Each loop was calibrated to hold 10 cc of mouth air. When collecting the 10-cc sample, a 50-cc polyethylene syringe was attached to one end of the loop while a ...
j. Soc.Cosmet. Chem.,44, 101-107 (March/April1993)
Relationshipbetweensensoryand instrumental evaluations of mouth odor HOLLANDRA
P. NILES and ABDUL GAFFAR,
Colgate-Palmolive Technical Center,909 RiverRoad, Piscataway, NJ 00854.
Received March20, 1992. Presented in part at theannualsession of the AADR, Washington, D.C., March1986. Synopsis
Two methodsarecurrentlyavailableto assess mouthodor:subjective evaluation (sensory rating)andgas chromatographic (GC) analysis of the volatilesulfurcompounds (VSC),hydrogen sulfide,methylruercaptan, anddimethyl sulfidein mouth air. The purposeof this studywasto examinethe correlationbetween thesetwo methodsin a humanclinicalstudy.Twentyadultsparticipatedin the study.The morningmouth
odorof eachsubjectwasassessed by two expertsensory evaluators on a scaleof 0 to 8 (showingincreasing odorintensity).Followingsensory evaluations, mouthair samplesfromeachsubjectwerethen obtainedand analyzedby GC for VSC. The studywasdividedinto threephases,viz., a control,a test, and a recovery phase.Eachphaselastedfor threedays,andtwo readings on the secondandthird dayof eachphasewere obtainedby both methods.During the test phase,the subjectsbrushedtheir teeth twice a day with a standardfluoride-containing dentifrice,and morning mouth odor determinationswere conductedas describedabove.During the controland recoveryphase,the subjectspracticedoral hygienead libitum. Correlationcoefficients (r) betweensensory ratingsandGC readings(VSC--ng/ml) of mouthair were0.22, 0.77, and 0.78 during the control,the test, and the recoveryperiods,respectively.The latter two were significantat p = 0.01.
The ratiosof the GC readingto the sensory rating wereconsistent overthe rangeof valuesobtained;an average factorof 3.2 wouldpredictthe corresponding GC readingfor a givensensory ratingin this study. For example,VSC by GC of 25.8 ng/ml of mouthodorprovideda sensory ratingof 8 (strongodor).This indicates that the objectiveevaluation of mouthodorby GC doescorrelate with a subjective sensory rating of mouthodor.The GC methodthereforeprovidesuswith usefulinstrumentalmeasurements (ng/ml) that can be translatedinto a consumer-perceivable odorintensity.
INTRODUCTION
It hasbeenpreviouslyshownthat the volatilesulfurcompounds (VSC) suchas H2S, CH3SH, and (CH3)2Sareresponsible for localmouthodorin healthyindividuals(1). TheseVSC components arisefrom the putrefactionof salivaryproteinsandaminoacids by gram-negative oralmicroorganisms (2) andcanbe detectedin directmouthair by a gaschromatographic methodor by a sensorymethod(3). Sincethe sensorymethodis subjective,it is highly desirableto developan objectiveinstrumentalmethodto assess offensive local mouth odor. lol
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We havepreviouslyshownthat the instrumentalmethod is capableof assessing the effectof bothrinseanddentifricetreatmentson mouthodorin humans(4). The purpose of this study was to determinethe correlationbetweenthe instrumentalmethod and sensoryevaluationin a controlledclinicaltrial.
MATERIALS
AND
METHODS
A 220 Tracorgaschromatograph equippedwith a flamephotometricdetector,housing a 394 millimicronfilter specificfor sulfur,wasusedto measurethe sulfurcompounds responsible for mouth odor. A 25-ft teflon (FEP) Supelcocolumnwasalsoused(containing 0.05% polyphenylether and 0.25% phosphoricacid on a 30/60 meshchromosorb T) to specifically separate the primarysulfurcomponents of mouthair. Air wasusedas a carriergasto eliminatethe air peakand to reducethe analysistime. The
isothermal oventemperature was60øC,with inlet anddetectortemperatures of 130øC. A standardgasSO2 permeationtube was usedto convertthe resultsinto nanograms/ milliliter
for the total sum of sulfides.
The mouthair samplewascollectedin a 24-ft teflonstorageloop(x/s-ino.d. x x/x6-in i.d.) that waswrappedwith teflontapeandhada Mininert teflonvalveattachedto each end (Figure 1). We haveshownin previousstudiesthat this loop is capableof storing mouth air without substantialsampleloss. Each loop was calibratedto hold 10 cc of mouth air. When collectingthe 10-ccsample,a 50-ccpolyethylenesyringewasattached to oneend of the loop while a 3-in sterilizedteflon tube wasattachedto the other end.
Figure 1. Storageloopfor mouthair samples.
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The tube was carefullyinsertedinto the panelist'smouth, both teflon valveswere opened,and the mouth air wasdrawn into the storageloop using the syringe.During the evaluationprocedure,panelistswereaskedto sit for 10 minuteswith their mouths closedprior to eachevaluationto allow for the buildupof the sulfurvolatiles.
The storedsamples weretransported to thelaboratoryin dry iceandanalyzedwithin five to sevenhoursfollowingcollection.Two sampleswere obtainedfrom eachsubjectfor analysis.The samplestorageloop was attachedto the gas chromatographinlet gas samplingvalve, wherethe storedmouth air samplewas introducedinto the GC. The gassamplingvalveapparatus wasalsocalibratedfor 10 cc of mouth air. A 50-cc syringewasattachedto the outlet portionof the valveon the GC in orderto draw the sampleinto the samplingvalve. When the valvewasturned, the air carriergasswept thesampleinto the column.The instrumentalreadingswereexpressed in nanograms per milliliter (ng/ml). The secondmethodfor the measurement of mouth odoris a subjectivesensoryevaluation. This evaluationwas conductedby two professional judgesfrom the La-Wall Harrison ResearchLaboratoriesimmediatelyfollowing the instrumentalevaluations. Panelistswaited for ten minutes with their mouths closed, similar to the instrumental
procedure.A specialboothwasconstructed that allowedthe paneliststo remainanonymous.A sterilethistle tube was insertedinto the panelists'mouthswhile the bellshapedend wasinsertedthroughthe screenwherethe judgeevaluatedthe mouth odor. Panelistswere askednot to breath during this evaluation.Each of the two judges evaluatedeachpanelist.The offensiveness of the odorwasratedon a scalefrom 0 to 8 (TableI), where0 is non-offensive and 8 is considered highly offensive.Two evaluations wereconductedon eachpanelist,oneevaluationby eachjudge.
STUDY
DESIGN
Twentypanelists (tenmenandten women)werequalifiedto participatein the studyby meetingthe inclusion/exclusion characteristics of the protocol.The inclusioncharacteristicsconsistedof beingof adult age, 20 yearsand above,havinggoodgeneralhealth, andhavinga total sulfurvolatilebaselinegreaterthan 5 ng/ml of mouthair. This level Table
I
SensoryEvaluationCriteria for Mouth Odor
Sensoryscore
Offensiveness of mouth odor
8
7
Strong
6 5
Definite
4 3
Faint
21
Doubtful
0
None
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wasdeterminedby a pre-studyscreening evaluation to identifythe qualifyingpanelists. The early morning screeningevaluationswere conductedusing the samemethod as describedbelowin the test phase.
The exclusioncharacteristics consisted of being a smoker,havinghard or soft tissue tumorsof the oralcavity,havingextensive cariesor periodontal problems,andusingfull or partialdenturesand/ororthodontic bands.Alsoexcluded from the studywerepanelistswho usedmedicationsuchasantihistamines, antibiotics,or othertypesof medicationfor two weeksprior to or duringthe study. The study consistedof three phases:control, test, and recovery.During the control phase,subjects usedtheir normalhygienepractices for threedays,with evaluations on the secondandthird days.During the testphase,subjects wereaskedto usea placebo dentifricein the morningandpriorto retiringat nightfortwo days(days1 and2), with evaluationson the secondand third day. Subjectswereaskedto brushwith the placebodentifricefor 60 seconds and rinsewith water.During the recovery phasesubjects resumedtheir normalhygienepractices for two daysasin the controlphase,andwereevaluated on the second andthird daysof the recoveryphase.The purposeof this phasewasto ascertain any carryover effectfrom the test phase.Spearman's rank correlationwascomputedto determinethe relationship betweenthe instrumental readingof VSC andthe sensory ratings.
In all phases, subjects wereaskedto usetheirtreatmenttwicea day,in themorningand at night beforeretiring(days1 and2). Followingthe final nighttimetreatmenton the morning of day 3, organolepticevaluationsand samplestoragecollectionswere conducted.Foreachmorningevaluation subjects reportedto thetestsitewithoutbrushing, rinsing,eating,or drinkingin ordernot to influenceor washawaythe VSC formedin the mouthair. At this time, two baselinesamples wereevaluatedorganoleptically and two storagesamples werecollectedfromeachsubject.Subjects werethengivenbreakfast and the appropriatetreatment:normalhygienein the controlphase,the placebodentifricein thetestphase,or normalhygienein therecovery phase.Followingthemorning treatments,the subjectsreturnedin threehoursfor the post-treatment organoleptic evaluationsand samplestoragecollections.During this 3-hr interim betweenevaluations, subjectsare askednot to eat or drink.
RESULTS
TableII showsboththe meaninstrumental andsensory mouthodorscores of subjects groupedaccordingto treatmentperiodof the study. For example,the meanand standard deviationfor the sensoryratingsfor day 3 of the controlperiodis 7.15 +- 0.42, while the meanfor the instrumentalevaluationsis 21.86 ng/ml +- 4.18.
The resultsshowthat during the controlperiodthe correlationwaspoor(r = 0.22), while, duringthe testand recoveryperiod,the valueshadimprovedwith an r valueof 0.77 and0.78, respectively. Theselattervaluesweresignificantat a p-valueof lessthan 0.01.
Table III showsthe instrumentalmeanvaluesthat correspond to eachmeanintegerof eachsensory rating(i.e., 5, 6, 7, 8). All subjects with meansensory ratingsequalto 5, 6, 7, or 8 were identified,and their corresponding instrumentalmeanvalueswere
MOUTH
ODOR
Table
EVALUATION
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II
Mean InstrumentalValues vs Mean SensoryRatings
Controlperiod
Determination
Test period
Recoveryperiod
(mean -+ S.D.)
N
Day 2
Day 3
Day 2
Day 3
Day 2
Day 3
Sensoryrating
20
7.15 + 0.42
6.98 + 0.76
6.43 + 0.73
6.45 -+ 0.82
6.38 + 0.70
6.55 + 0.69
Volatile
20
21.86 -+ 4.18
22.77 + 6.00
20.14 + 6.12
20.16 + 4.97
23.12 -+ 6.13
22.22 -+ 4•06
sulfur
ng/ml Correlation (r) coefficient
0.22 (N.S.)
0.77 (p (0.01)
Table
Calculation
Meansensory ratings A
8 7.0 6.0 5.0
0.78 (p (0.01)
III
of Prediction
Factor
Meaninstrumental readings (total sulfurvolatiles,ng/ml) N*
4O 31 21 28
B
Factor = B/A
25.85 22.70 19.78 16.59
3.23 3.24 3.29 3.32
Average = 3.27
* Total of 120 determinationsfrom all threephasesof the study.
averages for eachsensory rating(5, 6, 7, or 8). This allowsus to calculatea prediction factor(B/A). Note that this predictionfactoris quite consistentover the sensoryand instrumentationvaluesobtainedin this study, providinganotherindicationof correlation betweensensoryand instrumentalscores. A sensory ratingof 5 wasthe lowestratingreportedfor anysubjectby the judgesduring this study.FromTable I, a ratingof 5 is the lowestratingfor "definite"offensivemouth odor. From Table III, a rating of 5 corresponds to an instrumentalscoreof 16.59. This instrumental
value could be considered a threshold for odor offensiveness. In other
words, if the total sulfidemeasurement in nanograms/milliliterfor a given subjectis below 16.59 ng/ml, it couldbe ascertained that the subject'smouth odorwould not be of "definite"offensiveness (5 rating) but of "faint" offensiveness.
DISCUSSION
The organoleptic (sensory) ratingis currentlyan acceptable in vivoprocedure for determining the efficacyof mouth rinsesand dentifricesin mouth odorreduction. On the other hand, previouschemical,massspectrometric,and gaschromatographic methodshaveestablished VSC asbeingthe primarysourceof offensivelocalmouthodor (4). Therefore,a numberof investigations havebeenconducted to establisha correlation betweeninstrumentaltechniques anda subjectivesensory methodof evaluatingmouth
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odor. For example,Schmidtetal. (5) showedthat VSC are detectedinstrumentallyby GC and flamephotometry,and correlatedthesescores with organoleptic ratings. We havepreviouslyutilized the GC instrumentalprocedureto assess the efficacyof rinsesagainstmouth odorin a clinicalstudy.Antibacterialrinsessignificantlyreduced VSC in mouthair samplesobtainedthreehourspostrinsing,while the placeborinsedid not significantlyreduceVSC. Theseresultsindicatedthat thismethodcouldbe usedfor
assessing the effectsof activeagentsagainstlocalmouthodor(6). It wasof interestto correlatethis instrumental(objectivemethod) with the sensory(subjectivemethod) rating procedure.
There were severalobstaclesin transformingthis instrumentalmethodologyinto a practicaltool for clinicaltrials in the field. The instrumentalsetupis largeandcumbersomeand thus cannotbe transportedreadilyto a clinical site. Also, the evaluations are normallydonein the morningbeforesubjectseat or drink. This makesthe scheduling inflexibleand restrictsthe numberof subjectswho canparticipatein the study. Lastly,no publishedcorrelation studiesof thisnaturewereconducted in a clinicalstudy framework,betweenthe sensoryandinstrumentalevaluations.To facilitatethis type of study, it wasvery usefulto havea deviceto collect,store,and preservethe VSC mouth air samples.Sucha devicecanthenbeusedto analyzethe storedsamples uponreturning to the laboratory.The storageloopsdescribed in this studyfulfilledtheserequirements. The storagesystemalsoprovidedus an opportunityto conducta true clinical study offsiteand to correlatetheseinstrumentalmeasurements with a sensoryevaluationdone
by expertjudgesprior to the instrumentalVSC determination. We obtained strong positive correlation(r = 0.77-0.78) during test and recovery periodsof the studybut lessthan a robustcorrelation(r = 0.22) during the control
period.The latter is attributedto a lackof familiarityby subjects with the procedures in the earlyphaseof the study.Thesesubjectsinitially haddifficultywith the organolepticandstoragesamplingprocedures. Basedon previousstudies,we areconfidentthat the judges,samplecollector,and analystwerenot the factorscontributingto the low correlationin the controlphase.The noteworthyfeatureof this studyis the development of a factorto predictsensoryevaluations usingthe instrumentalreadingsin this study. This providesus a usefultool to translateinstrumentalmeasurements into consumerperceivableodor intensity. Recently,Rosenberg et al. (6) measured the correlationbetweenthe sulfidemeasurement in the mouth air and sensoryratingsin forty-onesubjectsusinga portablesulfide monitor.The overallcorrelationcoefficient obtainedby the investigators (r = 0.603) waswithin the rangeof what we observed,althoughthe instrumentusedwasnot of the sensitivityor specificityto sulfidesascompared to theGC systemusedto measure sulfur volatiles
in mouth
air.
CONCLUSIONS
The clinicalapproachusedto studythe correlationbetweensensory measurements and instrumental
measurements of mouth odor indicated that:
1. There wasa goodcorrelationbetweenthe instrumentaland sensoryevaluations for mouth odor. The correlation coefficients were 0.22 for the control and 0.77 and 0.78
for the test and recoveryphases,respectively, betweenthe two methods.
MOUTH
ODOR
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107
2. Resultsalsoindicatedthat the correlationimprovedasthe subjectsbecomefamiliar with the samplingprocedures. 3. Bothmethodswereusefulfor measuringthe intensityof offensiveness of mouthodor.
REFERENCES
(1) J. Tonzetich,Production andoriginoforalmalodor.A reviewof mechanisms andmethods of analysis, J. Periodont. 48, 13 (1977). (2) M. C. Solis-Gaffar, J. T. Fischer,andA. Gaffar,Instrumental evaluation of odorproductby specific oralmicroorganisms, J. Soc.Cosmet. Chem.,30, 244-247 (1979). (3) J. Tonzetich,Directgaschromatographic analysis of sulfurcompounds in mouthair in man, Arch. Oral Biol., 16, 587 (1971). (4) M. C. Solis-Gaffar,H. P. Niles, W. C. Rainieri, and R. C. Kestenbaum,Instrumentalevaluationof
mouthodorin a humanclinicalstudy,J. Dent.Res.,54, 35! (!975). (5) S.C. Schmidt,S. R. Missan,J. W. Tarbot, and A. Cooper,The correlation betweenorganoleptic mouthodorratingsandlevelsof sulfurcompounds, OralSurf.,45, 560 (!979). (6) M. Rosenberg, G. V. Kulkarni,A. Basy,andC. A. G. McCulloch,Reproducibility andsensitivity of oralmalodormeasurements with a portablesulfidemonitor,J. Dent.Res.,70, !436 (199!).
