UNIVERSIDADE ESTADUAL DE CAMPINAS FACULDADE DE ODONTOLOGIA DE PIRACICABA
THAYLA HELLEN NUNES GOUVEIA
EFEITO DO PERÓXIDO DE CARBAMIDA A 10% ASSOCIADO AO
POLÍMERO BIOADESIVO ARISTOFLEX NAS PROPRIEDADES
FÍSICAS E QUÍMICAS DO ESMALTE DENTAL
EFFECT OF 10% CARBAMIDE PEROXIDE ASSOCIATED WITH A
BIOADHESIVE POLYMER ARISTOFLEX ON THE PHYSICAL AND
CHEMICAL PROPERTIES OF DENTAL ENAMEL
PIRACICABA 2018
THAYLA HELLEN NUNES GOUVEIA
EFEITO DO PERÓXIDO DE CARBAMIDA A 10% ASSOCIADO AO POLÍMERO BIOADESIVO ARISTOFLEX NAS PROPRIEDADES FÍSICAS E QUÍMICAS DO
ESMALTE DENTAL
EFFECT OF 10% CARBAMIDE PEROXIDE ASSOCIATED WITH A
BIOADHESIVE POLYMER ARISTOFLEX ON THE PHYSICAL AND CHEMICAL PROPERTIES OF DENTAL ENAMEL
Tese apresentada à Faculdade de Odontologia de Piracicaba da Universidade Estadual de Campinas como parte dos requisitos exigidos para obtenção do título de Doutora em Clínica Odontológica, na Área de Dentística.
Thesis presented to the Piracicaba Dental School of the University of Campinas in partial fulfillment of the requirements for the degree of Doctor in Clinical Dentistry, in Restorative Dentistry area.
Orientadora: Profª. Drª. Débora Alves Nunes Leite Lima
PIRACICABA 2018 ESTE EXEMPLAR CORRESPONDE À VERSÃO FINAL DA TESE DEFENDIDA PELA ALUNA
THAYLA HELLEN NUNES GOUVEIA,
ORIENTADA PELA PROFª. DRª. DÉBORA ALVES NUNES LEITE LIMA.
Agência(s) de fomento e nº(s) de processo(s): Não se aplica.
Ficha catalográfica
Universidade Estadual de Campinas
Biblioteca da Faculdade de Odontologia de Piracicaba Marilene Girello - CRB 8/6159
Gouveia, Thayla Hellen Nunes,
G745e GouEfeito do peróxido de carbamida a 10% associado ao polímero bioadesivo Aristoflex nas propriedades físicas e químicas do esmalte dental / Thayla Hellen Nunes Gouveia. – Piracicaba, SP : [s.n.], 2018.
GouOrientador: Débora Alves Nunes Leite Lima.
GouTese (doutorado) – Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba.
Gou1. Dentes - Clareamento. 2. Análise espectral. 3. Química analítica. 4. Esmalte dentário. I. Lima, Débora Alves Nunes Leite, 1978-. II. Universidade Estadual de Campinas. Faculdade de Odontologia de Piracicaba. III. Título.
Informações para Biblioteca Digital
Título em outro idioma: Effect of 10% carbamide peroxide associated with a bioadhesive
polymer Aristoflex on the physical and chemical properties of dental enamel
Palavras-chave em inglês:
Teeth - Bleaching Spectrum analysis Chemistry, analytic Dental enamel
Área de concentração: Dentística
Titulação: Doutora em Clínica Odontológica Banca examinadora:
Débora Alves Nunes Leite Lima [Orientador] Maria Denise Rodrigues de Moraes Bezerra Maria Beatriz Freitas D'Arce
Giselle Maria Marchi Baron Luís Roberto Marcondes Martins
Data de defesa: 31-08-2018
Programa de Pós-Graduação: Clínica Odontológica
UNIVERSIDADE ESTADUAL DE CAMPINAS Faculdade de Odontologia de Piracicaba
A Comissão Julgadora dos trabalhos de Defesa de Tese de Doutorado, em sessão pública realizada em 31 de Agosto de 2018, considerou a candidata THAYLA HELLEN NUNES GOUVEIA aprovada.
PROFª. DRª. DÉBORA ALVES NUNES LEITE LIMA PROFª. DRª. MARIA DENISE RODRIGUES DE MORAES BEZERRA
PROFª. DRª. MARIA BEATRIZ FREITAS D'ARCE PROFª. DRª. GISELLE MARIA MARCHI BARON PROF. DR. LUÍS ROBERTO MARCONDES MARTINS
A Ata da defesa com as respectivas assinaturas dos membros encontra-se no processo de vida acadêmica do aluno.
DEDICATÓRIA
Esta trabalho é dedicado aos amores de minha vida: Meus pais, Francisco e Margarida, por serem apoio e fortaleza. Meus irmãos, Thaciano, Thaís e Talita, pela cumplicidade e amor. Meu noivo, Manoel Costa Neto, pela paciência e segurança. Sem vocês junto a mim, jamais teria conseguido!
AGRADECIMENTO ESPECIAL
À minha orientadora, Profa. Dra. Débora Alves Nunes Leite Lima, por me permitir viver o Doutorado. Por acreditar em mim, como uma mãe que tudo faz pelo seu filho, a senhora me escolheu novamente e lutou por mim. Por toda sua confiança e incentivo, MUITO OBRIGADA!
AGRADECIMENTOS
À Deus, pelas bênçãos concedidas, por todas experiencias e por nunca ter me abandonado. À Universidade Estadual de Campinas – UNICAMP, na pessoa de seu Magnífico Reitor Prof. Dr. Marcelo Knobel.
À Faculdade de Odontologia de Piracicaba - FOP, na pessoa de seu diretor, Prof. Dr. Guilherme Elias Pessanha Henriques e diretor associado Prof. Dr. Francisco Haiter Neto. À Profa. Dra. Karina Gonzales Silvério Ruiz, Coordenadora Geral do Programa de Pós-Graduação da FOP/UNICAMP.
Ao Prof. Dr. Valentim Adelino Ricardo Barão Coordenador do Curso de Pós-Graduação em Clínica Odontológica da FOP/UNICAMP.
Aos funcionários da FOP-UNICAMP sempre dispostos a ajudar e solucionar os problemas surgidos.
Aos Professores do Departamento de Odontologia Restauradora, Área de Dentística- FOP/UNICAMP, Prof. Dr. Luís Alexandre Maffei Sartini Paulillo; Prof. Dr. Luís Roberto Marcondes Martins; Prof. Dr. Flávio Henrique Baggio Aguiar; Profa. Dra. Giselle Maria Marchi Baron; Prof. Dr. Marcello Gianinni pelos ensinamentos concedidos durante a pós-Graduação.
Aos Professores da banca do meu exame de qualificação, Prof Dr. Flávio Henrique Baggio Aguiar; Prof. Dr. Waldemir Francisco Vieria Junior e Profa. Dra. Lúcia Trazzi Prieto, pela atenção e excelentes contribuições nesse estudo.
À minha turma de doutorado do Departamento de Odontologia Restauradora, Área de Dentística- FOP/UNICAMP por toda convivência, ensinamentos e amizade, Jéssica, Cristiane e Michelle, Waldemir, Mary, Suelem, Diogo, Marília e Isabel. Muito Obrigada. À todos meus colegas de pós-graduação mestrado e doutorado do Departamento de Odontologia Restauradora, Área de Dentística- FOP/UNICAMP, Danielle, Bruna, Carol, Mariana, Rodrigo, Renata, Josué e Laura. Muito Obrigada.
Ao amigo Bruno Vilela do Departamento de Ciências Fisiológicas- Área de Farmacologia, Anestesiologia e Terapêutica- FOP/UNICAMP, por toda ajuda durante o doutorado.
Ao amigo Ándre Condeles do Departamento de Química Analítica – Curso de Química da USP – Ribeirão preto, pela amizade e ajuda nos testes químicos durante meu Doutorado. À Danielle Ferreira, Aluna de Mestrado do Departamento de Odontologia Restauradora, Área de Dentística- FOP/UNICAMP, presente de Deus nessa reta final do Doutorado, por cuidar de mim e pela presença sempre disposta. Estarei sempre na torcida por suas conquistas.
As minhas amigas-irmãs, Jéssica Theobaldo e Michelle Lima, a vocês meu carinho e respeito. Obrigada por cuidarem de mim. Desejo que sejam muito felizes em todas as escolhas que optarem. Nossa amizada será eterna. Da pós-graduação para vida!
À todos os meus familiares, tios, primos e primas, presentes em todos os momentos da minha vida, que, apesar da distância física, estão sempre ao meu lado.
A minha faculdade querida, hoje Centro Universitário Católica de Quixadá - UNICATÓLICA, por ser meu amor primeiro na Odontologia.
À minha professora da Graduacão, Profa. Dra. Maria Denise Moraes, minha eterna orientadora, pela escolha de ser verdadeiramente mestre, pelo amor a profissão e pela amizade.
Aos amigos professores, Prof. Dr. Filipe Martins, Prof. Dr. Juscelino Freitas e Prof. Dr. José Filho, pelo incentivo e torcida, da Cátolica para o mundo, chegou minha vez migos!
À minha eterna dupla, Rafela Costa, por todo carinho e amizade. Seguimos conquistando nossos sonhos. Jaja Profa Ma Rafinha!
À Talita Malini, um prazer sua companhia e amizade, obrigada pelos anos de convivência na casa do cavalo. Desejo as melhores conquista para ti.
À Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), pela concessão de bolsa em determinado período deste Doutorado.
À Drogal Manipulações, na pessoa da farmacêutica Rafaela, por todo atendimento e colaboração durante toda minha pós graduação.
E a todos que, direta ou indiretamente, contribuíram para a realização deste trabalho, o meu sincero agradecimento.
“...Desistir dos sonhos é abrir mão da felicidade porque quem não persegue seus objetivos esta condenado a fracassar 100 vezes...”
RESUMO
Objetivo: Analisar in vitro o efeito do polímero bioadesivo, co-polímero do ácido sulfônico acriloildimetiltaurato e vinilpirrolidona neutralizado (Aristoflex® AVC ), no esmalte dental clareado por meio das propriedades de cor, microdureza, rugosidade, concentração do conteúdo mineral e morfologia de superfície. Métodos: Oitenta e quatro blocos dentais bovinos (4 × 4 × 2,5 mm) foram divididos em 7 grupos (n=12): PC 10% com carbopol – produto comercial FGM - Whiteness Perfect (WP); PC 10% com Aristoflex (A + CP 10%); PC 10% com carbopol - manipulado (C + CP 10%); gel de Aristoflex (A); gel de carbopol (C); Peróxido de Carbamida na concentração de 10% (CP 10%) e saliva artificial (controle). Os géis foram preparados e sua viscosidade e pH foram analisados por um reômetro modular e pHmetro digital respectivamente. O tratamento clareador foi realizado, diariamente, por 4 horas, em um período de 14 dias. Entre as sessões, as amostras permaneceram em solução remineralizadora. As análises de cor, rugosidade e microdureza foram realizadas antes e após os tratamentos. A cor foi obtida pelo método CIE Lab (ΔE, ΔL*, Δa* e Δb*) através de espectrofotômetro de reflectância; o perfil da rugosidade de superfície (Ra) foi avaliado por um perfilômetro de contato; e o microdurômetro codificou a microdureza Knoop superficial. As concentrações analíticas (mg/L) dos minerais Ca e P foram realizadas no 10,30,70 e 140 dia de tratamento, por meio do espectrômetro de emissão óptica com plasma acoplado indutivamente (ICP OES). Para este teste foram considerados os seguintes grupos: WP; A + CP 10%; C + CP 10%; A; C and CP 10% (controle sem espessante). A morfologia foi qualificada por microscopia eletrônica de varredura (MEV). Os dados de ΔE, ΔL* e Δb* foram submetidos a ANOVA e teste de Tukey. Os dados de ∆a* foram analisados por modelos lineares generalizados. A perfilometria, a microdureza e os dados de Ca e P foram analisados por modelos mistos para medidas repetidas no tempo (PROC MIXED) e teste de Tukey-Kramer (a = 0,05). Resultados: Todos os grupos que continham o PC 10%, independente do espessante utilizado, apresentaram significativa mudança de cor para ΔE, ΔL*, Δa* e Δb* comparado ao controle. O grupo A + CP 10% manteve os valores iniciais de rugosidade e microdureza após o clareamento. Os grupos que continham somente espessante e o grupo WP, apresentaram os menores valores de microdureza e maiores valores de rugosidade, diferindo estatisticamente do controle. Maiores quantidades de Ca e P foram encontradas no 10 dia tratamento para o grupo WP comparado aos grupos A + CP 10% e C + CP 10% mas, no 140 dia, nenhuma diferença estatística foi observada entre os grupos. Alterações morfológicas foram observadas em todos os grupos avaliados pelo MEV, exceto no grupo tratado com o A + CP 10%. Conclusão: O polímero bioadesivo Aristoflex® AVC associado ao peróxido de carbamida a 10% foi efetivo no tratamento clareador sem alterar as propriedades fisícas e químicas avaliadas. Além disso, este gel experimental causou menor perda mineral quando comparado ao grupo PC 10% com carbopol (produto comercial).
Palavras-Chaves: Clareamento dental. Peróxido de carbamida. Análise química. Esmalte. Microdureza. Perfilometria. Espectroscopia.
ABSTRACT
Objective: To analyze in vitro the effect of bioadhesive polymer, Ammonium Acryloyldimethyltaurate/VP Copolymer (Aristoflex® AVC), on enamel properties, color, microhardness, roughness, mineral content and morphology surface. Methods: Seven groups (n=12) of enamel blocks from bovine incisors (4 × 4 × 2.5 mm) were divided in: CP 10% with carbopol – commercial product FGM - Whiteness Perfect® (WP); CP 10% with Aristoflex (A + CP 10%); CP 10% with carbopol - manipulated (C + CP 10%); Aristoflex thickener (A); carbopol thickener (C); Carbamide peroxide 10% (CP 10%) and artificial saliva (control). The bleaching/thickener gels were prepared and Rheological characterizatio and pH were analyzed by a Modular Compact Rheometer and digital pHmeterrespectively. The bleaching treatment was performed daily for 4 hours during 14 days, between sessions the samples were kept in remineralizing solution. The color, roughness and microhardness analyzes were performed before and after the bleaching/thickener treatment. The color was obtained by CIE Lab method (ΔE, ΔL*, Δa* and Δb*) through a spectrophotometer; Surface roughness profilometry (Ra) was evaluated by a contact profilometer; The knoop microhardness was measured by microdurometer. The analytical concentrations (mg / L) of Ca and P minerals were performed only at 1st, 3rt, 7th and 14th day of treatment, by means of Inductively Coupled Plasma Optical Emission Spectrometry (ICP OES) for the groups WP, A + CP 10%, C + CP 10%, A, C and CP 10% (control without thickener). The morphology was qualified by scanning electron microscopy (SEM). The ΔE, ΔL and Δb data were submitted to ANOVA and Tukey's test. The Δa data were analyzed by generalized linear models. The roughness profile, microhardness and Ca and P data were evaluated by repeated measurements (PROC MIXED) and Tukey-Kramer test (a = 0.05). Results: All groups with PC 10%, regardless of the polymer used, differ statistically were found to ΔE, ΔL*, Δa* e Δb* compared to the control group. The group A + CP 10% kept initial values of roughness and microhardness after bleaching. The groups only thickener and the WP group presented the lowest values of microhardness and higher values of roughness, with statistical differences of the control. Higher amounts of Ca and P were found in the 1st day treatment in the WP group compared to the groups A + CP 10% and C + CP 10% but in the day 14th no statistical difference was observed between the groups. Morphological alterations were observed in all groups evaluated by SEM, except in the group treated with A + CP 10% compared to the control. Conclusion: The 10% CP solution associated with the bioadhesive polymer Aristoflex® AVC was effective on the bleaching treatment without changing the physical properties of the dental enamel. Also, this experimental gel caused less mineral loss than the group of 10% Carbamide peroxide 10% with carbopol (commercial product).
Keywords: Bleaching treatment. Carbamide peroxide. Chemical analysis. Enamel. Microhardness. Profilometry. Spectroscopy.
SUMÁRIO
1 INTRODUÇÃO ... 13
2 ARTIGO: Effect of Ammonium Acryloyldimethyltaurate/VP Copolymer on the physical and chemical properties of bleached dental enamel. ... 17
3 CONCLUSÃO ... 43
REFERÊNCIAS ... 44
Apêndice 1 - Metodologia detalhada ... 46
ANEXOS ... 62
Anexo 1 - Verificação de originalidade e prevenção de plágio por meio do software Turnitin ... 62
1 INTRODUÇÃO
Na construção de um sorriso esteticamente agradável, a uniformidade na cor dos dentes é um fator de extrema relevância. Assim, procedimentos clínicos que possibilitem de forma menos invasiva a recuperação da estética dental têm sido cada vez mais valorizados (Gupta et al., 2017). Dentro deste contexto, o clareamento dental é contemporaneamente eleito como um dos tratamentos mais procurado para a melhoria da aparência estética, sendo conceituado como um procedimento conservador e eficiente quando bem indicado (Corcodel
et al., 2017). Por outro lado, este procedimento pode ocasionar algumas alterações na
morfologia dos tecidos dentais mineralizados, influenciando negativamente as propriedades físicas do esmalte dental, como o aumento da rugosidade e a diminuição da microdureza (Tanaka et al., 2010; Sa Y et al., 2013; Ferraz et al., 2018).
O tratamento clareador de dentes com vitalidade pulpar pode ser realizado por meio das principais técnicas: a técnica de clareamento de consultório, realizada pelo cirurgião-dentista em ambiente clínico; ou de clareamento caseiro, em que o paciente realiza o procedimento fora do ambiente clínico, através da utilização de uma moldeira personalizada, confeccionada previamente pelo cirurgião-dentista que também fará a orientação e supervisão semanalmente do tratamento. Em ambas as técnicas, o processo clareador das estruturas dentais alteradas por manchas de natureza intrínseca ou extrínseca ocorre através da aplicação de géis à base de peróxido de hidrogênio e/ou do seu precursor, o peróxido de carbamida, em diferentes concentrações que são utilizados variando o tempo de aplicação (Eimar et al, 2012).
Quimicamente, o processo clareador ocorre por meio de uma reação de oxidorredução. O peróxido de carbamida (H2NCONH2H2O2), quando em contato com a saliva ou água, dissocia-se em peróxido de hidrogênio (H2O2) e uréia (H2NCONH2). O peróxido de hidrogênio decompõe-se de diferentes maneiras, formando íons, moléculas de oxigênio reativas, instáveis que se transformam em oxigênio (O*), ânion superóxido e/ou ânion de peróxido de hidrogênio) ou radicais livres (Hidroxil - 2HO*e Peridroxil - HO2*). Posteriormente, a uréia degrada-se em amônia (2NH3) e em dióxido de carbono (CO2) (Fonseca, 2008; Cavalli et al., 2004; Dahl e Pallesn, 2003).
Os radicais livres gerados nesta reação clivam os pigmentos que escurecem o dente. Esses pigmentos são cadeias de alto peso molecular e que, através de reações de oxirredução, são transformadas em moléculas menores, o suficiente para não serem detectadas no espectro visível e ainda podem ser eliminadas da estrutura dental pelo processo de difusão,
tornando assim, o dente mais claro (Plotino et al., 2008; Toledano et al., 2011; Ubaldini et al., 2013; Kwon SR, & Wertz PW, 2015). Entretanto, os radicais de peróxido de hidrogênio não agem de forma específica, são extremamente instáveis e podem reagir não somente com as duplas ligações de carbono contida nas moléculas cromógenas que escurecem os dentes, mas também com a matriz orgânica e inorgânica do esmalte e dentina (Basting et al., 2005; Kawamoto & Tsujimoto, 2004). Assim, os efeitos indesejáveis, como alterações da morfologia superficial, redução da microdureza e perda de minerais, podem ser observados após o tratamento clareador (Basting et al., 2005; Basting 2001; Sa Y et al., 2013; Tanaka et
al., 2010). Contudo, estudos vêm mostrando que estas alterações, sobretudo a redução da
microdureza do esmalte e da dentina, podem não estar relacionadas somente à ação dos radicais livres de peróxido, mas também devido a outros constituintes presentes na formulação do produto clareador, como os espessantes (Basting et al., 2005; Oliveira et al., 2007).
Na formulação dos produtos para o clareamento dental da técnica caseira, geralmente encontram-se os seguintes constituintes: o agente clareador, que é o conteúdo ativo, peróxido de hidrogênio ou precurssor peróxido de carbamida, e os demais componentes constituindo a parte inativa do produto, sendo eles: os umectantes, que ajudam a dissolver os componentes e auxiliam na difusão da parte ativa, exemplo, o propileno glicol e a glicerina; os conservantes, como o propilparabeno, benzoato de sódio, que são usados em pequenas quantidades na formulação, pois podem acelerar a degradação do peróxido de hidrogênio pela liberação de alguns metais como o ferro, cobre e magnésio, impedem também o crescimento bacteriano (Alqahtani, 2014); os estabilizantes, que são utilizados para retardar a degradação dos peróxidos ao longo do tempo e, com isso, aumentar o tempo de validade dos produtos, sem afetar a sua eficiência como o edeato dissódico de cálcio (EDTA) ou ácido dietileno-triamino-pentaacético (DTPA); agentes tamponantes, utilizados para permitir um pH do produto o mais neutro possível, a fim de tentar impedir uma desmineralização do tecido dental como o aminometil propano (AMP -95) (Benbachir et al., 2008); agentes aromatizantes, incluídos para permitir uma utilização agradável, como a menta ou hortelã e, ainda, adoçante tipo sacarina; agentes dessensibilizantes, com a função de diminuir uma possível sensibilidade dentária induzida pelo tratamento, sendo os mais comumente encontrados são o fluoreto de sódio e o nitrato de potássio (Silva et al., 2012); e um ou mais agente de viscosidade, que são polímeros responsáveis pelo espessamento da solução, os quais atuam dando a consistência de gel ao produto e agem como modulador da reação química de oxidação e redução (Basting et
al., 2005; Oliveira et al., 2007; Alqahtani 2014). São contemporaneamente investigados e
denomindados como polímeros bioadesivos, devido à capacidade de interagir, por meio de ligações iônicas, com a estrutura dental (Ávila et al., 2017).
Um dos espessantes mais utilizados na composição dos géis clareadores caseiros é o polímero carboxipolimetileno denominado, carbopol (Basting et al., 2005; Oliveira et al., 2007; Soares et al., 2016). Este espessante possui natureza ácida, sendo derivado de um ácido carboxílico. Mas, para uso intrabucal como espessante em géis clareadores ou soluções de saliva artificial, deve ser tamponado a um pH neutro para ser um agente inerte (Pozzobon et al., 2003). Entretanto, esse espessante tem sido descrito na literatura como capaz de causar diminuição na microdureza do esmalte dental. Basting et al., 2005 relataram que o carbopol inibiu completamente o crescimento cristalino do esmalte, após o tratamento clareador, em todos os grupos que o continha como espessante, mostrando uma significativa redução nos valores de microdureza. Segundo estes autores, isso ocorreu devido à sua elevada capacidade de ligação com o cálcio, o que consequentemente impossibilitou uma provável remineralização pelos minerais presente na saliva artificial. Além disso, os autores acreditam que este espessante não apresente uma estabilidade de pH durante o tempo do tratamento clareador e que, em contato com o meio aquoso (saliva), se torne ainda mais ácido, mesmo tendo sido tamponado previamente, e isso pode contribuir ainda mais com o processo de desmineralização dental.
Desta forma, este estudo propõe avaliar o efeito de um gel clareador com o espessante à base do co-Polímero do ácido sulfônico acriloildimetiltaurato e vinilpirrolidona, denominado Aristoflex® AVC, o qual apresenta características opostas ao carbopol. O Aristoflex é um polímero sintético pré-neutralizado, catiônico, que permite a formação de géis cristalinos com boa consistência. As características mais importantes deste espessante é não ser dependente de pH e, por isso, mostra uma alta estabilidade em pHs ácidos (Lindsey et al., 2014). Na indústria farmacêutica é utilizado com estabilizante e agente de consistência para emulsões leves. Na indústria química, tem sido empregado na confecção de dentifrícos, nas concentrações de até 1%, mostrando não ter risco toxicológico para a saúde humana (Montgomery, 2011; Golding et al., 2014).
Tendo o exposto acima apresentado, justifica-se a necessidade de estudar com maior propriedade as interações dos espessantes nos géis clareadores caseiros com o esmalte dental, a fim de avaliar a eficácia desses produtos, como meio de qualificar a viabilidade de novas formulações clareadoras. Portanto, o objetivo desse estudo, in vitro, foi analisar o efeito
do polímero bioadesivo Aristoflex® AVC no clareamento dental à base de peróxido de carbamida a 10% sobre as propriedades físicas de cor, rugosidade superficial, microdureza e análise química do conteúdo mineral, por meio das concentrações de P e Ca no esmalte dental.
Esta tese será apresentada no formato alternativo de acordo com o Art. 2º da Informação CCPG/001/2015, § 2°.
2 ARTIGO: Effect of Ammonium Acryloyldimethyltaurate/VP Copolymer on the physical and chemical properties of bleached dental enamel.
Effect of polymers on bleaching gels on the properties of enamel. Artigo Submetido ao periódico Juornal of Denristry (Anexo 2)
Thayla Hellen Nunes GOUVEIA1, PhD student Flávio Henrique Baggio AGUIAR1, PhD Gláucia Maria Bovi AMBROSANO2, PhD Débora Alves Nunes Leite LIMA1, PhD
1 Department of Restorative Dentistry, Piracicaba Dental School, State University of Campinas – FOP/Unicamp, Piracicaba, SP, Brazil.
2 Department of Social Dentistry/Statistics, Piracicaba Dental School, State University of Campinas – FOP/Unicamp, Piracicaba, SP, Brazil.
Abstract
Objective: To analyse in vitro the effect of a bioadhesive polymer, Ammonium Acryloyldimethyltaurate/VP Copolymer (Aristoflex® AVC), which is associated with at-home bleaching and is based on a 10% carbamide peroxide (CP 10%) solution, on the physical properties of colour, surface roughness and microhardness and chemical analysis by quantifying phosphorus (P) and calcium (Ca). Methods: Seven groups (n=12) of enamel blocks from bovine incisors (4 × 4 × 2.5 mm) were divided into the following groups: CP 10% with carbopol -Whiteness Perfect®, FGM (WP); CP 10% with Aristoflex (A + CP 10%); CP 10% with carbopol, manipulated (C + CP 10%); Aristoflex (A); carbopol (C); carbamide peroxide 10% (CP 10%); and control (no treatment). Before treatment, the viscosity of the gels was assessed by a modular rheometer, which demonstrated acceptable viscosity values for all study gels. The samples were then submitted to daily treatment applications for 4 hours during 14 days, stored in a remineralizing solution. The colour analyses (ΔE, ΔL*, Δa* and Δb*), profilometry (Ra) and surface microhardness (KHN) were performed before and after the bleaching treatment. The analytical concentrations of Ca and P were performed on the 1st, 3rd, 7th and 14th days by inductively coupled plasma optical emission spectrometry (ICP-OES) for the groups WP, A + CP 10%, C + CP 10%, A, C and CP 10% (control without thickener). The morphology was qualified by scanning electron microscopy (SEM). The ΔE, ΔL* and Δb* data were submitted to ANOVA and Tukey’s tests. Generalized linear models analysed the Δa* data. The roughness profile, microhardness and Ca and P data were evaluated by repeated measurements (PROC MIXED) and a Tukey-Kramer test (a = 0.05). Results: All groups with CP 10%, regardless of the polymer used and differing statistically, were found to include ΔE, ΔL*, Δa* and Δb* compared to the control group. The group A + CP 10% maintained low values of roughness and microhardness after bleaching. Higher concentration of Ca and P were found in the 1st day of treatment for the WP group compared to the groups A + CP 10% and C + CP 10%. However, at Day 14, no statistical difference was observed. Conclusion: The 10% CP solution associated with the bioadhesive polymer Aristoflex® AVC was effective on the bleaching treatment without changing the physical properties of the
dental enamel. Also, this experimental gel caused less mineral loss than the grupo 10% carbamide peroxide with carbopol (commercial product).
Clinical Relevance: Because the change of Carbopol by Aristoflex, a bioadhesive polymer, does not interfere with the effectiveness of bleaching treatment, it is a promising agent associated to carbamide to maintain the physical properties of enamel after bleaching.
Keywords: Dental bleaching, carbamide peroxide, chemical analysis, enamel, microhardness, profilometry, spectroscopy.
Introduction
Dental bleaching has been contemporaneously elected as one of the most popular treatments for the improvement of aesthetic appearance and is considered as a conservative and efficient procedure.[1] On the other hand, this procedure can cause alterations in the morphology of mineralized dental tissues, negatively influencing the physical and chemical properties of the dental enamel as it can result in an increase in roughness, loss of calcium (Ca) and phosphorus (P) as well as a decrease in microhardness. [2-4]
These changes occur due to the action of free radicals, which are generated by the breaking of hydrogen peroxide into oxygen (O*), hydroxyl (2HO*) and perhydroxyl (HO2 *). These radicals, which are extremely electrolytic and diffuse, act in a non-specific way in highly unsaturated organic macromolecules, resulting in dental darkening, as well as with the organic and inorganic matrix of enamel and dentin. [5,6]
Moreover, studies have shown that these changes, especially the reduction of enamel and dentin microhardness, are not only be related to the actions of free radicals but also to other constituents present in the formulation of whitening products, such as bioadhesive polymers. [5,7,8] The functions of the polymers in product formulations are to make the liquid composition of the bleaching agent viscous until it ultimately turns into a gel, prolong the release time of oxygen ions and maintain the product’s intimate contact with the dental surface [7], a result of its bioadhesive property. Carboxypolymethylene (carbopol), often used as an at-home bleaching technique, [9,10] has been associated with decreased enamel microhardness after bleaching treatment. [2,5, 11]
Considering the importance of the polymers in the formulation of bleaching gels and in possible changes in the properties of dental enamel, this study proposed to incorporate another polymer with characteristics opposite to carbopol, Ammonium
Acryloyldimethyltaurate Copolymer (Aristoflex® AVC). This thickening allows the formation of crystalline gels with good consistency and is not dependent on pH, therefore showing high stability at acidic pHs. In the oral cavity, it has been used in the preparation of dentifrices in concentrations of up to 1%, showing no toxicological risk to human health. [26,27] Thus, the polymer is a promising novel bleaching gel for new formulations.
Therefore, the objective of the study was to evaluate the effect of the bioadhesive polymer, Ammonium Acryloyldimethyltaurate/VP Copolymer (Aristoflex® AVC) in a bleaching gel based on 10% carbamide peroxide (10% CP) evaluating the physical properties of colour, surface roughness, and microhardness, as well as in chemical analyses by means of the quantification of minerals Ca and P in dental enamel. The null hypotheses tested were the following: 1) The bleaching gel formulation containing Aristoflex® AVC would not affect the effectiveness of tooth whitening, and 2) the bleaching gel formulation containing Aristoflex® AVC would not alter the physical-chemical properties of dental enamel.
Materials and Methods Preparation of the Specimens
Newly collected bovine incisors that did not have stains or enamel fractures were selected and stored in 0.1% thymol at 4°C until use. A total of 84 bovine incisors were used.
After the selection, blocks of teeth with the following dimensions were obtained: 4 mm in length, 4 mm in width and 2.5 mm in height. For this, the tooth crown was separated from the root and the blocks were obtained by section with a double diamond disc (Extec 4” x 0.12 x 1/2) coupled in a precision metallographic cutter (Isomet 1000, Buehler). The enamel surface was treated with silicon carbide (SiC) granules #600, #2500 and #4000 (Buehler, Lake Bluff, IL, USA) under constant water irrigation using a rotary polymer (Arotec Ind. Com., Cotia, SP, Brazil) to flatten the surface. For the final polishing, felt disks and spray solutions containing diamonds suspended in different granulations of 3 µm, 1 µm and 0.25 µm (Buehler, Lake Bluff, IL, USA) were used. Among the polishing procedures, the samples were placed in an ultrasound device (Marconi, Piracicaba, SP, Brazil) with distilled water and deionized for 15 minutes to remove debris. In the end, the samples were sonicated with a cleaning solution (UltraMetTM 2 SonicCleaningSolution - Buehler, Lake Bluff, IL, USA) to remove any waste.
All specimens were marked on the side with a #1014 spherical drill (KG Sorensen Cotia, São Paulo, Brazil) to standardize the colour and profile (start and end) readings to always be in the same position. The specimens were randomly divided into 7 groups according to the bleaching/thickening treatment: WP (CP 10% solution with carbopol [FGM - Whiteness Perfect]), C + CP 10% (CP 10% with carbopol, manipulated), A + CP 10% (CP 10% with Aristoflex [manipulated and experimental]), C (carbopol gel only), A (Aristoflex gel only), CP 10% only and no treatment (control) (Table 1).
Twenty-four hours before as well as during the experiment, all prepared specimens were stored in artificial saliva in an oven at 37°C, changed daily. The saliva composition was made up of the following: 1.5 µM Ca, 0.9 µM P, 150 µM KCL and 0.1 M Tris buffer with pH 7.0. [2]
Table 1: Composition of bleaching/thickeners agents used in the study.
Bleaching/thickeners agents Composition
10% CARBAMIDE PEROXIDE WITH CARBOPOL (MANUFACTURED) *WHITENESS PERFECT ® - FGM
10% carbamide peroxide, neutralized carbopol, potassium nitrate, sodium fluoride, glycol humectant, deionized water.
10% CARBAMIDE PEROXIDE WITH ARISTOFLEX THICKENER
(EXPERIMENTAL)
10% Carbamide peroxide; 0.2% sodium fluoride; 3% potassium nitrate; 2% Aristoflex AVC®; 0.1% methylparaben; 7% propylene glycol; deionized water qsp 12 g.
10% CARBAMIDE PEROXIDE WITH CARBOPOL THICKENER
(MANIPULATED)
10% Carbamide peroxide; 0.2% sodium fluoride; 3% potassium nitrate; 2% Carbopol 940; 0.1% methylparaben; 7% propylene glycol; aminomethylpropane; Deionized water qsp 12g.
ARISTOFLEX THICKENER 2% Aristoflex AVC®; 0.1% methylparaben; 7% propylene glycol; deionized water qsp 50 g.
CARBOPOL THICKENER 2% Carbopol 940; 0.1% methylparaben; 7% propylene glycol; aminomethylpropane; deionized water qsp 50 g.
10% CARBAMIDE PEROXIDE 10% Carbamide peroxide; 0.2% Sodium fluoride; 3% Potassium nitrate; deionized water qsp 10 mL.
* Bleaching agent formulated by the FGM dental products industry (Joinville, SC, Brazil). Formulation according to manufacturer's data.
Specimen-staining protocol.
Staining the samples was performed by immersing the samples in a solution of tea, which was produced by mixing 1.8 g of black tea (Dr. Oetker Ltda, São Paulo, SP, Brazil) in 100 mL of distilled water, boiled for 3 minutes and infused for 5 minutes. The tea solution was changed every 24 hours for 6 days. After this period of immersion in the solution, the samples were stored in artificial saliva solution for one week and was changed daily for colour stabilization [12]. Prior to the spectrophotometric readings, the black tea dregs that formed on the enamel and dentin surfaces were removed by a single operator using a rubber cup with a mixture of pumice and water (2:1 ratio) at a low speed for 30 s on each side. [16] Preparing the thickening gels and experimental bleaching gels.
The required concentrations and quantities of each component of the product were defined in the pilot study, which experimented with adding the required amount of polymer to ensure the experimental bleaching gels had an acceptable viscosity and close-to-neutral pH. A stable pH for gel formation for carbopol occurred between pH 4.5 to pH 7.0 and for Aristoflex between pH 4 and pH 9.
In this way, 0.2 g of each polymer (carbopol or Aristoflex) added to a mixture of 0.1 g of nipagin and 9 g of deionized water was used to make 10g of each thickening gel. At the end, 0.79 g of the propylene glycol moistener was added to the mixture. To prepare 10 g of the bleaching gel, 8.68 g of the thickening gel (carbopol or Aristoflex) was used, which was added to a mixture of 1 g of carbamide peroxide crystals, 0.02 g of sodium fluoride and 0.03 g of potassium nitrate.
A rheometer (Modular Compact Rheometer-Anton-Paar MCR-102) analysed the viscosity of the gels. The tests were conducted using 50-mm diameter plate geometry (PP50-1) and a 1-mm gap. All tests were performed at 25°C using a solvent barrier to prevent evaporation. The viscosity measurements were performed in the range of 0.01 to 1000s – 1, and the values obtained from the average of three readings for each gel are shown in Table 2.
The pH of the bleaching/thickener gels were measured in duplicates by using a digital pH meter for approximately 3 g of each gel (Table 2).
Table 2: Viscosity (η-) and initial pH of bleaching and thickening gels.
bleaching /thickeners gels Viscosity (𝜼 −) Pa.s
pH
CP 10% with carbopol -FGM 0.243 (0.00751) 5.8
CP 10% with Aristoflex (experimental) 0.191 (0.00252) 6.5 CP 10% with carbopol (compound) 0.179 (0.00265) 6.2
Aristoflex thickening 0.211 (0.00751) 5.3 Carbopol thickening 0.203 (0.00153) 5.9 CP 10% - 7.2 CP 10% - 10% Carbamide peroxide.
Bleaching and thickening treatment protocol
Prior to the bleaching treatment, bulkheads were made with a 2:1 acrylic resin base In the centre, a silicone block of addition (Elite HD + normal setting- © ZermackSpA- BadiaPolesine (RO), Italy) in the dimensions of 5x5 mm was added to create enough space for accommodation for the specimen. After the polymerization of the resin, the block was removed and the specimen was placed with the aid of wax (sticky wax on a stick, ASFER
0.000 0.050 0.100 0.150 0.200 0.250 0.300 FGM A+CP C+CP A C Po wer-l aw i nd ex ( -)
Figure 1. Bar graph of viscosity values (η-) of bleaching and thickeners gels in Pa.s.
Indst. Química Ltda, São Caetano of Sul, São Paulo, SP, Brazil), leaving only the surface of the dental enamel exposed. The samples that received treatment with CP 10% were positioned 1 mm below the level of the bulkhead to prevent the bleaching solution containing CP 10% without any thickener from flowing. In this way, the bulkhead was attached to one end of a stainless steel wire in individual plastic containers in order to facilitate handling and to avoid contamination of the specimen.
The bleaching treatment was performed for 14 consecutive days. The gels were individually weighed and standardized on each analytical balance sample (Shimadzu AUW 220 d, Kyoto, Japan) until reaching a weight of approximately 0.01 g [11]. This gel remained for 4 hours on the surface of the specimen and reserved in hermetically sealed containers and at a relative humidity at 37 o C ± 2 to simulate the oral cavity.
The specimens were washed abundantly in deionized water except days 1st, 3rd, 7th and 14th because for these specific days, the wash water was collected for subsequent chemical analysis of the mineral content. The specimens were then dried with absorbent paper (Kleenex - Kimberly-Clark, Brazil) and stored in 3.0 ml of artificial saliva solution (pH = 7.0) in an oven at 37°C ± 2.
Collecting the rinse solution
On the 1st, 3rd, 7th and 14th days, all gel samples (n = 12) from each group were carefully removed by washing with 3 mL of deionized water in closed bottles and submerging in an ultrasonic bath for 1 minute (Marconi, Piracicaba, São Paulo, SP, Brazil) and 1 minute of vigorous stirring (Solution SizerAP56, Phoenix Luferco, Araraquara, SP, Brazil) for better gel separation from the enamel surface and homogenization of the rinse solution. [11, 13] After sonicating and vigorously stirring the specimens, the specimens were removed from the rinse solution and replaced in individual containers containing artificial saliva. The solution of this rinse was collected and stored at a controlled temperature (-18°C ± 2) to later analyse its concentrations of Ca and inorganic P via inductively coupled plasma optical emission spectrometry (ICP-OES).
Determining the concentration of calcium and phosphorus in the rinse solution.
For the quantification of Ca and P, an ICP-OES (ICap 7400 Duo, Thermo Scientific) was used. In this analysis, groups WP, A + CP 10%, C + CP 10%, A, C and CP 10% (control without thickener) were considered.
In this technique, the sample is dispersed in a gas phase (argon) using a nebulizer to form droplets. These droplets are transported to a region of high temperature plasma between 5500 and 7000 k.
During this passage, the droplets are dried and microcrystals are formed to give rise to the atoms in the elements that make up the sample. When the sample is in plasma, it is possible to excite atoms and gaseous ions. As a result of the excitation process, emission spectra of the atomic and ionic lines of the elements of interest are obtained to identify the elements by their characteristic wavelengths (nm) and calculate concentration from the corresponding emission line intensities.
A solution containing five known concentrations of Ca and P (0.01, 0.1, 1, 2.5 and 5 mg/L) were used to calibrate the apparatus and define specific atomic emission spectra for each mineral. The values for Ca were evaluated at wavelength 393,366 and for P at 213.618. The data were obtained with the aid of software according to the operational parameters: frequency of the generator at 40 MHz; radio frequency power at 1.2 kW; plasma gas flow at 12 L min-1; auxiliary gas flow at 0,5 L min-1; nebulizer gas flow at 0.6 L min-1; sample flow at mL min-1; observation height at 12 mm; pump speed at 60 RPM; integration time of 15s; and replica number of 1.
Colour Measurement
The colour analyses were performed only on the enamel surface before and after the bleaching/thickening treatments. The samples were removed from the apparatus of the acrylic resin then placed in a Teflon device (sample port) inside a GTI Mini Matcher MM1e (GTI Graphic Technology Inc., Newburgh, NY, USA) to standardize the environment of each reading. A previously calibrated Konica Minolta CM-700d spectrophotometer was used according to the manufacturer’s instructions.
The values obtained from the average of three readings for each sample were quantified in three coordinates (L *, a *, b *) of the CIE Lab System, which defined the colour of an object within a three-dimensional space of colours through specific software. The coordinate L * represents the degree of luminosity ranging from 0 (black) to 100 (white). The coordinate a * evaluates the presence of red (a * +) and green (a * -) pigments, and the coordinate b* evaluates the presence of yellow (b * +) and blue (b * -) pigments. [13,14] The differences in the L *, a * and b* values between the initial reading and final (after treatment) were expressed in ΔL, Δa and Δb, and the general colour change was calculated using the following equation: ΔE = [(L1 - L0) 2 + (a1 - a0)2 + (b1 - b0)2]1/2. Before the initial and final readings, the samples were kept in artificial saliva for 24 hours in an oven at 37°C ± 1 for complete hydration.
Surface profile analysis.
The analysis was performed via a profilometer (DEKTAK 150 - Veeco, NY, USA). Three scans were performed at different positions for all specimens by means of a metal tip with a diameter of 12.5 µm. Each scan generated a profile of height as a function of the width travelled in 500 µm of length, with a load of 1.0 mg for a duration of 25 s. Thus, the average roughness (Ra) values of the three scans were considered to be the Ra of each sample. The data were processed in the software Dektak version 9.2. The values were generated in angstrom then converted to millimetres.
Microhardness Knoop analysis
A Knoop microhardness analysis was performed before and after all treatments (whitening/thickening). A microhardness tester (HMV-TW, Shimadzu Corporation, Tokyo, Japan) was used with a Knoop diamond indenter under a load of 25 g for 5 s on the enamel surface in each sample. Five Knoop measurements were performed in the central region of the block with a distance of 100 µM between them. The average of the 5 values obtained were calculated as the surface microhardness (KHN) value: KHN =!",!" ! !"! ! !! ! where C = static load (in g) and a = indentation length.
After determining their microhardness, 84 blocks of enamel that had approximate hardness values were selected. A one-factor analysis of variance (ANOVA) of the obtained data was performed. Because all groups showed similarities (ρ > 0.05), the specimens were randomly divided into 7 experimental groups (Table 1).
Scanning Electron Microscopy
In order to observe the surface enamel of the samples, two samples from each group were randomly selected at the end of the experiment. Samples were plated with gold alloy, and photomicrographs of representative areas with a 1000X magnification were performed by using scanning electron microscopy (SEM; JEOL-JSM 5600 LV-Tokyo, Japan).
Statistical analysis
After the exploratory analysis, the ΔE, ΔL and Δb data were analysed by a one-way ANOVA and Tukey’s test. The Δa data did not meet ANOVA presumptions and were instead analysed by generalized linear models. The profile, microhardness and Ca and P data were analysed by mixed models for repeated measures in time (PROC MIXED) and Tukey-Kramer tests. All analyses were performed using SAS software (Release 9.2, 2010, SAS Institute Inc, Cary, NC, USA) at a level of significance of 5%.
Results Colour
Regarding the ΔE values described in Table 3, the highest averages were found in the treated groups that contained CP 10% regardless of the presence or type of thickener used, with a statistically significant difference compared to the control and to groups that did not contain CP 10% (Aristoflex gel and carbopol gel) (ρ < 0.001). The latter does not differ from each other, but they do differ from the control (ρ < 0.001).
The values of ΔL showed that the groups containing CP 10% (WP, A + CP 10%, C + CP 10% and CP 10%) obtained the highest values of luminosity, differing statistically from the control group (ρ < 0.001). However, the groups treated only with the Aristoflex thickener
and carbopol thickener did not show any statistically significant differences in relation to the control.
Values for Δa and Δb showed that the groups treated with CP 10%, regardless of the presence of thickener (WP, A + CP 10%, C + CP 10% and CP 10%) showed a negative variation, with a statistically significant difference compared to the control (ρ < 0.001). The groups treated only with the Aristoflex thickener and carbopol thickener did not differ statistically from the control group.
Table 3. Mean (standard deviation) of ΔE, ΔL, Δa and Δb as a function of treatments. Bleaching/thickener
Treatment ∆E ∆L ∆a ∆b
CP 10% with carbopol - WP - FGM 8.90 (1.60) a 6.6 (2.1)a -1.1(0.7) c -5.4 (1.9) bc CP 10% with Aristoflex (experimental) 7.49 (1.80) a 3.9 (2.0)ab -1.1 (0.6) c -6.0 (1.8) c CP 10% with carbopol (compound) 8.30 (2.36) a 5.4 (1.3)a -1.2 (0.6) c -6.0 (2.6) bc Aristoflex thickening 4.78 (1.91) b 1.0 (3.5) c -0.4 (0.7)abc -3.5 (1.3) ab Carbopol thickening 4.53 (1.62) b 1.5 (2.5) bc -0.3 (0.6) ab -3.2 (2.3) ab CP 10% 8.24 (1.79) a 6.1 (1.8) a -1.0 (0.5) bc -5.1 (1.9) bc Control 2.69 (1.65) c -0.1 (1.6) c 0.0 (0.8) a -1.7 (2.1) a Means followed by distinct letters differ from each other (p≤0.05) in the column within each coordinate. CP 10% - 10% Carbamide Peroxide.
Surface profilometry
The results described in Table 4 refer to the profile values of the specimens before and after the different treatments. All groups before the treatment had no statistically significant differences. However, after the application of the different treatments, only A + CP 10%, CP 10% and the control maintained their initial profile values, showing no statistically significant differences.
After applying the treatments, it was observed that the highest values of roughness, detected by the profilometer, was in the WP group, with a statistically significant difference compared to the control group (ρ < 0.001). The groups treated only with thickeners (Aristoflex and carbopol) also presented high values in their profiles after treatment, differing statistically from the control (ρ < 0.001), but did not differ much from each other. The group
treated with CP 10% with Aristoflex obtained statistically low values of roughness; it did not differ much from the control nor from C + CP 10% and CP 10%.
Table 4. Mean (standard deviation) of profilometry (mm) as a function of treatment and time. Bleaching/thickener Treatment Time Before After CP 10% with carbopol -WP - FGM 0.06 (0.01) Ba 0.12 (0.04) Aa CP 10% with Aristoflex (experimental) 0.06 (0.01) Aa 0.07 (0.02) Acd CP 10% with carbopol (compound) 0.05 (0.01) Ba 0.09 (0.02) Aabc
Aristoflex thickening 0.05 (0.01) Ba 0.10 (0.01) Aab Carbopol thickening 0.06 (0.01) Ba 0.11(0.04) Aab
CP 10% 0.06 (0.01)Aa 0.09 (0.02) Abc
Control 0.05 (0.01)Aa 0.05 (0.01) Ad
Means followed by distinct letters (uppercase in horizontal and lower case in vertical) indicate statistical differences (p≤0.05). CP 10% - 10% Carbamide Peroxide.
Surface microhardness
All groups before the treatment had no statistically significant differences. However, after the application of the different treatments, only A + CP 10% and the control maintained their initial microhardness values, showing no statistically significant differences.
In the final evaluation, the lowest microhardness values were observed to be in the groups containing only thickeners, which differed statistically from each other and in comparison to the other groups (ρ < 0.001). The groups that contained carbopol as a thickener (WP and C + CP 10%) and the group treated with CP 10% only had intermediate microhardness values compared to the control group and other groups. On the other hand, A + CP 10% presented the highest value of microhardness compared to other groups, without statistically differing much from the control group.
Table 5. Mean (standard deviation) of microhardness (KHN) as a function of treatment and time. Bleaching/thickener Treatment Time Before After CP 10% with carbopol WP -FGM 357.41 (19.06) Aa 297.48 (20.62) Bb CP 10% with Aristoflex (experimental) 357.10 (17.72) Aa 335.43 (20.12) Aa CP 10% with carbopol (compound) 357.35 (17.43) Aa 304.61 (28.30) Bb Aristoflex thickening 357.04 (17.32) Aa 187.42 (18.43) Bc Carbopol thickening 357.35 (17.45)Aa 110.89 (7.32) Bd
CP 10% 357.39 (15.35) Aa 294.20 (32.25) Bb
Control 357.23 (15.95) Aa 349.40 (13.30) Aa
Means followed by distinct letters (upper case in horizontal and lower case in vertical) indicate statistical differences (p≤0.05). CP 10% - 10% Carbamide Peroxide.
Calcium and phosphorus concentration
When comparing the samples between different days, only the CP 10%-treated groups were observed to be statistically significant different between the 3rd and 14th day of treatment (ρ < 0.001). On the last day (14th), Ca loss increased. In the other groups, no statistically significant differences were observed (Table 6).
On the first day of whitening, we observed that the WP group had a higher Ca mineral loss with a statistically significant difference compared to A + CP 10%, C + CP 10% and CP 10% only (ρ < 0.001) but did not differ statistically from groups A and C.
On the 3rd and 7th days of treatment, statistical differences were not observed between the WP group and the groups with experimental or manipulated bleaching gels (i.e., A + CP 10% and C + CP 10%). However, differences were observed in CP 10%, which had higher concentrations of Ca loss compared to the WP group (ρ < 0.001). On the 14th day, no statistically significant differences were found.
Table 6. Mean (standard deviation) of Caconcentration (mg/L) in rinsing water as a function of treatment and time.
Bleaching/thickener Treatment
Ca (mg/L) in rinsing water
1st day 3rd day 7th day 14th day CP 10% with carbopol
-WP - FGM 1.33 (0.49) Aa 1.13(0.27)Aa 0.98 (0.36)Aab 0.81 (0.37)Aa CP 10% with Aristoflex
(experimental) 0.58 (0.17) Ad 0.67(0.37) Aab 0.67 (0.36)Ab 1.05 (0.67)Aa CP 10% with carbopol
(compound) 0.69(0.15) Abcd 0.63(0.23) Aab 0.96 (0.21)Aab 1.13 (0.58)Aa Aristoflex thickening 1.25 (0.36) Aab 1.18(0.25) Aa 1.35 (0.33)Aa 1.71 (0.79)Aa Carbopol thickening 1.10(0.21) Aabc 1.10 (0.52) Aa 0.98 (0.31)Aab 1.39 (0.45)Aa CP 10% 0.65(0.16) ABcd 0.36 (0.09) Bb 0.72(0.27)ABb 1.10 (0.55)Aa Means followed by distinct letters (uppercase in horizontal and lower case in vertical) indicate statistical differences (p≤0.05). CP 10%- 10% Carbamide Peroxide.
The concentration of P in the rinse solution during the bleaching treatment are shown in Table 7. When comparing the solutions between the different days, statistically significant differences were found only in the group treated with CP 10%. Between the 3rd and 7th day, increasing concentrations of P loss (ρ < 0.001) were found for this group.
The WP group showed statistically significant differences from the 1st, 3rd and 7th days compared to the other treatments. No statistically significant differences were found.
Table 7. Mean (standard deviation) of P concentration (mg/L) in rinsing water as a function of treatment and time.
Bleaching/thickener Treatment
P (mg/L) in rinsing water
1st day 3rd day 7th day 14th day CP 10% with carbopol
-WP - FGM 2.57 (0.30) Aa 2.04(0.30) Aa 2.44(0.68)Aa 2.10 (0.86)Aa CP 10% with aristoflex
(experimental) 0.66 (0.07) Abc 0.63(0.29) Ab 0.56 (0.14)Ab 1.25 (0.75)Aa CP 10% with carbopol
(compound) 0.74 (0.21) Ab 0.47(0.07) Ab 0.95(0.24)Ab 0.97 (0.51)Aa Aristoflex thickening 0.53 (0.12) Abc 0.45 (0.16) Ab 0.71 (0.46) Ab 1.09 (0.65)Aa Carbopol thickening 0.39 (0.06) Ac 0.57 (0.49) Ab 0.70 (0.30)Ab 0.66 (0.37)Aa CP 10% 0.57 (0.18) ABbc 0.47 (0.11) Bb 1.04 (0.27) Ab 1.17 (0.50)ABa Means followed by distinct letters (uppercase in horizontal and lower case in vertical) indicate statistical differences (p≤0.05). CP 10%- 10% Carbamide Peroxide.
By observing images of the samples, some changes in enamel morphology after the application of the different treatments (Figure 3) were found when compared to the control group (Figure 2).
When compared to the control group (Figure 2), the group treated with CP 10% with carbopol-WP (Figure 3a) presented a surface with evidence of an enamel prism, suggesting demineralization with an interprismatic dissolution. A similar finding was observed in the C + CP 10% group (Figure 3c) but with less evidence of dissolution when compared to Figure 3a. No demineralization patterns were observed in A + CP 10% (Figure 3b), but a similarity of this group to the control group was observed (Figure 2).
Surfaces treated with the Aristoflex thickener presented with a surface full of pores and depressions (Figure 3d). However, the surface treated with the carbopol thickener presented with regions of evident prisms covering the entire observed surface (Figure 3e). It did have some pores but to a lesser extent to the Aristoflex group. Pores were also found on the surface of the group treated with CP 10% only; however, no prismatic dissolution patterns were observed in this group.
Figure 2. Scanning electron microscopy analysis of the enamel surface of the control (no
a
b
c
d
e
f
Figure 3. Scanning electron microscopy analysis of the enamel surface after bleaching/ thickening treatment. a) WP (CP 10% with carbopol -FGM); b) A + CP 10% (CP
10% with Aristoflex (experimental)); c) C + CP 10% (CP 10% with carbopol (manipulated)); d) A (Aristoflex thickening); e) C (carbopol thickening); f) CP 10% (10% carbamide peroxide).
Discussion
The first null hypothesis, which tested whether the bleaching gel formulation containing Aristoflex® AVC would affect the effectiveness of tooth whitening, was accepted. All groups containing CP 10% regardless of the type of thickener used showed significant changes in the general colour of the specimens and in all the colour dimensions of the CIE system (L *, a * and b *) after bleaching procedures.
In this study, the specimens were stained with a solution of black tea to standardize their initial colour. [15] This solution darkened the specimens in reddish-brown shades. In doing so, any reduction in the colour of these shades would prove the effectiveness of the bleaching method. The values of Δa and Δb, the parameters describing the chroma of an object, tended to have negative values in the three-dimensional colour system. The changes occurred from the a * + (red) axis to the a * - (green) axis, showing that the samples were less reddish. The specimens could also change from the b * + (yellow) axis to the b * - (blue) axis to become, in other words, less yellowish. The values for ΔL revealed an increase in the luminosity of all specimens treated with CP 10% regardless of the type of polymer used. [14,15] The ΔE parameter, which provides information about the overall colour change of an object, was also observed. The scientific literature states that a threshold of perception in colour variation occurs when ΔE is 3 (ΔE = 3.7). [16] In the findings of this study, all groups that contained CP 10% presented a ΔE value of 7.4, which would be clearly accepted and shows a noticeable change in colour. This value differs widely from the control group, whose variation was 2.69. The process of dental bleaching occurs through the penetration and diffusion of bleaching agents into the enamel and dentin via the structures of prisms. Hydrogen peroxide dissociates into free radicals, which oxidize long-chain organic molecules responsible for the coloration of dental tissue. [18,19] These agents are able to penetrate the permeable tissue of the dental enamel through the structures of prisms to reach and break down the dentin and organic molecules, thus promoting bleaching. [14, 18- 22]
Nevertheless, the groups treated only with the thickeners Aristoflex gel and carbopol gel presented a ΔE value of 4.7 and 4.2, respectively, and a change in colour was perceived and accepted. However, in the isolated evaluation of each coordinate (ΔL, Δa and Δb), no statistically significant differences were found between the two groups and the control group. Despite this difference in colour change, thickeners are known to lack the ability to whiten the tooth structure. Thickening polymers consist of large molecules with a high molecular weight used in formulations with a viscosity agent, which acts to give a gel consistency to the
product. In the bleaching process, they act as modellers in the oxidation and reduction chemical reaction process to release hydrogen peroxide, and the latter is responsible for the bleaching process. [5, 7] This change may be related to the acidic nature of viscosity agents, which may cause surface demineralization and alter the morphology of the enamel. As a consequence, the surface reflectance pattern of the specimens may have been altered.
Carbopol, or carboxypolymethylene, comprises synthetic, water-soluble polymers used as a gelling agent in aqueous systems and emulsions and given as a solution for viscosity. [23,10] Changes in the dental surface caused by this thickening have been attributed to carbopol’s low pH and high viscosity. Studies have shown that this thickener is derived from a carboxylic acid, has an acidic pH and may contribute to dental demineralization. [24,5,25] The Ammonium Acryloyldimethyltaurate/VP Copolymer (Aristoflex® AVC) is a polymer of a similar acidic nature, derived from synthetic (cationic) polysulfonic acid. This agent is used as a gelling agent or thickener in personal care formulations and is one of the constituents of oral hygiene products such as whitening toothpaste and oral rinses. [26, 27]
Hence, these thickeners promote a superficial demineralization to stimulate the leaching of the chromogenic pigments in the superficiality of the enamel, which influences the values of ΔE. Yet, for a viscosity agent to be incorporated into a bleaching product formulation, it must be buffered at a neutral pH and thus act as a compound so as not to negatively influence the bleaching reaction. [23] This step was carefully evaluated before starting the manipulation of the experimental and manipulated bleaching gels.
The second null hypothesis, which tested whether bleaching gel formulations containing Aristoflex® AVC would alter the physical and chemical properties of dental enamel, was accepted. The surface roughness and microhardness values of the specimens were not altered after applying the bleaching treatment with Aristoflex. In addition, the analytical concentrations of Ca and P were lower for this group on the 1st day of bleaching compared to the WP group, which presented higher concentration for both minerals.
Ávila et al., [8] a study on in vitro chemical erosion, demonstrated the interaction of some polymers with the dental structure. The study showed that polymers used in various oral products increase the viscosity of the formulations, increase the bioavailability of the drugs and/or active substances in the products to effectively act on the oral environment and interact with the dental structure due to their bioadhesive capacity. This bioadhesive capacity is related to possible ionic bonds between the polymers, negatively charged centres forming what the authors called “film” (i.e., a layer of deposited polymer on the dental structure). In
the findings of the Ávila et al., [8] a film formed with the carbopol thickener was very strong and thick, able to resist subsequent erosive cycles. When the other thickeners tested (natrosol and Aristoflex) were used, the authors suggested that the bonds were not strong enough to resist the acid challenge, that is, the formed film was not as effective as with carbopol. This shows the great affinity of carbopol with the dental structure.
The result of the cited study concur with this study’s results on microhardness and profilometry; we observed reduced values of microhardness and increased roughness in the groups that contained carbopol alone or were associated with CP 10% (commercial and manipulated). These findings can also be observed in the SEM images in Figures 5a, 5c and 5e. A possible explanation may be linked to the formation of a film; in a bleaching treatment, this formed film acts as a barrier to prevent salivary remineralization, which is necessary to reduce or prevent the mineral loss caused by hydrogen peroxide throughout the treatment. [28, 2]
For the treated group with Aristoflex gel alone, the A group, which had no association with CP 10%, a significant reduction in the values of microhardness and increased roughness, in addition to a porous surface with an erosive appearance (Figure 5d), were observed. This could have occurred because of the cationic nature of this polymer, which has a negatively charged phosphate attraction. However, if we hypothesize that this bond with the enamel is inferior to the carbopol bond, which has an anionic nature with a positively charged Ca centre, [8] the amount of Ca in the hydroxyapatite would be greater than the amount of phosphate ions (Ca10(PO4)6(OH)2). However, even with a reduced number of bonds, the formed film would prevent salivary remineralization. As a result, this is why changes in the physical properties were reduced in the Aristoflex group.
However, when evaluating Aristoflex in association with CP 10%, the microhardness and roughness values were maintained even after treatment and did not differ from the control group. A uniform surface, absent from demineralization patterns or pores, was observed in the SEM image (Figure 5b). An explanation for this finding may be due to a weakly formed film by Aristoflex with the enamel surface, as previously discussed. Due to its cationic nature, this polymer may have attracted fluoride ions, which are available in enamel, to bind with its positive sites, further decreasing the number of possible connections between the polymer and the dental structure and consequently decreasing the formation of this film but allowing the dental surface to be free for salivary remineralization.
It is also important to highlight the findings of the group treated only with CP 10%, whose microhardness values were also reduced in relation to the control group and roughness values were presented intermediately. This finding can be attributed to the absence of the reaction modeller; without the thickeners, peroxide degradation occurs faster and more directly on the dental surface. [29] Hydrogen peroxide can produce different types of reactive oxygen species. Under alkaline conditions, hydrogen peroxide works via the hydroperoxyl anion (HO2-), and, under different pH conditions, certain free radicals, such as the superoxide anion (O2-) and the hydroxyl anion (OH-), are formed. Therefore, these radicals may have interacted with the organic and inorganic matrix of the enamel, encouraging demineralization and consequently, reducing microhardness values and alterations in surface morphology, as observed in Figure 5f.
Because Ca and phosphate ions are the main constituents of the crystals in hydroxyapatite, the presence of these minerals was observed when collecting the rinse solution during the whitening treatment, indicating that a demineralization process may have occurred. [11, 13] When comparing the 1st, 3rd, 7th and 14th days of bleaching treatment, statistical differences between the treatments performed were not observed, except for the CP 10% (without thickener) group, which showed an increase in mineral concentration for Ca between the 3rd and 14th days and for P between the 3rd and 7th days.
However, to quantify Ca on the 1st day of bleaching, we observed significant differences between the WP group, which had a higher concentration of CA compared to A + CP 10%, C + CP 10% and CP 10% only. For P, the CP 10% group with carbopol - WP presented significantly higher concentration of P compared to all other treatments. Cavalli et
al., [11] when studying the efficacy of commercial and experimental agents containing additives such as Ca and fluorine in the maintenance of the physical and chemical properties of the enamel, found that a bleaching product with a CP 10% carbopol-WP solution showed a high percentage of mineral loss and a decrease in microhardness values. It was unable to reestablish mineral loss through salivary remineralization throughout the bleaching treatment. In addition, due to the product’s high viscosity, acidic pH and thick film formation, which is formed by polymer and dental minerals, greater demineralization occurred in this group, and, consequently, a greater concentration of Ca and P was observed.
Although the values of Ca and P minerals were found on the 1st day of treatment, A + CP 10% did not present significant differences compared to the C + CP 10% group; this corresponds to an observable trend of a lower concentration of Ca and P in the rinse solution
