Oral Health of Patients Treated with Acrylic Partial Dentures Using a Toothpaste Containing Bee ...

13 Pages·2017·1.33 MB·English

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Evidence-Based Complementary and Alternative Medicine
Volume 2017, Article ID 4034179, 12 pages
https://doi.org/10.1155/2017/4034179
Research Article
Oral Health of Patients Treated with Acrylic Partial Dentures
Using a Toothpaste Containing Bee Product
KarolinaWiatrak,1TadeuszMorawiec,1RafaBRój,2AnnaMertas,3
AgnieszkaMachorowska-Pienidhek,4PatrykKownacki,1MartaTanasiewicz,5
MaBgorzataSkucha-Nowak,5StefanBaron,6TomaszPiekarz,1MaciejWrzoB,1
MateuszBogacz,1 JacekKasperski,2andIwonaNiedzielska7
1DepartmentofOralSurgery,SchoolofMedicinewiththeDivisionofDentistryinZabrze,MedicalUniversityofSilesiainKatowice,
PlacAkademicki17,41-902Bytom,Poland
2DepartmentofProstheticDentistry,SchoolofMedicinewiththeDivisionofDentistryinZabrze,
MedicalUniversityofSilesiainKatowice,PlacAkademicki17,41-902Bytom,Poland
3DepartmentofMicrobiologyandImmunology,SchoolofMedicinewiththeDivisionofDentistryinZabrze,
MedicalUniversityofSilesiainKatowice,Jordana19,41-808Zabrze,Poland
4DepartmentofOrthodontics,ChairofMasticatoryDysfunctionandOrthodontics,SchoolofMedicinewiththeDivisionof
DentistryinZabrze,MedicalUniversityofSilesiainKatowice,PlacTraugutta2,41-800Zabrze,Poland
5DepartmentofConservativeDentistrywithEndodontics,SchoolofMedicinewiththeDivisionofDentistryinZabrze,
MedicalUniversityofSilesiainKatowice,PlacAkademicki17,41-902Bytom,Poland
6ChairofMasticatoryDysfunctionandOrthodontics,SchoolofMedicinewiththeDivisionofDentistryinZabrze,
MedicalUniversityofSilesiainKatowice,PlacTraugutta2,41-800Zabrze,Poland
7DepartmentandHospitalofCraniomaxillofacialSurgeryandOralSurgery,SchoolofMedicinewiththeDivisionof
DentistryinZabrze,MedicalUniversityofSilesiainKatowice,PlacAkademicki17,41-902Bytom,Poland
CorrespondenceshouldbeaddressedtoRafałRo´j;[email protected]
Received21October2016;Accepted23November2016;Published6February2017
AcademicEditor:VassyaBankova
Copyright©2017KarolinaWiatraketal. This is an open access article distributed under the Creative Commons Attribution
License,whichpermitsunrestricteduse,distribution,andreproductioninanymedium,providedtheoriginalworkisproperly
cited.
Thisstudywascarriedouttoinvestigatetheinfluenceofapropolisandteetreeoil-containinghygienicagentonselectedoralhealth
parameters,oralmicroflora,andtheconditionofperiodontalhealth.Thirty-sevenpatientswhounderwentoralrehabilitationwitha
removableacrylicdenturewereselectedandrandomlyassignedintotwogroups:studygroup(A)whichreceivedanewlyformulated
propolisandteetreeoil-containingtoothpasteoracontrolgroup(C)withoutanactiveingredient.API,S-OHI,andmSBIwere
assessedinthreesubsequentstages.Duringeachexaminationswabswereemployedformicrobiologicalinoculation:inthestudy
groupafter4weeksuseoftheactivetoothpasteshowedadecreaseinthenumberofisolatedmicroorganisms.Inthecontrolgroup,
after4weeksuseofthetoothpastewithoutactiveingredientsresultedinincreaseinthenumberoftheisolatedmicroorganisms.
Improvementsinhygieneandtheconditionofperiodontiumwereobservedinpatientsusingactivetoothpastes.Inthestudygroup
theoralfloradiversitywasreducedbythedecreaseinthenumberofculturedmicroorganismspecies,whileinthecontrolgroup
anincreaseinthenumberofculturedmicroorganismsandtheirspecieswasobserved.
1.Introduction numerousreportsonoralbacteriabeingthecauseofbacterial
endocarditis,pneumonia,gastricinfections,chronicobstruc-
Oral health and condition reflect body’s general health. tivepneumonia,andotherdiseases[1].Theriskofcontracting
Condition of the oral cavity, level of hygiene, and residing theabove-mentioneddiseasesincreaseswithage.Moreover,
pathogens affect the whole human organism. There are the amount of patients who use dentures, which are yet
2 Evidence-BasedComplementaryandAlternativeMedicine
anotherplaceofaccumulationofmicroorganismsassociated occurringcompoundswithabroad-spectrumantimicrobial,
withtheseinfections,riseswithageaswell.Absencesofteeth antiviral,antifungal,andantiprotozoalactivity.Additionally,
are closely related to both oral and general health [2]. Loss theyexhibitantiphlogisticandantioxidantproperties[19,20].
of a single tooth may cause rotation, shifting, and tipping Propolis,so-calledbeeglue,ismadebybeesmostlyfrom
of neighbouring teeth towards the empty space, which will resinous substances gathered from buds of some trees. Its
in turn lead to adverse loads on these teeth, not along the propertiesdependonthespeciesofbeeswhichproduceitas
long axis of the tooth [3–5]. Partial lack of teeth, especially well as on the plants used for the production [21]. Propolis
in lateral teeth, may cause the loss of mandibular occlusal is a very complex substance with high contents of active
support,whichleadstochangesinitspositioninrelationto ingredients like flavonoids, phenols, aromatic acids, esters,
themaxilla[6].Suchashiftinmandibularpositionmayresult wax substances, aldehydes, coumarin, sterols, enzymes, or
inmusclefatigueandotherperpetuatedocclusalrelationsin fattyacids,aswellasproteins,microelements,andvitamins
temporomandibularjoints[7,8]. [22,23].Propolisiswell-solubleinethylalcohol;thusmedical
Prosthetic treatment is not only designed to restore extractscaneasilybeprepared[23].Indentistrypropolisis
partial lacks of teeth. It also serves a prophylactic purpose usedinthetreatmentofcaries[24]andperiodontaldiseases
by preventing the formation of pathological changes in [20, 25]. It also exhibits analgesic activity and accelerates
the stomatognathic system and removing already existing woundhealing,whichisusedindentalsurgery[26].Thereare
changes[9].Itiscommontoemployacrylicpartialdentures. numerousscientificreportsonitsanticancer,antiphlogistic,
Suchdenturesshifttheloadsthroughmucousmembraneand and immunomodulating properties [19, 27]. Tea tree oil is
periosteumtothebone,restoringmasticatoryfunctiontoa obtainedfromleavesofatreegrowinginAustraliaandNew
limited extent, do not protect the residual dentition, cause Zealand, Melaleuca alternifolia, using cold water extraction
periodontitisandmucositis,createfavourableconditionsfor [28].
theaccumulationofbacterialplaque,andacceleratetheloss Tea tree oil is a mixture of terpene compounds, such
of alveolar process. Due to the adverse impact of partial as terpinen-4-ol, 𝛾-terpinene, p-cymene, 𝛼-terpinene, and
dentures on the denture medium and residual dentition it 1,8-cineole, which are largely responsible for tea tree oil’s
isnecessarytomaintainproperoralhygiene,removeplaque properties [29, 30]. TTO exhibits strong antiseptic activ-
carefully, and take preventive measures against caries and ity, selective for pathogenic microorganisms, and is highly
periodontopathy[10,11].Regardlessofthetypeofprosthetic effective against oral cavity pathogens [31]. It has also been
restoration chosen, the maintenance of proper denture and demonstrated that TTO has inhibitory properties against
oralhygieneisindispensabletoachieveasuccessfultreatment Candidaalbicans[32].
outcome[12]. The study aims to assess the influence of toothpastes
The oral cavity constitutes a natural environment for containing ethanolic extract of propolis and tea tree oil on
microflora,whosecompositionisanindividualfeature.Itis oralhygieneinpatientswithpartialdentures.Thenumberof
influenced by, inter alia, age, topography of the oral cavity, scientificreportsonsynergisticactivityofthesesubstancesin
conditionofteeth,nutritionalandbreathinghabits,tobacco theimprovementoforalhygieneisinsufficient[33,34].The
smoking,oralhygiene,andlossofteethorusageofprosthetic aimofthisstudywastoevaluatetheinfluenceoftoothpaste
restorations[13].Theextentofsuchchangesdependsonthe with active substances of plant origin, such as ethanolic
type of dentures: fixed dentures cause local changes in the extractofpropolisandteatreeoil,onmicrofloraoftheoral
microflora, while removable dentures cause more complex cavityandtoevaluatethefollowingindices:OHI-S,denture
changes.Itisassociatedwiththeformationofnewreservoirs plaqueindex,mSBI,andAPIinpatientswithpartialdentures.
of microorganisms such as the intramucosal surface of the
denture base or clasps on retaining teeth. Acrylic base of
2.MaterialsandMethods
partial dentures covers a substantial surface of the mucosa,
creatingfavourableconditionsforthebuildupofbacteriaand
This study was conducted between September 1, 2015, and
fungiintheformofdentureplaque.Itisaplacecharacterised
March 1, 2016, in the Department of Prosthetic Dentistry,
by high humidity, elevated temperature, reduced oxygen
Akademickie Centrum Stomatologii i Medycyny Specjal-
supply, and impaired conditions for salivary self-cleaning.
istycznej (Bytom, Poland), and in Specialist Dental Clinic
Moreover, the acrylic material, due to its heterogeneous
(Katowice, Poland), which provide comprehensive dental
and porous structure, absorbs water and swells in the oral
careforpatientswithremovabledentures.
cavity,furtherfacilitatingtheaggregationofmicroorganisms
[14–16]. Perfect oral hygiene is also the basic method for
prevention of caries and diseases of the periodontium and 2.1. Toothpaste Preparations with Propolis and Tea Tree Oil.
oral mucosa. Caries and periodontal diseases are the most Twosamplesoftoothpastecoveredanonymouslywithblank
commoncauseofteethloss.Therefore,itiscrucialtoprotect whitetagandmarkedonlywithletterAorCwerecompared:
residualdentitioninpatientswithpartialdenturesfromthese the studied toothpaste with the active ingredient (study
diseases.Thebasichygienictreatmentismechanicalremoval group,A),1,5%contentofethanolextractofPolishpropolis,
ofplaque,supportedby,forexample,antimicrobialchemical and with no active ingredients, placebo (control group, C).
agents [17, 18]. The positive effect of ethanolic extract of Thesecondnaturalproductvaluedforitspropertieswastea
propolis and tea tree oil on the maintenance of proper oral treeoil(TTO).ItisobtainedfromtheMelaleucaalternifolia
hygienehasbeenknownforalongtime.Theyarenaturally plant.Twootherspecies(MelaleucalinariifoliaandMelaleuca
Evidence-BasedComplementaryandAlternativeMedicine 3
dissitiflora)arealsousedtoobtainessentialoil,onlyifthey (iv)patients with class V fillings or acrylic/ceramic
containmorethan30%terpinen-4-olintheessentialoil.The crownsaroundtheretainingteeth,
majorcomponentofteatreeoilisterpinen-4-ol.Itscontent
(v)patientswithcancer,
is29–45%.Thecompositionofactiveandplacebotoothpaste
hadthefollowing: (vi)psychosomaticdisorders,
(vii)patientsaftertraumawithinthecraniofacialregion,
ActiveToothpaste
(viii)pregnantandlactatingfemales,
aqua(upto100%ofweight),
glycerine(5–12%), (ix)patients suffering from asthma or atopic dermatitis,
allergictofoods,drugs,orhoneyanditsproducts,or
silica(10–14%),
havingotherallergy-relatedailments.
sorbitol(10–20%),
hydroxyethylcellulose(0.1–1%), The study was conducted with the prior approval from the
Bioethics Committee of the Silesian Medical Chamber in
titaniumdioxide(0.5–2%),
Katowice, Poland, Resolution number 8/2015 of March 23,
xanthangum(0.3–1%), 2015.
ethanolicextractofpropolis(EEP)(1.0%),
teatreeoil(TTO)(1.0%), 2.3. Patients. 37 patients aged 41–82 (57.0 ± 10.49) were
qualified for the study, including 20 females (54%) and
mentholoil(0.2%),
17 males (46%). Patients were randomly divided into two
rosemaryoil(0.1%).
subgroups.Thestudygroupconsistedof18personsaged41–
79 (55.05±11.09), who received the active toothpaste with
PlaceboToothpaste
propolisandteatreeoil,whereasthecontrolgroupconsisted
aqua, of19patientsaged41–82(66.79±11.19),whoweregiventhe
negativecontroltoothpaste.
glycerine,
silica,
2.4. Clinical Examination Protocol. The studies were per-
sorbitol,
formed in two stages. The first stage consisted of a general
hydroxyethylcellulose, medicalanddentalinterviewbasedonasurveyandaform
titaniumdioxide, writtenparticularlyforthispurpose.Thestudywasextended
byasurveyconcerningmedicalhistoryandcurrentdiseases,
xanthangum.
allergies,takenmedication,andenvironmentalrisk.Mobility
of the retaining tooth was also measured with Periotest
2.2. Study Groups Inclusion and Extrusion Criteria. Subject
duringthefirstvisit,asacriterionqualifyingthepatientfor
qualificationforthestudywasbasedonmedicalanddental thestudy.Thevaluesmaybeinthefollowingranges:−08to
history,interview,andreviewofclinicalrecords.Theexclu-
+09,physiologicalteethmobility;+10to+19,perceptibleteeth
sion and inclusion criteria from the investigation were as
mobility; +20 to +29, moderate teeth mobility under inves-
follows:
tigator’s pressure; +30 to +50, severe teeth mobility under
pressurefromthelipsortongue[35,36].Patientsqualifiedfor
InclusionCriteria
the study were evaluated to determine the value of selected
(i)writtenparticipationconsent, oral hygiene indices included in the form, that is, OHI-S,
denture plaque index, mSBI, and API. The measurement
(ii)age(40–85years),
procedure according to the above-mentioned indices was
(iii)patientslacking5–8teethinthemaxillaandmandible
carriedoutwithuseofastandardWHOperiodontalprobe.
underprosthetictreatment,
The measurementwas based on the assessment ofpresence
(iv)patients with remaining posterior occluding pairs and size of plaque on selected tooth surfaces and bleeding
withaminimumoftwosupportingzonesaccording fromtheinterdentalpapilla.
toEichnerindex. Approximal Plaque Index (API), Lange and Ainamo,
1988:Presenceofplaqueisassessedonlyonocclusalsurfaces.
ExtrusionCriteria Thepresenceofbacterialplaqueisexaminedinquadrants1
and3onocclusalsurfacesonthesideoftheoralcavityproper
(i)lackofawrittenparticipationconsent,painfulmasti-
andinquadrants2and4onocclusalsurfacesonthesideof
catorydysfunction,
thevestibule.TheformulausedtocalculatethevalueofAPI
(ii)edentulouspatientsorpatientswithresidualdentition
[%]=thesumofinterdentalspaceswithplaque/thesumof
(EichnersubgroupsC1–C3),patientswithfulldental allexaminedsurfaces×100%.Indexvalueswereinterpreted
archesorlacking1-2teethinanarch,
as follows: API 100–70%, bad oral hygiene; API 70–40%,
(iii)patientswithretainingteethmobilityover+20PTV averagehygiene,improvementrecommended;API39–25%,
testedwiththePeriotestinstrument, quitegoodoralhygiene;API<25%,optimaloralhygiene.
4 Evidence-BasedComplementaryandAlternativeMedicine
Simplified Oral Hygiene Index determines the amount sets: ENTEROtest 24N, NEFERMtest 24N, STREPTOtest
of soft debris or calculus on the four buccal surfaces of the 24, STAPHYtest, ANAEROtest 23, OXItest, PYRAtest, and
selected teeth: upper right first molar, upper right central TNW lite 6.5 computer program for species identification
incisor,upperleftfirstmolar,andlowercentralincisor,and of microorganisms (Erba-Lachema, Brno, Czech Republic).
onthelingualsurfacesofthelowerleftfirstmolarandlower Also the following Biomerieux (Marcyl’Etoile, France) bio-
rightfirstmolar. chemical tests were used: Katalaza, Slidex Staph Kit, and
Denture plaque index (Budtz-Jo¨rgensen, 1978) provides API Candida. Performance and interpretation of results of
the following description for the amount of denture plaque the tests was carried out according to the manufacturer’s
onthefittingsurface:0=nonvisible,1=lessthanone-third recommendationswithdiagnosticreagentsets.
covered,2=one-thirdtotwo-thirdscovered,and3=more
thantwo-thirdscovered.
2.6. Statistical Analysis. The first stage of statistical analysis
The periodontal status (gingival health) was evaluated
consistedofverificationofthecompatibilityoftheobtained
with use of the modified Sulcus Bleeding Index (mSBI,
index values and the number of bacteria with normal dis-
Muhlemann-Son,1971)andbyrecordingonly“bleedingpres-
tribution with use of the Shapiro-Wilk test. Variables with
ence”or“bleedingabsence”forallexistingteeth.Theindex
normal distribution were presented with arithmetic mean
describes the presence of localised bleeding in interdental
and standard deviation, whereas nonparametric variables
spaces and interdental papilla. In quadrants 1 and 3 the
were presented with median and interquartile range. One-
vestibular side is examined, while in quadrants 2 and 4 the
way analysis of variance (ANOVA) and Levene’s test were
lingualoneisexamined.
used to compare the results of the study group with the
After clinical examination acrylic partial dentures were
controlgroupforOHI,dentureplaqueindex,andthenumber
madeforall37patients.Bothgroups(studyandcontrol)were
of bacteria. The comparison of results between groups was
subjectedtohygienisationoftheoralcavityandinstructedon
performedwiththeTukey-Kramermethod.Resultsobtained
howtoproperlybrushtheirteethwithuseofFonescircular
for OHI and denture plaque index were compared with
technique. Fones circular technique consists of putting the
Student’s𝑡-testfordependentandindependentsamples.For
toothbrushperpendicularlytoslightlyopeneddentalarches
unrelated variables of API and SBI the results of study and
and performing circular movements. In the roll technique
controlgroupswerecomparedwithMann–Whitney𝑈test,
thetoothbrushisplacedonattachedgingivaattheangleof
∘ Wilcoxon signed-rank test, and Friedman’s ANOVA with
45 withthebristlespointedtowardstheroot.Patientswere
Kendall’scoefficientofconcordance.Theresultsweredeemed
alsoshowntheproperdenturehygiene.Theywereinstructed
tobestatisticallysignificantif𝑝<0.05.
to wash their dentures after every meal with use of a soft
denturebrushorwithhandsandhardsoap.Afterbrushing
thedenturesweretobecarefullyrinsedunderrunningwater 3.Results
and put in the oral cavity. Patients were advised not to use
dentures on a 24/7 basis. Dentures were supposed to be 3.1. Oral Health Conditions. The general distribution range
removed for the night and kept in a dry place after prior for API was presented in Table 1. Statistically significant
washing. All patients were examined in three subsequent differencesbetweengroupswerereported7and28daysafter
stages: preliminary qualification at baseline before hygiene theinitialexamination(𝑝 < 0.05and𝑝 < 0.001).Improve-
procedure (1st assessment), a follow-up after 7 days (2nd ment of hygiene was reported both in the study group (A)
assessment),andafollow-up28daysaftertheinitialexamina- and in the controlgroup(C). It maybe associated with the
tion(3rdassessment).Patientswereexaminedforregression reduction of interproximal spaces in the lateral segments
of lesions, frame of mind, and possible side effects, which ofmaxillaand/ormandible.Asignificantimprovementwas
wasrecordedonthecontrolform.Duringeachexamination onlyobservedinthestudygroup(A)(Table1).
swabs were used for microbiological examination of oral Another assessed index was the simplified Greene-
cavitymicroflora.Themicrobialmaterialforsmeartestswas Vermilion hygiene index. Statistically significant improve-
collectedfromthefloorofthemouth. ment of hygiene in the study group (A) was observed
(Table2).Inthestudygroup(A)theindexwasdetermined
2.5. Bacterial Strains Isolation and Microbiological Investi- after7and28daysofusage(𝑝=0.013221,𝑝=0.000468).No
gation. Microbiological examinations were performed with statisticalsignificancewasobservedinthecontrolgroup(C)
use of classic methods employed in microbiological diag- (Table2).Significantreductionwasobservedbycomparing
nostics. Material collected from the patients was cultured OHI7and28.
on proper growth media for proliferation and subsequent Basedontheconductedanalysisofdentureplaqueindex
isolationofpurecultures.Aerobicbacteriawereproliferated values no significant reduction in the values was observed
on Columbia agar solid medium with 5% sheep blood at aftertheperiodofuseoftoothpastesinstudy(A)andcontrol
∘
37 C. Anaerobic bacteria were proliferated on Schaedler (C)groupscomparedtotheoutputvalue(Figure1).
∘
K3 solid medium with 5% sheep blood at 37 C in anaer- Statistically significant reduction of gingival bleeding
obic conditions obtained with Biomerieux GENbag anaer indexafter7and28daysoftheuseoftoothpastescomparedto
generators (Marcy-l’E´toile, France). Species identification thegingivalbleedingindexatthebeginningofthetreatment
was performed after isolation and proliferation of cultured wasobservedbothinstudy(A)andinthecontrolgroup(C)
microorganism strains with use of the following reagent (Table3).Thestudyandcontrolgroupdidnotdifferinterms
Evidence-BasedComplementaryandAlternativeMedicine 5
Table1:APIranges:assessmentforstudyandcontrolgroups.
Oralhygieneassessment(interproximalspaces)
Friedman’sANOVAtest
Theassessmentcriteria T1 T2 T3 (𝑝)/Kendall’scoefficient Wilcoxontesstig𝑝ned-rank
ofconcordance
Optimal 11% 39% 56%
(1):(2)=0.000438
Studygroup(A) Quitegood 15% 28% 44% 𝑝=0.0000/0.95894 (2):(3)=0.000293
Average 53% 33% 0% (1):(3)=0.000196
Bad 21% 0% 0%
Optimal 17% 21% 42%
(1):(2)=0.231060
Controlgroup(C) Quitegood 11% 37% 37% 𝑝=0.00228/0.32012 (2):(3)=0.064186
Average 72% 42% 21% (1):(3)=0.000327
Bad 0% 0% 0%
Mann–Whitney“𝑈”test(𝑝) 0.784483 0.046554 0.001032 — —
T1(1):preliminaryexaminationbeforehygieneprocedure;T2(2):examinationafter7days;T3(3):28daysaftertheinitialexamination.
Table2:OHIranges:assessmentforstudyandcontrolgroups.
Oralhygieneassessment(interproximalspaces)
Mean± Mean± Mean±
T1 standard T2 standard 𝑝 T3 standard 𝑝
deviation deviation deviation
0–0.5 55.5% 0–0.5 66.6% 0–0.5 88.8%
Studygroup(A) 0.6–1 44.5% 0.41±0.25 0.6–1 34.4% 0.33±0.23 0.013221 0.6–1 11.1% 0.20±0.17 0.000468
1.1–2 0% 1.1–2 0% 1.1–2 0%
2.1–3 0% 2.1–3 0% 2.1–3 0%
0–0.5 58% 0–0.5 63% 0–0.5 68.4%
Controlgroup 0.6–1 21% 0.64±0.50 0.6–1 21% 0.58±0.30 0.352356 0.6–1 31.6% 0.57±0.23 0.382331
(C) 1.1–2 16% 1.1–2 16% 1.1–2 0%
2.1–3 5% 0.006370 0.000002 2.1–3 0%
Study(A)versus
0.097301 0,006370 0,000002
control(C)(𝑝)
T1(1):preliminaryexaminationbeforehygieneprocedure;T2(2):examinationafter7days;T3(3):28daysaftertheinitialexamination.
oftheassessmentoftheSBIvalueuponinitialexamination. They are all part of the oral flora and, more specifically,
After 7 and 28 days of use the control group exhibited a dentalbiofilmbacteria.Lactobacillusacidophilusisaknown
significantlyhigherlevelofSBIincomparisonwiththestudy pathogen responsible for the development of caries. Acti-
group(Table3). nomyces spp. belong to pioneer bacteria responsible for the
formation of plaque, while A. israeli may be an etiological
3.2.MicrobiologicalInvestigation. Thefollowingobservations factor for actinomycosis. The following species were also
in study group (A) constituted by 18 patients were made: eliminated:BacteroidesuniformisandParabacteroidesdista-
77 microorganisms of 31 species were isolated in the first sonisclassifiedasGram(−)anaerobes;Acinetobacterfreundii,
microbiologicaltest.Thesecondtest(after7daysofuseofthe Burkholderiacepacia,andNeisseriasicca(Gram(−)aerobe),
toothpastewithEEPandTTO)resultedintheisolationof78 as well as Staphylococcus epidermidis MSCNS (Gram(+)
microorganismsof27species.Thethirdtestafter4weeksof aerobe),whichcancauseopportunisticinfections,including
useoftheactivetoothpasteshowedadecreaseinthenumber sepsis.Themicrofloragainedthefollowingspecies,Gram(+)
of isolated microorganisms to 65, of 24 species (Table 4). anaerobes:AtopobiumminutumandClostridiumbutyricum,
Theoralmicrofloradiversitywasreducedbythedecreasein Gram(−)anaerobes:CapnocytophagaochraceaandPrevotella
thenumberofculturedmicroorganismspecies.Thefollow- melaninogenica,Gram(+)aerobes:Microccusspp.andStrep-
ing bacterial species were eliminated: Actinomces viscocus, tococcusoralis,andGram(−)aerobes:Providenciarustigiani
Actinomyces israelii, Atopobium parvulum, Bifidobacterium and Stenotrophomonas maltophilia. Micrococcus spp. and S.
ovatus,Clostridiumbotulinum,Eubacteriumsaburreum,Lac- oralis are classified as microorganisms which participate
tobacillus acidophilus, Lactobacillus fermentum, and Propi- in plaque formation as one of the first. Capnocytophaga
onibacterium granulosum, classified as Gram(+) anaerobes. ochracea belongs to the green bacterial complex, associated
6 Evidence-BasedComplementaryandAlternativeMedicine
Table3:SBIranges:assessmentforstudyandcontrolgroups.
SulcusBleedingIndexassessment
Friedman’sANOVAtest
Theassessment T1 T2 T3 (𝑝)/Kendall’scoefficient Wilcoxonsigned-ranktest𝑝
criteria
ofconcordance
Normalgingiva
50% 72.5% 100% (1):(2):0.003283
Studygroup(A) SBI<10% 𝑝=0.00000/0.71010 (2):(3):0.001474
Bleedingonprobing 50% 27.5% 0% (1):(3):0.000982
Normalgingiva
53% 65.5% 65.5% (1):(2):0.726768
Controlgroup(C) SBI<10% 𝑝=0.09760/0.12247 (2):(3):0.286004
Bleedingonprobing 47% 34.5% 34.5% (1):(3):0.572082
Mann–Whitney𝑈test(𝑝) 0.346197 0.012713 0.000001 —
T1(1):preliminaryexaminationbeforehygieneprocedure;T2(2):examinationafter7days;T3(3):28daysaftertheinitialexamination.
Type of the group; expected average edge 4 weeks of use of the toothpaste without active ingredients
Wilk’s Lambd；=,96734,F(3,33)=,37141,p=,77414 resulted in the culture of 92 microorganisms of 26 species
Decomposition of effective hypotheses (Table 4). An increase of the number of cultured microor-
Vertical bars represent confidence intervals .95 ganismsandaslightincreaseinthenumberofspecieswere
2,1
observed. The following species were eliminated: Gram(+)
2,0 anaerobes: Clostridium sporogenes, Eubacterium saburreum,
1,9 andLactobacillusfermentum,andGram(−)anaerobes:Bac-
es) 1,8 teroides ovatus, Bacteroides uniformis, Fusobacterium mor-
alu 1,7 tiferum,andPrevotellaoralis,aswellasGram(+)aerobicbac-
x (v 1,6 terium Staphylococcus aureus MSSA. The microflora gained
de thefollowingGram(+)anaerobes:Bifidobacteriumdentium,
n 1,5
e i Bifidobacterium ovatus, Clostridium chauvoei, Gemella mor-
u 1,4
aq billorum, and Propionibacterium propionicum. Bifidobac-
pl 1,3
e terium spp. are probiotic bacteria dwelling in human intes-
ntur 1,2 tine. Gemella morbilorum is a pathogen isolated from cysts
De 1,1 whichwerenotresolvedafterthesecondattemptofendodon-
1,0 tic treatment. The flora also gained Gram(−) anaerobic
0,9 bacterium Veillonella parvula, Gram(+) aerobic bacterium
0,8 Streptococcusmutans,andGram(−)aerobicbacteriumNeis-
Study group Control group seriasicca.Streptococcusspp.participateinplaqueformation,
Type of the group while S. mutans is a known etiological factor for caries. V.
parvula, which belongs to the purple complex according to
Denture plaque index T1 Socransky,isisolatedinpatientswithhealthyperiodontium.
Denture plaque index T2 ThenumberofculturedCandidaalbicansalsoincreased.As
mentioned before, this fungus is often isolated in patients
Denture plaque index T3
with removable dentures. The number of the following
Figure1:ANOVAEstimatedMarginalMeansfordentureplaque bacteria declined in the control group: Blautia producta,
indexstudyandcontrolgroup.
Clostridium clostridiforme, Lactobacillus acidophilus, Cap-
nocytophaga ochracea, and Streptococcus sanguinis, and the
number of the following bacteria increased: Actinomyces
withadvancedperiodontitisaccordingtoSocransky.Inour israelii,Actinomycesnaeslundii,Campylobactergracilis,Strep-
study the number of the following bacteria declined: Bifi- tococcusmitis,Streptococcussalivarium,Escherichiacoli,and
dobacterium adolescentis, Veillonella parvula, Streptococcus Neisseriasubflava,aswellasfungiCandidaalbicans(Table5).
salivarius, and Streptococcus sanguinis. They all belong to
physiologicalbacterialfloraandparticipateintheformation
3.3.SideEffects. Noserioussideeffectshavebeenreported.
ofdentalbiofilms.AreductionofCandidaalbicanswasalso
Somepatientsreporteddiscolorationoftheirtoothbrush.
observed(Table5).C.albicansisoftenisolatedfromtheoral
cavity, although most frequently in patients using dentures,
anddoesnotalwayscauseinfectionsymptoms. 4.Discussion
In the control group (C) constituted by 19 patients,
79 microorganisms of 25 species were isolated in the first Naturalextractsusedinthetestedtoothpasteswereintended
microbiologicaltest.Thesecondtestresultedintheisolation to provide chemical assistance in the mechanical removal
of83microorganismsof28species,whilethethirdtestafter of plaque and thus improve oral hygiene. The first one was
Evidence-BasedComplementaryandAlternativeMedicine 7
Table4:Themicroorganismsspeciesisolatedfrompatients,whousedtoothpastewithEEPandTTO(studygroup,A)orwithoutEEPand
TTO(controlgroup,C).
Numberofpatientsfromstudy Numberofpatientsfromcontrol
Isolatedmicroorganisms group(A) group(C)
Initial After7days After28days Initial After7days After28days
Gram(+)anaerobicbacteria
Actinomycesviscosus 1 1 0 0 0 0
Actinomycesisraelii 1 1 0 2 3 3
Actinomycesnaeslundii 1 2 1 3 7 7
Anaerococcusprevotti 0 0 0 0 1 0
Atopobiumminutum 0 0 1 0 1 0
Atopobiumparvulum 1 0 0 0 0 0
Bifidobacteriumadolesccentis 2 1 1 1 0 1
Bifidobacteriumdentium 1 0 1 0 1 3
Bifidobacteriumlongum 4 5 4 0 1 0
Bifidobacteriumbreve 0 0 0 5 2 5
Bifidobacteriumovatus 3 2 0 0 0 1
Blautiaproducta 1 0 2 2 0 1
Clostridiumbarati 1 1 1 0 2 0
ClostridiumbotulinumbiovarA 1 1 0 0 0 0
Clostridiumbutyricum 0 0 1 0 0 0
Clostridiumchauvoei 0 1 0 0 0 1
Clostridiumclostridiforme 0 0 0 2 1 1
Clostridiumhistolyticum 0 1 0 0 0 0
Clostridiumperfringens 0 1 0 1 1 1
Clostridiumsporogenes 0 0 0 1 0 0
Eubacteriumsaburreum 1 1 0 1 0 0
Gemellamorbillorum 0 0 0 0 0 1
Lactobacillusacidophilus 1 0 0 3 3 1
Lactobacillusfermentum 1 1 0 1 1 0
Propionibacteriumgranulosum 1 1 0 0 1 0
Propionibacteriumpropionicum 0 0 0 0 1 1
Gram(−)anaerobicbacteria
Bacteroidesovatus 0 0 0 1 0 0
Bacteroidesuniformis 1 0 0 1 0 0
Campylobactergracilis 0 0 0 1 1 2
Capnocytophagaochracea 0 1 1 3 2 1
Fusobacteriumnucleatum 1 1 1 0 1 0
Fusobacteriummortiferum 0 0 0 1 0 0
Mitsuokellamultacida 3 3 4 5 4 5
Parabacteroidesdistasonis 1 1 0 0 0 0
Prevotellamelaninogenica 0 0 1 0 0 0
Prevotellaoralis 0 0 0 1 0 0
Veillonellaparvula 2 3 1 0 1 1
Gram(+)aerobicbacteria
Micrococcusspp. 0 0 1 0 0 0
StaphylococcusaureusMSSA 1 0 2 1 0 0
StaphylococcusepidermidisMSCNS 1 0 0 0 0 0
Streptococcusmitis 7 10 7 6 7 9
Streptococcusoralis 0 1 1 0 0 0
8 Evidence-BasedComplementaryandAlternativeMedicine
Table4:Continued.
Numberofpatientsfromstudy Numberofpatientsfromcontrol
Isolatedmicroorganisms group(A) group(C)
Initial After7days After28days Initial After7days After28days
Streptococcusmutans 0 0 0 0 0 2
Streptococcussalivarius 9 5 8 7 4 10
Streptococcussanguinis 4 5 2 5 7 3
Gram(−)aerobicbacteria
Acinetobacterfreundii 1 0 0 0 0 0
Burkholderiacepacia 1 0 0 1 0 1
Enterobacteramniogenus 0 1 0 0 0 0
Enterobactercloacae 0 1 0 0 0 0
Enterobacterspp. 0 0 0 0 1 0
Escherichiacoli 0 0 0 1 1 2
Neisseriasicca 1 0 0 0 2 1
Neisseriasubflava 15 18 16 18 16 19
Providenciarustigiani 0 0 1 0 0 0
Pseudomonasspp. 0 0 0 0 1 0
Stenotrophomonasmaltophilia 0 0 1 0 0 0
Fungi
Candidaalbicans 8 8 6 5 9 9
Numberofmicroorganismsstrains 77 78 65 79 83 92
Altogether: 220 254
ethanolic extract of propolis. There are numerous studies Brazilianpropolis,inwhichtheyconfirmeditsantiphlogistic
confirmingitsstrongantimicrobial,antiviral,antifungal,and activity using three recognised methods. The same study
antiphlogisticactivity[37–39].Propolisexhibitsawidespec- includedanassessmentoftheinfluenceoftoothpastewith3%
trumofantimicrobialactivities.ResearchconductedbyKoru ethanolic extract of Brazilian propolis on oral hygiene with
et al. demonstrated that propolis is more effective against useoforalcavityindices:API,OHI,andSBI.Researcherscol-
Gram(+)aerobesthanagainstGram(−)aerobes[40].Intheir lectedalsosmearsfrompatientsinordertoassessantimicro-
study Feres et al. compared antimicrobial activity of 11% bialactivity.Patientsweredividedintotwogroups.Thestudy
propolisandthepopular0.12%chlorhexidineagainstbacteria group (I) was using an active toothpaste (T) with propolis,
insalivainhealthypatientsandinpatientswithperiodontal whilethecontrolgroup(II)wasusingatoothpastewithno
diseases. Both substances exhibited statistically significant activesubstance(G).Patientswereexaminedonthefirst,ini-
antimicrobial activity [41]. Antimicrobial activity of 10% tialvisit,secondvisitafteraweek,andthirdoneafter4weeks.
propolis and 0.12% chlorhexidine against S. mutans and L. After4weeksofusingtheTtoothpastetherewasastatistically
acidofilus,knownpathogensresponsibleforthedevelopment significant decrease in OHI in both groups I and II, which
ofcaries,wascomparedin2011.Bothprovedtobeeffective may indicate that daily mechanical plaque removal, even
[42].Anotherstudyfrom2013compared0.12%chlorhexidine withoutactivesubstances,canbeeffectiveinhealthypatients.
with propolis at a much lower concentration, namely, 2%. API in group II did not change and patients were qualified
ItsantimicrobialactivityagainstS.mutansandL.acidofilus forthegroupwithaveragehygiene,whereasAPIingroupI
was confirmed [43]. These studies proved the cariostatic improvedafter7daysandreturnedtobaselineuponthelast
properties of propolis, antiperiodontal pathogen activity, examination,patientwithquitegoodhygiene.SBIimproved
and efficiency at lower concentrations. In the present study in group I, but in a statistically insignificant way. It did not
we employed 1.5% ethanolic extract of propolis. Hitherto, changeingroupII[45].Studyoftoothpasteswith3%ethano-
there have been no studies confirming the antimicrobial lic extract of propolis in patients undergoing orthodontic
effectiveness at the concentration of 1.5%. However, in the treatmentwasconductedin2013andin2016.Apartfromthe
toothpaste tested it was combined with tea tree oil, which smears,itcomprisedalsoAPIaccordingtoLange,Gingival
hassimilarproperties.Thus,theconcentrationofEEPcould Index(GI),andOrthodonticPlaqueIndex(OPI)uponfirst
be lowered. At the same time, lower concentration of EEP examination and after 35 days of using a toothpaste with
minimisestheriskofsideeffectssuchasallergicreactionsand propolisandCTgel.Thecontrolgroupconsistedofpatients
burningsensationintheoralcavity[34].Animportantaspect using a toothpaste without propolis and CC gel. The first
istheantiphlogisticpropertiesofethanolicextractofpropolis examinationdidnotdemonstrateanystatisticallysignificant
[44].Skabaetal.conductedananalysisofethanolicextractof differencesintheassessedindices.Uponexaminationafter35
Evidence-BasedComplementaryandAlternativeMedicine 9
Table5:Thechangesoforalmicroflorafrompatients,whousedtoothpastewithEEPandTTO(studygroup,A)orwithoutEEPandTTO
(controlgroup,C).
Changesofmicroorganisms
Studygroup(A) Controlgroup(C)
speciesafter28daysofstudy
Gram(+)anaerobicbacteria
Actinomycesviscosus
Actinomycesisraelii
Atopobiumparvulum Bifidobacteriumovatus
Clostridiumsporogenes
Clostridiumbotulinum
Eubacteriumsaburreum
Eubacteriumsaburreum
Lactobacillusfermentum
Lactobacillusacidophilus
Lactobacillusfermentum
Propionibacteriumgranulosum
Eliminatedspecies Gram(−)anaerobicbacteria
Bacteroidesovatus
Bacteroidesuniformis Fusobacterium
Parabacteroidesdystasonis mortiferum
Prevotellaoralis
Gram(+)aerobicbacteria
Staphylococcusaureus
StaphylococcusepidermidisMSCNS
MSSA
Gram(−)aerobicbacteria
Acinetobacterfreundii
Burkholderiacepacia —
Neisseriasicca
Gram(+)anaerobicbacteria
Blautiaproducta
Clostridium
Bifidobacteriumadolescentis
clostridiforme
Lactobacillusacidophilus
Gram(−)anaerobicbacteria
Declinedspecies Capnocytophaga
Veillonellaparvula
ochracea
Gram(+)aerobicbacteria
Streptococcussalivarius
Streptococcussanguinis
Streptococcussanguinis
Fungi
Candidaalbicans —
Gram(+)anaerobicbacteria
Bifidobacteriumdentium
Bifidobacteriumovatus
Atopobiumminutum Clostridiumchauvoei
Clostridiumbutyricum Gemellamorbillorum
Propionibacterium
propionicum
Gram(−)anaerobicbacteria
Gainedspecies
Capnocytophagaochracea Prevotella
Veillonellaparvula
melaninogenica
Gram(+)aerobicbacteria
Micrococcusspp.
Streptococcusmutans
Streptococcusoralis
Gram(−)aerobicbacteria
Providienciarustigiani
Neisseriasicca
Stenotrophomonasmaltophilia
Gram(+)anaerobicbacteria
Actinomycesisraeli
Blautiaproducta
Actinomycesnaeslundii
Gram(−)anaerobicbacteria
Increasedspecies Mitsuokellamultiacida Campylobactergracilis
Gram(+)aerobicbacteria
Streptococcusmitis
StaphylococcusaureusMSSA
Streptococcussalivarius
10 Evidence-BasedComplementaryandAlternativeMedicine
Table5:Continued.
Changesofmicroorganisms
Studygroup(A) Controlgroup(C)
speciesafter28daysofstudy
Gram(−)aerobicbacteria
Neisseriasubflava
Neisseriasubflava
Escherichiacoli
Fungi
— Candidaalbicans
daystherewasastatisticallysignificantdecreaseinthevalues Inthepresentstudytherewasanincreaseinthenumber
ofGIandOPIcomparedtothecontrolgroup.Nosignificant of cultured C. albicans colonies in patients using both the
differencewasshowninthedecreaseofAPI[46,47]. active toothpaste and the placebo. Both active substances
Melaleucaalternifoliateatreeoil(TTO)isalsoaknown employed in the active toothpaste exhibit antifungal activ-
antiseptic preparation with antimicrobial and antifungal ity. However, it should be remembered that the group of
activity. It has been successfully used for many centuries in examinedpatientswasusingacrylicpartialdentures,which
dermatology, particularly in the treatment of acne vulgaris are an iatrogenic factor for C. albicans infection [55]. The
[48], accelerates wound healing, and has anticancer prop- use of acrylic partial dentures caused changes in the oral
erties [49]. The basic composition of the oil is terpinen- microflora.TheacrylicmaterialisareservoirforC.albicans,
4-ol, 𝛾-terpinene, 𝛼-terpinene, 1,8-cineole, terpinolene, 𝜌- whose amount depends on denture hygiene and its quality,
cymene,𝛼-pinene,𝛼-terpineol,aromadendrene,𝛿-cadinene, particularlyroughness[56,57].The2011studyofbeeprod-
limonene, sabinene, globulol, and viridiflorol. They are ter- uctsincomparisonwithfluconazoleconfirmedtheantifungal
penecompoundswithantisepticandantiphlogisticactivities activityofpropolisandthepossibilityofitsalternativeusein
which apart from dermatology can also be employed in antifungaltherapy.ItexhibitsactivityagainstC.albicans,C.
dentistry, particularly in the maintenance of proper oral glabrata,C.krusei,andTrichosporonspp.[58].Therearemany
hygiene [50]. In 2003 Hammer et al. examined in vitro reports on the antifungal activity of tea tree oil. The 2003
the antimicrobial activity of tea tree oil against 161 isolated invitrostudyexaminedtheantifungalactivityofparticular
oralpathogensanddetectedMICandMBCfor15ofthem. components of the oil in concentrations between 0.5% and
MIC and MBC values were lowest for Porphyromonas spp., 2%.Theresultsdemonstratedthatallcomponentsoftheoil,
Prevotella spp., and Veillonella spp. and highest for isolated apart from 𝛽-myrcene, exhibit antifungal activity [59]. The
Streptococcus spp., Fusobacterium spp., and Lactobacillus studyfrom2004confirmedthehypothesisofthemechanism
spp. Time necessary for the death of Streptococcus mutans of action of tea tree oil against C. albicans, C. glabrata, and
and Lactobacillus rhamnosus in ≥0.5% concentration was 5 S. cerevisiae. The oil influences the properties and disrupts
minutes. It was thus demonstrated that oral pathogens are thefunctionsoffungalcellmembrane[60].Studyconducted
susceptibletotheactivityofTTO[51]. by Sudjana et al. demonstrated that TTO inhibits adhesion
Studiesontoothpasteswithnaturalextractsagainstthree of C. albicans to human cells and polystyrene and inhibits
species of bacteria S. mutans, P. aeruginosa, and E. faecalis theformationofbiofilmthroughreductionofsurfacetension
comparedtoatoothpastewithoutantimicrobialingredients [61].
were conducted in 2015. S. mutans is a known etiological
factor for caries, while P. aeruginosa and E. faecalis are
5.Conclusions
pathogens isolated in periodontopathies and endodontic
infections. 8 toothpastes with the following composition
(1)Improvementsinhygieneandtheconditionofperi-
were used: sorbitol (I), tocopherol (II), mentha (III), cin-
odontium were observed in patients using tooth-
namon/mentha (IV), propolis/tea tree oil (V), mentha/ac¸a´ı
pasteswithethanolicextractofpropolisandteatree
berry (VI), mentha/guarana (VII), and propolis (VIII). The
oil.
controlgroupwasatoothpastewithnoantimicrobialproper-
ties(group(IX),negativecontrol),whilegroup(X)(positive (2)Thetoothpastewithethanolicextractofpropolisand
control) was a dental gel with triclosan and formaldehyde. tea tree oil had no influence on the reduction of
TheresultsdemonstratedthatE.faecalisexhibitedresistance dentureplaque.
to the toothpaste with propolis and tea tree oil and sus-
(3)Thestudiedtoothpastewithactiveingredients(etha-
ceptibilitytothetoothpastewithpropolisonly.Theauthors
nolicextractofpropolisandteatreeoil)demonstrates
explain it with a different concentration of propolis in the
thebeneficialinfluenceonmicroorganismscomposi-
toothpastes. Propolis/tea tree oil toothpaste exhibited the
tioninoralmicroflora.
strongestantimicrobialactivityagainstS.mutanscompared
to other toothpastes. P. aeruginosa proved to be resistant to
all tested natural substances [52]. Some other researchers CompetingInterests
demonstratealternativemethodstoreducepathologicaloral
microflora,byadditiontoresincompositesonelementions The authors declare that there is no conflict of interests
withnanogoldandnanosilver[53,54]. regardingthepublicationofthispaper.