstitutedHydroxyapatite
0.2
0 Copper is an essential micronutrient of almost all living
5 5 5 5 3 6 8 9 2 1 + organismsbecauseitisinvolvedinmanymetabolicprocesses.
2. 1.2 0.62 0.31 0.15 0.07 0.03 0.0 0.0 c However, higher concentrations of copper ions may have
Concentration (mg/mL) toxic effects, inter alia, because of their ability to generate
ROSs.Theantibacterialpropertiesofcopperwerediscovered
E. coli ATCC 25922 K. pneumoniae 2968
E. coli 714 B. Subtilis very early on; we know of Egyptian texts written around
C. krusei 963 2600–2200BCwhichpresentmethodsforthesterilizationof
drinkingwaterandwoundstothechestusingcopper[76].
Figure 6: Antibacterial activity of Ag-HAp (𝜒 = 0.1) on E.
Ag Themechanismoftheantibacterialactivityofhydroxya-
coliATCC25922,E.coli714,Klebsiellapneumoniae2968,Bacillus
patitesdopedwithcopperionsisnotyetfullyunderstood.It
subtilis,andCandidakruseistrains(adaptedfrom[69]withpermis-
isbelievedthatcopperionsformstrongbondswiththiolic,
sion).
imidazole,amine,andcarboxylicgroupsofproteins,causing
structuralchangesandincreasesinpermeabilityand,hence,
of K. pneumoniae and C. krusei. In contrast, antibacterial membrane transport dysfunction and cell death [77]. In
activityagainstE.coliandB.subtilisrequiresacontentof0.5% addition, copper ions also form a bonding with amine and
weightofsilverintheapatite[69](seeFigure6).Thegrowth amide groups, as well as with the disulphide bridges of
ofS.aureusandStaphylococcusepidermidisisclearlyinhibited proteins and enzymes of bacteria, causing damage to DNA
withasilvercontentof1%ofweight[71]. andRNAandresultingintheinhibitionofthereproduction
Excessive silver content in the hydroxyapatite may be ofbacteriaortheirdeath[78].
toxic, not only to microorganisms but also to mammalian The antibacterial properties of copper are also used
tissues,andmayleadtotheinhibitionoftheirgrowth[61,71, by macrophages which, by copper concentration, increase
72].Therefore,theselectionofanappropriatesilvercontent their ability to inactivate microorganisms through the oxy-
in Ag-HAp is an important issue, since it should be high gen burst [79]. Therefore, some microorganisms, such as
enoughtobeabletoeffectivelyfightmicroorganisms,while Mycobacteriumtuberculosis,S.aureus,andSalmonellaenter-
at the same time it should also be limited in order that it ica, have produced mechanisms protecting them from the
doesnotadverselyaffecttheconditionofmammaliantissues. toxic effect of copper in order to be able to survive the
Numeroustoxicologicalstudieshaveshownthat,alongwith intervention of macrophages [80–83]. In addition, copper
the increase of the content and the rate of release of silver deficiencyreducesthebactericidalactivityofneutrophilsand
ions from Ag-HAp, we can also observe an increase in invitromacrophages[84].
theirnegativeimpactontissues,inhibitingtheirgrowthand Studies have confirmed the antibacterial efficiency of
+
development,andinhigherAg concentrations—leadingto hydroxyapatitesdopedwithcopperionsagainstE.colibacte-
deathofthecells.Inordertodeterminetheoptimumsilver ria.Hydroxyapatitesweresynthesizedusingthewetmethod.
content, one performed in vitro toxicity studies on human Copperionswereintroducedintothestructureusingcopper
osteoblasts [71, 73, 74], human stem cells [61], and mouse acetateofCu(CH3COO)2⋅H2O.Hydroxyapatitesdopedwith
fibroblasts [75], as well as in vivo studies in rats involving copper ions to the amount 3.3% of weight inhibited the
theimplantationofAg-HAp.Invitrostudiescarriedouton growthofE.colibacteria,whileacontentof0.66%ofweight
2+
humanstemcellsshowthatahydroxyapatitecontaining0.3% ofCu wasdeemedtobeinsufficienttoinhibitbacteria[85].
silver weight has no negative effect on the growth of these Studieshavealsoshowntheinhibitoryeffectofhydrox-
cellsoverthecourseofsevendaysofculture,whileacontent yapatitesdopedwithcopperionsonthegrowthofCandida
of 0.7% silver slightly inhibits their growth. In contrast, a albicansfungi[77](seeFigure8).
concentrationof8.3%weightexhibitsasignificantcytotoxic Inthiscase,hydroxyapatiteswerealsoobtainedusingthe
effect, specifically in inhibiting the growth of human stem wet method, while the copper ion source was copper oxide
cells [61]. In vitro experiments on human osteoblasts have (II)CuO.Theresultingmaterialmorestronglyinhibitedthe
shownthatasilvercontentof6%weightsignificantlyinhibits growthoffungithaninthecaseofstudiesofhydroxyapatites
the growth of osteoblasts and leads to the death of some of dopedwithzincions.Copperionsdonotinhibitthemulti-
them,while2%and4%weightsrepresentcontentsthatenable plicationofS.aureusbacteria,whichisprobablyduetothe
agoodbalancebetweeneffectiveantimicrobialandcytotoxic factthatS.aureusisagram-positivebacteria.
activity [73]. In addition, in vitro studies conducted on rats Cooperions,incontrasttootherdivalentmetalcations
confirmedtherelativelylowtoxicityofAg-HAp,containing (Zn, Co, Ni, Mn, Fe, and Sn), cause selective changes in
upto4.3%weight(seeFigure7)[72]. thepermeabilityofthecellmembraneoftheSaccharomyces
The complexity of the problem of the balance between cerevisiaeyeasts,withnochangesinthevacuolemembrane
effectiveantimicrobialactivityandcytotoxicactivityincludes permeability[86].
6 BioMedResearchInternational
20𝜇m 20𝜇m
(a) (b)
20𝜇m 20𝜇m
(c) (d)
Figure7:FESEMmicrographsillustratingthehFOBcellmorphologyafterthreedaysofculture:(a)HA,(b)HA-2Ag,(c)HA-4Ag,and(d)
HA-6Ag.Thehairlinearrowindicatesalivecellandthethickarrowindicatesadeadcell(reprintedfrom[72]withpermission).
Inthecaseofhydroxyapatitesdopedwithcopperions,a ofbacteriaandfungi,includingE.coli,S.aureus,C.albicans,
problemmaybecausedbythecytotoxicpropertiesofcopper, and Streptococcus mutans [92]. Studies on a hydroxyapatite
bothagainstbacteriaandosteoblasts[77].Suchaneffectcan doped with zinc, synthesized using the wet method and a
beobservedinthecaseofhydroxyapatiteswithacontentof hexahydrate zinc nitrate of Zn (NO3)2⋅6H2O (wherein the
0.66%ofweightofCu2+. zinc ion content was 1.6%), have shown its real impact on
the reduction of the numbers of S. aureus bacteria [93]. In
2+ addition, it was proved that the small amount of zinc ions
4.Zn -SubstitutedHydroxyapatite
introducedintothehydroxyapatitestructureplaysanimpor-
Zinc, one of the essential microelements, is present in the tant role in the processes of the growth and differentiation
active centres of more than 300 enzymes involved in the ofthecells;however,themechanismoftheions’impacton
metabolismofbones.Italsoexhibitsantibacterialactivity,the biologicalprocesseshasnotyetbeendiscovered.
mechanism of which is similar to that of copper ions. Due Theuseofhydroxyapatitesdopedwithzincionsasacoat-
to its antibacterial properties, one started to introduce zinc ingfortheremineralizationofteethhelpstoreducebacterial
ionsintothemoleculeofhydroxyapatite.Moreover,zincions adherence and the growth of tartar. Studies confirmed the
havealsohadadirectimpactontheproliferativeproperties efficiencyofhydroxyapatitesdopedwithzincionsinreducing
ofosteoblastsandaninhibitoryeffectonboneresorptionby thegrowthofthethreemostcommonoralpathogens,includ-
osteoclasts. ing Aggregatibacter actinomycetemcomitans, Fusobacterium
Some papers have presented studies on hydroxyapatites nucleatum,andS.mutans[94](seeTable1).
dopedwithsmallamountsofzincions(lessthan1%),which Hydroxyapatites are also used in the treatment of bone
confirmed effective bioactivity and antibacterial properties cancerandchronicosteomyelitis.Intheseandotherdiseases
[77, 85–90]. In contrast, 1.3% content of zinc ions causes ofthebones,infectionsposeseriousproblems.Unfortunately,
anincreaseinosteoblastresponses[91].Inadditiontotheir themostcommonlyusedantibioticshaveverylowconcen-
biologicalproperties,onehasalsoconfirmedtheinhibitory trationsinthebonetissue.Therefore,itiscrucialtolookfor
effectofhydroxyapatitesdopedwithzinconthedevelopment asolutionwhichachieveshighconcentrationsofantibiotics
BioMedResearchInternational 7
(a) (b)
CuHAP1
CuHAP2
(c)
Figure8:PhotographsoftheantimicrobialtestresultsofCuHAPsamplesagainstE.coli:(a)CuHAP1,(b)CuHAP2,andC.albicans:(c)
CuHAP1andCuHAP2(reprintedfrom[77]withpermission).
Table1:Antibacterialactivitiesofzinc-loadedcalciumphosphatenanostructures.ThesamplesA,B,andCwerepreparedbydifferentroutes:
∘
A—theobtainedprecipitatewasdriedandheatedat200Cfor3hbeforeZn-loading;B—thecollectedwetproductwasdirectlyusedfor
∘
Zn-loading;andC—preparedat70Cfor10h(rewrittenfrom[94]withpermission).
Sample Bacterium Inhibitionrate% Bacterium Inhibitionrate% Bacterium Inhibitionrate%
A 99.82 8.91 17.25
Aggregatibacter Fusobacterium
B 95.98 99.97 Streptococcusmutans 86.87
actinomycetemcomitans nucleatum
C >99.99 >99.99 >99.99
in the bone tissue while reducing the toxic effect of drugs zinc ions is greater than the drug released from the “pure”
on the human body. Studies have been performed on the hydroxyapatite. The use of such a solution in the treatment
efficiencyofhydroxyapatitesdopedwithzincionsascarriers of infections of bone tissue allows for the shortening of
for the controlled release of ciprofloxacin, which exhibits therapy—increasingitsefficiency—andthereductionofthe
antibacterial activity against S. aureus and Pseudomonas doseofciprofloxacin,whichcontributestothereductionof
aeruginosa(thesebeingthemostcommonpathogenscausing the formationof resistant strains. As in the case of hydrox-
diseasesofthebonesandjoints).Theyhaveshownthatthe yapatitesdopedwithcopperions,theuseofhydroxyapatites
release of ciprofloxacin by the hydroxyapatite takes place containing zinc ions may be limited by the cytotoxicity of
in connection with the release of zinc ions and increases zinc,whichisrevealedwhenthecontentexceeds1.2%[93].
2+
in the drug concentration and number of Zn ions used Duetothepossiblyidenticalmechanismoftheantibac-
increases the antibacterial activity [95]. Furthermore, the terial activity of zinc and copper ions and their similar
release of ciprofloxacin from hydroxyapatite doped with efficiency as regards the treatment of microorganisms, the
8 BioMedResearchInternational
studiesonthesimultaneousintroductionoftheionsofthese caused by the presence of strontium and not fluoride, as
bothmetalsintothehydroxyapatitestructureareconducted. was previously thought. In turn, in the paper of [111], it
wasprovedthattheantibacterialpropertiesofstrontiumare
2− verypoorbutthatitscombinationwithfluorideimprovesits
5.SeO -SubstitutedHydroxyapatite
3 antibacterialpotentialinthetreatmentoftoothdecay.
Onealsostudiedtheantibacterialactivityofhydroxyap-
Selenium is one of the microelements necessary for the
atiteincorporatedwithstrontiumions.Intheelaborationof
proper development of the human body. Yet, until recently,
[112],whereSr-hydroxyapatitewassynthesizedusingthesol-
it was considered a highly toxic agent [96, 97]. However,
gel-supercriticalfluiddryingmethod(SCFD),itwasproved
studieshaveshownthatitisacomponentofselenoproteins
thatthematerialcontainingstrontiumionsexhibitedantibac-
and the glutathione peroxidase enzyme, responsible for the
terial activity against E. coli, S. aureus, and Lactobacillus.
protection of cell membranes against harmful factors [97].
In turn, in the elaboration of [113], Sr-HAp was prepared
Many studies have proved that selenium is a factor which
usingtheacceleratingactionofmicrowaves.Asmallamount
protects against oxidative stress and carcinogenesis. There-
of strontium in the received materials caused a bactericidal
fore, one performs research on the use of selenium com-
effectonE.coliandS.aureus.
pounds in the production of multifunctional biomaterials
which, in addition to the scaffold function, also represent
3+ 3+
the anticarcinogenic functions [98]. Studies conducted by 7.Ce -andEu -DopedHydroxyapatite
Tranetal.[99]andRodr´ıguez-Valenciaetal.[100]havealso
shownfavourableantibacterialproperties.Seleniumionsof Cerium and europium are rare earth elements from lan-
2−
SeO3 wereintroducedintothecarbonatedhydroxyapatite thanides,whicharepresentinthehumanbodyonlyasaresult
structure using the pulsed laser deposition method (PLD). of the cumulative effect of the environment. They have the
Thematerialformedthiswaywasusedtocreateacoatingfor abilitytoaccumulateinsmallamountsinthebonesandliver.
titanium.ItsantibacterialactivityonthestrainsofS.aureus Wedonotknowthebiologicalroleofceriumoreuropium,
and P. aeruginosa was found [100]. Even at a concentration butrecentstudieshaveshownthatceriumsaltsmaystimulate
of 0.6% weight of selenium in the coating, there was an metabolism[114].
inhibitionofbiofilmformationbythetwobacterialstrains. Inrecentyears,researchershavebeguntoinvestigatethe
It is worth noting that the concentration of selenium was biological properties of the cerium oxide of CeO2. It has
nontoxic and had no adverse effect on the adhesion of beenproventhatceriumoxideisabletoinduceangiogenesis
the cells or their proliferation. The literature discusses the through its direct effect on the modulation of oxygen in
antibacterial mechanism of selenium [100–103]. It is most intracellularenvironments[115].Inturn,Lordetal.[116]have
likely associated with oxidative stress, resulting in damage studied the impact of cerium oxide on human monocytes,
to the cell walls of bacteria. Microorganisms metabolize based on its ability to scavenge ROSs. The antibacterial
0
selenium to a harmless form of elemental selenium of Se properties of cerium oxide have also been studied in a
−
orHSe (hydrogenselenide),whichinturncanbeincluded number of elaborations, especially of strains of E. coli, B.
inselenocysteine.Asaresultofmetabolicreactions—which subtilis, Salmonella typhimurium, and Enterococcus faecalis
−
are oxidation-reduction reactions—H2O2 or even O2 may [117,118].
becreated,whichwouldconfirmthetheoryoftheoxidative
Therearefewworksdevotedtotheantibacterialproper-
antibacterialmechanismofselenium.
tiesofceriumionsandtheiruseinbiomaterialsengineering.
However, the literature contains reports on the synergistic
2+ effectoftheactivityofzincandceriumionsinnanomaterials
6.Sr -SubstitutedHydroxyapatite
ofalpha-zirconiumphosphateandtitanium[119,120].
Strontium is a microelement that cumulates in the bone In the available literature, only two elaborations are
mineral, playing an important role in it. A small amount devoted to hydroxyapatite doped with cerium ions [121,
of strontium induces the formation of bone tissue and the 122]. Feng et al. synthesized hydroxyapatite containing a
inhibition of its resorption. The available literature also maximumof10%weightofceriumusingthehydrothermal
containsinformationaboutthereliefofbonepainsandeven method[121].Theintroductionofceriumionstothehydrox-
attempts to treat bone cancer [104, 105]. There are many yapatite structure was manifested by the reduction in the
elaborations devoted to the introduction of strontium ions crystal size to about 20nm. Unfortunately, this elaboration
into the hydroxyapatite structure [106–109]. Due to their didnotconsiderresearchintotheantibacterialpropertiesof
similar properties and small differences in size, calcium theresultingmaterial.Linetal.[122]synthesizedhydroxya-
cationsmaybecompletelyreplaced withstrontiumcations. patitecontainingceriumionsusingthesol-gel-supercritical
The introduction of strontium ions into the hydroxyapatite fluiddryingmethod(SCFD).Theintroductionofceriumions
improves its biocompatibility and bioactivity, streamlining intothestructureresultedinthereductionofthecrystallinity
the process of osseointegration. In addition, strontium ions of the materials and a change in crystal morphology: from
improvethemechanicalpropertiesofthehydroxyapatite. rod-shapedHAptoneedle-shapedCe-HAp.Theelaboration
Recent reports also show that strontium can be used was devoted to the antibacterial activity of the resulting
as an antibacterial agent. Guida et al. [110] suggested that materialsbasedonthefollowingbacterialstrains:E.coli,S.
theantibacterialactivityofglass-ionomercementsismainly aureus,andLactobacillus.Itturnedoutthatthematerialswere
BioMedResearchInternational 9
2 Eu: HAp (xEu=0.2) reactions.Withthisproperty,galliumisusedasadiagnostic
1.8 andtherapeuticagentinmetabolicdisordersofsoftandhard
1.6 tissues[125].
1.4 Recentstudieshavealsoprovedtheantibacterialactivity
m 1.2 of gallium ions, which is based on the exchange of iron
n 1 ions in protein metabolism [126, 127]. One showed the
0
62 0.8 antibacterial activity of phosphorus, silica, and phosphorus
0.6 glasscontaininggallium.
0.4
Theavailableliteraturecontainsinformationonhydrox-
0.2
yapatite synthesis, enriched with gallium ions III [128].
0
The content in the material amounted to 11% of weight.
1 5 5 5 2 1 +
0. 0.2 0.12 0.06 0.03 M However,onehasnotexaminedtheantibacterialactivityof
Concentration (mg/mL) thismaterial.Inouropinion,suchstudiesshouldbecarried
out.
C. albicans E. faecalis
S. aureus E. coli
P. aeruginosa 9.Ti4+-ContainingHydroxyapatite
Figure 9: Antimicrobial activity of as-prepared Eu:HAp samples
(𝜒 =0.2)onE.coliATCC25922,P.aeruginosa1397,S.aureus0364, Inrecentyears,alotofattentionhasbeengiventotitanium
E.EfuaecalisATCC29212,andC.albicansATCC10231(adaptedfrom oxide TiO2, due to its photocatalytic properties [129, 130].
One uses its photooxidative activity on organic materials,
[123]withpermission).
suchasproteinsandlipids.Themechanismofphotocatalysis
of titanium oxide is described in the elaboration of [131]. It
takesplaceaccordingtothefollowingreactions:
characterizedbyahighantibacterialcapacity—thegreaterthe
dtheegrheeighoferthtehesuabnsttiibtuactitoenriaolfccaaplcaicuitmy. iAotnsablyowcerciounmteniotnosf, TiO2+h]󳨀→e−CB+h+VB (1)
ceriuminasample,theantibacterialactivitywasbasedonly − +
Free electrons (e ) and positively charged holes (h )
on the dynamic contact of the material with bacteria at a CB VB
have strong oxidizing and reducing properties. When elec-
concentrationof0.1g/mL.
Whenitcomestotheantibacterialactivityofeuropium tronsandvacanciesmigratetothesurface,TiO2canpartici-
pateinthereactionsofoxidation/reductionwithmolecules
ionsinthecontextofbiomaterials,ithasbeendescribedin
absorbed on the surface (i.e., water, oxygen, and organic
[123,124].Onestudiedtheantibacterialactivityofmaterials
andinorganicmolecules)(Figure10).Electronsmayreduce
synthesized using the coprecipitation method (i.e., the wet
Ti(IV)toTi(III),whichinturnreactswiththeoxygentoform
method) on the following strains of bacteria and fungi: S. −∙ −
aureus,E.coli,P.aeruginosa,E.faecalis,andC.albicans[123] O2 radicals.VacanciesmayreactwithwaterorionsofOH
andformradicalsof∙OH:
(seeFigure9).Thesestudieshaveshownthatevenwithalow
contentofEu3+,thematerialshadanantibacterialeffecton H2O+h+VB 󳨀→∙OH+H+
strainsofP.aeruginosa,S.aureus,andE.faecalis.Incontrast, (2)
these materials showed no bactericidal capacity in relation OH−+h+ 󳨀→∙OH
VB
totheE.colistrain.Fungicidalactivitywasexhibitedbythe
materialswithahighercontentofeuropium. In turn, the radicals of O2−∙ and ∙OH may react together,
causingtheformationofH2O2.
8.Ga3+-SubstitutedHydroxyapatite These factors, the so-called “reactive oxygen species”
(ROSs), may react with the wall or membrane of the cell
Gallium is an element from the group of semimetals. Its of bacteria. Unsaturated phospholipids—contained in these
efficacyinthetreatmentofmanytypesofdisordershasbeen cellelements—participateinreactionswithROS,causingthe
developedinthelate20thcentury.Awiderangeofpotential deathofthebacterialcell.
applications have been described in an extensive review of TheantibacterialpropertiesofTi(IV)areusedinbioma-
Bernstein [125]. Gallium inhibits bone resorption and thus terialsengineering.Inadditiontotheuseoftitaniumoxide
lowers the calcium concentration in plasma. In addition, as a component of biomaterial components, the literature
4+
it induces osteoblasts and helps bone formation. It has a describes hydroxyapatites doped with Ti ions [133, 134].
4+
positiveeffectontheadoptionofimplants.Numerousstudies Wakamura[133]provedthatTi inhydroxyapatite—where
havealsoproveditsimmunomodulatoryactivity.Inaddition, therearesmallamountsofTi(molarratioofCa/Pto100)—
2+ 2+
gallium—duetoitsantiproliferativeandantimitoticeffect— formsdivalentcations[Ti(OH)2] and[TiHPO4] ,which
may positively influence the treatment of certain types of replace calcium cations. Titanium contained in hydroxyap-
cancer[125]. atite crystals has a similar antibacterial mechanism to tita-
3+
DuetothehighsimilarityofFe ions(i.e.,similarionic niumoxide.Hydroxyapatitehasahighaffinityforproteins;
radius,electronegativity,coordinationnumber,etc.),gallium therefore, hydroxyapatite modified with Ti4+ ions has an
3+
ions of Ga may replace them in a variety of metabolic antibacterialeffectnotonlyafterUVirradiation.
10 BioMedResearchInternational
𝜆>390nm O2 medicineanddentalimplantology.However,wecanobserve
a trend of a broadening of the spectrum of the apatite
−∙
UV light O2 materials, with additional biological, physicochemical, or
biomechanicalproperties,despitethescaffoldforthenewly
Ele ctrone− ROS formedbone.Itseemsveryreasonabletolookforthepossi-
bilityofthesubstitutionofionswithantibacterialproperties.
+organic
So far, the best-known and most commonly used material
compounds
is hydroxyapatite enriched with silver ions. Much is also
Hole h+ ROS CO2+H2O ksunboswtintutaebdoubtytthheeainontisbaocftecroiaplpearctiavnitdyzoinfch.yOdrnoxthyaepoattihteers
TiO2
hand, it seems that the other materials still require a great
∙OH deal of research. In our opinion, one should also examine
H2O the relationship between the method of the preparation of
thesubstitutedapatitematerialwithitsantibacterialactivity
Figure 10: Photocatalytic properties of TiO2 and its antibacterial
and its efficiency. Such studies remain very rare or else
potential(adaptedfrom[132].
incomplete.
2+
10.Co -SubstitutedHydroxyapatite
ConflictofInterests
Cobaltisanessentialelementfortheproperfunctioningof
The authors declare that there is no conflict of interests
the human body. First of all, it is a component of vitamin
regardingthepublicationofthispaper.
B12,whichisnecessaryfortheregulationoftheproduction
ofredbloodcells,DNAsynthesisincells,andtheformation
Acknowledgments
of the myelin sheath, protecting the cells of nerves and
neurotransmitters[135].
JoannaKolmasthankstheFoundationforPolishSciencefor
The literature presents information on the use of Co
supportingherwiththeParent-BridgeProgrammethatwas
(II) ions as an antibacterial and antiviral agent in various
cofinancedbytheEUEuropeanRegionalDevelopmentFund.
organiccomplexes[136,137].In2013,twoelaborationswere
Ewa Groszyk and Dagmara Kwiatkowska-Ro´z˙ycka thank
developed concerning hydroxyapatites partly doped with
the Medical University of Warsaw for the financial support
cobalt ions (II) [138, 139]. In their work, Ignjatovic´ et al.
(FW23/NM1/13).
[138] presented the hydrothermal method for obtaining
2+
hydroxyapatite containing Co at the amount of 5–15% of
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