tooth color is an important issue for esthetic dentistry professionals and for patients 
 that wish to enhance the appearance of their smiles .
nowadays , at - home vital tooth 
 bleaching with custom trays is well accepted as a safe , conservative , low cost and 
 effective esthetic procedure for treatment of discolored teeth .
considering the active products available for vital 
 tooth bleaching , carbamide and hydrogen peroxide are the most commonly used agents for 
 different bleaching modalities , which vary according to peroxide concentration , mode of application 
 and time of exposure .
the whitening products available for vital tooth bleaching can be applied on tooth 
 surface at the office by a dentist or at home by the patient , under the dentist 's 
 supervision .
the only 
 concentration that so far received the seal of acceptance by american dental association 
 ( ada ) was the 101% carbamide peroxide , which assures its safety and efficacy for 
 at - home tooth bleaching with custom trays .
however , randomized clinical trials 
 showed that the increase of the carbamide peroxide concentration for home - use did not 
 increase the efficacy and longevity of the tooth whitening effect .
manufacturers have introduced many different bleaching agents ' concentrations and 
 methods with the premise of " improving " the whitening effect . as a consequence
, there was a significant increase of 
 the variety of in - office tooth bleaching products available in the market , which usually 
 uses high hydrogen ( 15 to 40% ) or carbamide ( 30 to 37% ) peroxide concentrations combined 
 or not with a supplementary light source ( i.e. laser , led , halogen lamp or plasma 
 arc ) . despite the method of activation of these agents , the data 
 available in the literature are conflicting . while some studies have reported that the 
 use of light sources can enhance the whitening effect , others have 
 shown that the use of light - activation with in - office tooth bleaching did not affect the 
 outcome of 35% hydrogen peroxide treatment .
considering the peroxide concentration and its time of application as the main factors 
 that can affect the effectiveness of tooth whitening , the aim of the present study was 
 to evaluate the effectiveness of three carbamide peroxide concentrations used in tooth 
 bleaching treatments .
the null hypothesis for this study was that the high - concentration 
 carbamide peroxide gel evaluated is not more effective than the low - concentration 
 carbamide peroxide bleaching agents .
sixty recently extracted bovine incisors without cracks and surface defects ( 40x 
 magnification ) were used in this study .
the roots were cut with a water - cooled 
 low - speed diamond saw ( isomet 1000 , buehler , lake bluff , il , usa ) and , the pulp 
 tissue was removed .
then , teeth were stored in 0.5% chloramine - t solution for one 
 week .
after disinfection , each crown was sectioned in both buccolingual and mesiodistal 
 directions to obtain one enamel - dentin slab ( 6x6x3 mm ) , which were measured with a 
 digital caliper ( dc 500 , mitutoyo , suzano , sp , brazil ) .
each slab was individually 
 embedded in self - curing acrylic resin in a ring model and then , enamel surfaces were 
 flattened with 600- and 1200-grit silicon - carbide paper under water cooling to create 
 a standardized buccal enamel surface .
samples were washed and stored in distilled 
 water at 37c for 1 week until starting the experimental procedures . in an attempt to standardize the tooth shade , an artificial staining of the specimens 

enamel surfaces were etched 
 with 35% phosphoric acid gel for 60 s and rinsed with distilled water for 30 s. 
 specimens were immersed daily over a 1-week period in a standardized coffee solution 
 at room temperature ( 252c ) in screw capped plastic universal containers .
the daily 
 renewed coffee solution was produced by boiling 12 g of coffee ( melitta , avar , sp , 
 brazil ) in 200 ml of distilled water for 5 min followed by filtering in a coffee 
 maker ( melitta ) .
after the immersion period , the specimens were washed and stored in 
 distilled water at room temperature .
tooth shades of the stained specimens were assessed with a digital spectrophotometer 
 ( vita easyshade , vita zahnfabrik , bad sackingen , germany ) after 1 , 3 and 7 days of 
 immersion in coffee solution ( baseline ) .
three measurements were done with the active 
 point of the spectrophotometer in the center of each sample .
the instrument 
 automatically averaged the three readings for each sample , which were use for overall 
 data analysis .
the digital spectrophotometer measures the tooth shade based on two 
 methods : by grouping shades according to the 16 shade tabs in the guide ( vitapan 
 classical , vita zahnfabrik ) , which were numbered from 1 ( highest value - b1 ) to 16 
 ( lowest value - c4 ) ( figure 1 ) and , the ciel*a*b * color system , 
 which was defined by the international commission on illumination . in the cielab 
 color system ,
the l * axis represents the value ( lightness or darkness ) , the a * axis 
 is a measure of redness ( positive a * ) or greenness ( negative a * ) , and the b * axis is 
 a measure of yellowness ( positive b * ) or blueness ( negative b * ) . the difference 
 between the color coordinates was calculated as : 
 e*=[(l*)+(a*)+(b * ) ] .
color scores by sequence of colors in the shade guide ( from lighter to darker 
 colors ) the same protocol used to determine the tooth shade at baseline ( t0 ) was used in all 
 other periods evaluated : after 1 week of treatment ( t1 ) and 1 week post - bleaching 
 ( t2 ) .
all teeth shades measurements were performed objectively by an experienced 
 examiner , which was blinded to the agent concentration . the same examiner responsible for the staining procedure carried out all bleaching 
 treatments .
the coffee - stained samples were randomly allocated into four groups 
 ( n=15 ) , according to the bleaching agent concentration : distilled water ( control 
 group ) , 10% ( cp10 ) , 16% ( cp16 ) or 37% ( cp37 ) carbamide peroxide .
the manufacturer and 
 chemical composition of each bleaching agent used in this study are listed in figure 2 .
description of the bleaching agents used in this study the control group was stored in distilled water at 37c , which was daily renewed 
 during 2 weeks .
samples of cp10 and cp16 groups were covered with 0.2 ml of the 
 respective bleaching gels , which were applied on enamel surface for 4 h/ day during 2 
 weeks .
samples of cp37 group were covered with 0.2 ml of the bleaching gel for 20 
 min .
the gel was light activated by two 40-s applications ( led radii , sdi , bayswater , 
 victoria , australia ) spaced by 10-min intervals .
this cycle was repeated at 3 sessions 
 with 5 days of interval between them . after bleaching treatments ,
the gels were 
 carefully removed with sterile gauze and tap water and then the samples were stored 
 in distilled water at 37c until the next application .
tooth shade means were analyzed statistically by kruskal - wallis and friedman 's tests 
 and color parameters were analyzed by two - way analysis of variance ( anova ) and 
 tukey 's test .
sixty recently extracted bovine incisors without cracks and surface defects ( 40x 
 magnification ) were used in this study .
the roots were cut with a water - cooled 
 low - speed diamond saw ( isomet 1000 , buehler , lake bluff , il , usa ) and , the pulp 
 tissue was removed .
then , teeth were stored in 0.5% chloramine - t solution for one 
 week .
after disinfection , each crown was sectioned in both buccolingual and mesiodistal 
 directions to obtain one enamel - dentin slab ( 6x6x3 mm ) , which were measured with a 
 digital caliper ( dc 500 , mitutoyo , suzano , sp , brazil ) .
each slab was individually 
 embedded in self - curing acrylic resin in a ring model and then , enamel surfaces were 
 flattened with 600- and 1200-grit silicon - carbide paper under water cooling to create 
 a standardized buccal enamel surface .
samples were washed and stored in distilled 
 water at 37c for 1 week until starting the experimental procedures .
in an attempt to standardize the tooth shade , an artificial staining of the specimens 
 was carried out following the method proposed by sulieman , et al .
enamel surfaces were etched 
 with 35% phosphoric acid gel for 60 s and rinsed with distilled water for 30 s. 
 specimens were immersed daily over a 1-week period in a standardized coffee solution 
 at room temperature ( 252c ) in screw capped plastic universal containers .
the daily 
 renewed coffee solution was produced by boiling 12 g of coffee ( melitta , avar , sp , 
 brazil ) in 200 ml of distilled water for 5 min followed by filtering in a coffee 
 maker ( melitta ) .
after the immersion period , the specimens were washed and stored in 
 distilled water at room temperature .
tooth shades of the stained specimens were assessed with a digital spectrophotometer 
 ( vita easyshade , vita zahnfabrik , bad sackingen , germany ) after 1 , 3 and 7 days of 
 immersion in coffee solution ( baseline ) .
three measurements were done with the active 
 point of the spectrophotometer in the center of each sample .
the instrument 
 automatically averaged the three readings for each sample , which were use for overall 
 data analysis .
the digital spectrophotometer measures the tooth shade based on two 
 methods : by grouping shades according to the 16 shade tabs in the guide ( vitapan 
 classical , vita zahnfabrik ) , which were numbered from 1 ( highest value - b1 ) to 16 
 ( lowest value - c4 ) ( figure 1 ) and , the ciel*a*b * color system , 
 which was defined by the international commission on illumination . in the cielab 
 color system ,
the l * axis represents the value ( lightness or darkness ) , the a * axis 
 is a measure of redness ( positive a * ) or greenness ( negative a * ) , and the b * axis is 
 a measure of yellowness ( positive b * ) or blueness ( negative b * ) .
the difference 
 between the color coordinates was calculated as : 
 e*=[(l*)+(a*)+(b * ) ] .
color scores by sequence of colors in the shade guide ( from lighter to darker 
 colors ) the same protocol used to determine the tooth shade at baseline ( t0 ) was used in all 
 other periods evaluated : after 1 week of treatment ( t1 ) and 1 week post - bleaching 
 ( t2 ) .
all teeth shades measurements were performed objectively by an experienced 
 examiner , which was blinded to the agent concentration .
the coffee - stained samples were randomly allocated into four groups 
 ( n=15 ) , according to the bleaching agent concentration : distilled water ( control 
 group ) , 10% ( cp10 ) , 16% ( cp16 ) or 37% ( cp37 ) carbamide peroxide .
the manufacturer and 
 chemical composition of each bleaching agent used in this study are listed in figure 2 .
description of the bleaching agents used in this study the control group was stored in distilled water at 37c , which was daily renewed 
 during 2 weeks .
samples of cp10 and cp16 groups were covered with 0.2 ml of the 
 respective bleaching gels , which were applied on enamel surface for 4 h/ day during 2 
 weeks .
samples of cp37 group were covered with 0.2 ml of the bleaching gel for 20 
 min .
the gel was light activated by two 40-s applications ( led radii , sdi , bayswater , 
 victoria , australia ) spaced by 10-min intervals .
this cycle was repeated at 3 sessions 
 with 5 days of interval between them . after bleaching treatments ,
the gels were 
 carefully removed with sterile gauze and tap water and then the samples were stored 
 in distilled water at 37c until the next application .
tooth shade means were analyzed statistically by kruskal - wallis and friedman 's tests 
 and color parameters were analyzed by two - way analysis of variance ( anova ) and 
 tukey 's test .
tooth shade means and tooth color parameters ( l * , a * , b * , e * ) for the treatment 
 groups at the different evaluation periods are presented in tables 1 and 2 , 
 respectively . at t1 and t2 evaluations , cp10 , cp16 and cp37 groups showed a 
 statistically significant decrease in tooth shade means when compared with baseline and 
 control group ( p<0.001 ) . at t1 ,
the group treated with cp37 presented lower tooth 
 shade means than the other bleaching treatments tested in the same period ( p<0.01 ) . 

however , the tooth shade means resulting from the 1-week post - bleaching procedures ( t2 ) 
 were not statistically different between cp10 , cp16 and cp37 groups ( p>0.05 ) .
tooth shade means and standard deviations ( sd ) for the different treatment groups 
 according to the evaluation periods different capital letters indicate statistically significant differences only 
 among the evaluation periods ( p<0.001 ) .
different lowercase letters indicate 
 statistically significant differences only among the treatment groups 
 ( p<0.001 ) .
tooth color parameter means and standard deviations ( sd ) for the different 
 treatment groups according to the evaluation periods at baseline , l * , a * , b * means were shown in absolute values .
different capital 
 letters indicate statistical significant differences only between evaluation 
 periods ( p<0.05 ) .
different lowercase letters indicate statistical 
 significant differences only between treatment groups ( p<0.05 ) . considering tooth color parameters at t1 and t2 , all treatment groups showed an 
 increasing of the lightness ( l * ) , a decreasing of the redness ( a * ) and yellowness 
 ( b * ) compared with baseline ( p<0.001 ) . however , the increasing of the lightness 
 ( l * ) in the group treated with cp37 was lower than cp10 and cp16 at both evaluations 
 periods ( p<0.001 ) .
although a * values decreased for all treatment groups , at t2 , 
 there was no statistically difference among cp10 , cp16 , cp37 and control ( p>0.06 ) . 

the decrease of b * values in cp37 was significantly lower than in the other treatment 
 groups ( p<0.001 ) . despite e * values , there was no difference among the treatment 
 groups at the end of the treatment ( p>0.1 ) .
previous studies have suggested that the two main factors that determine the tooth 
 whitening efficacy are the peroxide concentration and its time of application .
there is a belief that the increase of the bleaching agent 
 concentration can produce a higher and faster whitening effect compared with whitening 
 products with low - concentration bleaching agents .
however , this study showed that after 
 the first week of treatment , 10 and 16% carbamide peroxide concentrations were 
 significantly more effective than the 37% concentration . from this study
, it appears 
 that the factor contact time of bleaching agent was more important than the 
 concentration of gels .
although this result might seem surprising , similar effects in 
 tooth shade improvement were achieved with longer treatment times , independently of the 
 peroxide concentration used .
a review that compared 9 clinical 
 trials and the effectiveness of 25 tooth whitening products showed that tooth whitening 
 is more effective when the bleaching agent is placed in trays .
as the present in vitro study followed the manufacturers ' 
 recommendations in order to evaluate the effectiveness of carbamide peroxide - based gels 
 available for in - office bleaching ( 37% ) or home - use ( 10 and 16% ) , the mode and duration 
 of applications were different among the groups .
certainly , these different 
 characteristics of application for each bleaching agent might have influenced the 
 outcome of tooth whitening . a clinical trial that evaluated the effectiveness of 8 in - office 
 tooth whitening systems verified that the contact time between bleaching gel and tooth 
 surface is an important factor that can improve the outcome of the bleaching 
 treatment .
the necessity of light - activation of peroxide - based gels has already been questioned by 
 some authors . even though the use of light in 
 bleaching procedures has been considered unnecessary because of the presence 
 of other agents ( i.e. , carbopol ) that assure the catalyze of peroxides , 
 light - activation of 37% carbamide peroxide
was performed in our study to accelerate the 
 whitening process , following the manufacturer 's instructions . despite the subjectivity of visual tooth color assessment ,
it has been well documented 
 that the tooth color changes can be successfully assessed using both shade guides and 
 digital spectrophotometry .
nevertheless , when employing an 
 objective method for measuring the tooth color changes , it is also possible to evaluate 
 color parameters ( l * , a * and b * ) based on the cielab system , an three - dimensional color 
 space . according to 
 spectrophotometer data , the whitening improvement occurs mainly by increasing the 
 lightness ( higher l * ) and reducing the redness ( lower a * ) and yellowness ( lower 
 b * ) . at 1 week post - bleaching
, 
 all treatment groups showed an increase of the l * and a decrease of the a * and b * color 
 parameters .
there are clearly limitations in comparing in vitro results 
 with clinical studies once that changes in l * , a * and b * need to be interpreted with 
 caution . nevertheless , the color parameters data of this in vitro study 
 is in agreement with the results of clinical trials , indicating that lower 
 concentrations of tooth bleaching gels are as effective as higher 
 concentrations .
the control group showed an increase of the lightness ( l * ) and also a decrease of the 
 yellowness ( b * ) and redness ( a * ) during water storage .
these changes in the parameters 
 are probably due to the removal of tooth surface pigments , as a consequence of the 
 coffee staining procedure .
the overall color change ( e * ) values are often used in tooth whitening studies in order 
 to indicate the perceptible tooth color changes after treatment . at 1 week post - bleaching
clinically , such finding could demonstrate that the lower 
 concentration of peroxide - based gels would achieve the same perceptible results as the 
 37% carbamide peroxide , frequently used for in - office tooth bleaching .
besides , previous 
 studies have stated that color reversal after bleaching treatment occurs quicker for 
 in - office products than home tray - based bleaching agents .
considering that all carbamide peroxide gels evaluated in this study resulted in a 
 similar tooth shade improvement , it could be suggested the use of low carbamide peroxide 
 concentrations for vital tooth bleaching , in accordance with the american dental 
 association guidelines , which only assures the safety and efficacy of bleaching 
 treatment using trays with 10%1% carbamide peroxide application .
the null hypothesis for this study was accepted as the high - concentration carbamide 
 peroxide produced similar tooth color improvement than the bleaching agents containing 
 low carbamide peroxide concentration .
further studies and clinical trials are needed to 
 determine if the same color improvement is obtained with in - office or home - use carbamide 
 peroxide bleaching agents .
within the limitations of this study , it may be concluded that at - home ( 10 or 16% 
 carbamide peroxide ) or in - office ( 37% carbamide peroxide ) bleaching agents produced a 
 similar color improvement after the end of the treatment .
tooth bleaching with high or low concentrated carbamide peroxide agents were 
 similarly effective after treatment period .
practitioners must be informed about 
 bleaching techniques in order to provide an evidence - based choice for their 
 patients .
tooth bleaching with high or low concentrated carbamide peroxide agents were 
 similarly effective after treatment period .
practitioners must be informed about 
 bleaching techniques in order to provide an evidence - based choice for their 
 patients .
 objectives this in vitro study evaluated the effectiveness of three 
 carbamide peroxide concentrations used for tooth bleaching treatments . 
 material and methods sixty bovine dental slabs ( 6x6x3 mm ) were obtained , sequentially polished , 
 submitted to artificial staining ( baseline ) and randomized into four groups 
 ( n=15 ) , according to the bleaching agent concentration : distilled water ( control ) , 
 10% ( cp10 ) , 16% ( cp16 ) or 37% ( cp37 ) carbamide peroxide . 
 cp10 and cp16 were 
 covered with 0.2 ml of the respective bleaching gels , which were applied on enamel 
 surface for 4 h / day during two weeks . 
 samples of cp37 were covered with 0.2 ml of 
 the bleaching gel for 20 min . 
 the gel was light activated by two 40-s applications 
 spaced by 10-min intervals . 
 the gel was renewed and applied 3 times per clinical 
 session . 
 this cycle was repeated at 3 sessions with 5 days of interval between 
 them . 
 tooth shade evaluations were done with a digital spectrophotometer at t0 
 ( baseline ) , t1 ( after 1-week of treatment ) and t2 ( 1-week post - bleaching ) . 
 tooth 
 shade means were statistically analyzed by kruskal - wallis and friedman 's tests and 
 color parameters were analyzed by two - way anova and tukey 's test ( p<0.05 ) . 
 results at t1 and t2 evaluations , tooth shade was significantly lighter than at baseline 
 for all treatment groups , considering the color parameters l * , a * , b * , e * 
 ( p<0.001 ) or tooth shade means ( p<0.001 ) . 
 cp37 group showed lower shade mean 
 change than cp10 and cp16 at t1 ( p<0.01 ) , but this difference was not 
 statistically significant at t2 ( p>0.05 ) . 
 conclusions one week after the end of the treatment , all carbamide peroxide concentrations 
 tested produced similar tooth color improvement . 
INTRODUCTION
MATERIAL AND METHODS
Specimen preparation
Staining procedure
Tooth shade measurement
Bleaching procedure
Statistical analysis
RESULTS
DISCUSSION
CONCLUSIONS
Clinical relevance