LETÍCIA CABRERA CAPALBO 

 

 

 

 

 

 

 

ANÁLISE IN VITRO DA CAPACIDADE DE SOLUÇÕES CONTENDO 

FLUORETO E/OU HEXAMETAFOSFATO NA REMINERALIZAÇÃO 

DA DENTINA 
 

 

 

 

 

 

 
 

ARAÇATUBA - SP 
2021

Campus de Araçatuba 



LETÍCIA CABRERA CAPALBO 

 

 

 

 

 

 
 

ANÁLISE IN VITRO DA CAPACIDADE DE SOLUÇÕES 

CONTENDO FLUORETO E/OU HEXAMETAFOSFATO NA 

REMINERALIZAÇÃO DA DENTINA 
 

 

Dissertação apresentada à Faculdade de 

Odontologia de Araçatuba da 

Universidade Estadual Paulista “Júlio de 

Mesquita Filho” – UNESP, como parte dos 

requisitos para a obtenção do título de 

Mestre em Ciência Odontológica – Área 

Saúde Bucal da Criança. 

 

 

 

Orientador: Prof. Assoc. Juliano Pelim Pessan 

Coorientador: Prof. Tit. Alberto Carlos Botazzo Delbem 

 
 
 

ARAÇATUBA - SP 
2021



 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Catalogação-na-Publicação (CIP) 

Diretoria Técnica de Biblioteca e Documentação – FOA / UNESP 

 
 Capalbo, Letícia Cabrera. 
C236a  Análise in vitro da capacidade de soluções  
 contendo fluoreto e/ou hexametafosfato na remi- 
 neralização da dentina / Letícia Cabrera Capalbo. - 
 Araçatuba, 2020 
  63 f. : il. ; tab.  
 
  Dissertação (Mestrado) – Universidade Estadual  
 Paulista, Faculdade de Odontologia de Araçatuba 
  Orientador: Prof. Juliano Pelim Pessan 
  Coorientador: Prof. Alberto Carlos Botazzo Delbem 
 
  1. Dentina 2. Flúor 3. Fosfatos 4. Remineralização 
 dentária I. T. 
 
    Black D27 
    CDD 617.645 
 

Claudio Hideo Matsumoto – CRB-8/5550 
 
 
 
 



 

Dados Curriculares 
LETÍCIA CABRERA CAPALBO 

 
Nascimento 

 

05/08/1994 Araçatuba-SP 

 

Filiação Marcos Roberto Capalbo 

Lúcia Sanchez Cabrera Capalbo 

 

2012/2016 Curso de graduação em Odontologia na Universidade Estadual 
Paulista “Júlio de Mesquita Filho” - UNESP 

 

2016/2019 Especialização em Ortodontia – FACSETE (NEC) 

 

2019/Atual Curso de Pós-Graduação em Ciência Odontológica – área Saúde 
Bucal da Criança, nível de Mestrado, na Faculdade de Odontologia 
de Araçatuba- UNESP 

  

 

Associações 

CROSP- Conselho Regional de Odontologia de São Paulo 

SBPqO- Sociedade Brasileira de Pesquisa Odontológica 

IADR – International Association for Dental Research 

 

  



 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Dedicatória 
_______________



Dedicatória 

Letícia Cabrera Capalbo 

 

 

Aos meus pais, Marcos e Lúcia: 
Por todo apoio, suporte, dedicação e amor por mim. Por serem meus 

maiores incentivadores e sempre me ensinarem a batalhar pelos meus sonhos. 

Por sempre cuidarem de mim com tanto amor e carinho e me ensinarem que a 

humildade, perseverança, honestidade e fé são nossas maiores virtudes. Sou 

grata a Deus por ter me dado vocês como pais. Admiro a força e dedicação de 

vocês e me orgulho de ser sua filha! Obrigada por tudo, amo vocês! 

 

À minha irmã, Bruna: 
Por ser minha primeira e melhor amiga, apesar da distância e de nossas 

brigas. Por ser minha companheira de todas as horas e por nunca medir 

esforços para me apoiar e me ver bem e feliz. Meu grande exemplo como 

Cirurgiã-dentista, mãe e pessoa. Tenho muito orgulho de você e sei que jamais 

estarei sozinha, pois tenho você ao meu lado! Obrigada por tanto, amo você! 

 

Aos meus avós: Maria, Manoel (in memoriam) e 

Mafalda (in memoriam), 
Pelos ensinamentos, carinho e amor incondicional. Quem me dera vocês 

fossem eternos! Sou grata por cada palavra de carinho, por todo apoio e 

sabedoria de vida a mim passados. Quanto orgulho tenho em fazer parte da 

família que vocês construíram! Obrigada por terem me ensinado os reais 

valores da vida. Infelizmente já não posso ter todos vocês em vida comigo, mas 

sei que de onde estiverem, seguem olhando e cuidando de mim! Amo vocês! 

  



Dedicatória 

Letícia Cabrera Capalbo 

 

  

À minha afilhada, Maria Clara: 
Meu maior e melhor “presente”. Sou grata a Deus por ter te colocado em 

nossa família, trazendo ainda mais alegria para nós! Minha modelo para aula 

de Odontologia para bebês e que sempre quer escovar os dentinhos com a 

Dinda! Cheia de saúde e inteligência, não tem como não sorrir ao seu lado. A 

dinda sempre estará com você, te apoiando e incentivando! Meu amor por você 

é incondicional! 

 

 

Ao meu noivo, Renan: 
Por todo amor, carinho, respeito e companheirismo que tem comigo! Sou 

grata a Deus por ter te colocado em minha vida! Depois de 8 anos juntos posso 

dizer que a vida não tem graça sem você! Meus dias são mais felizes ao seu 

lado e não há distância nesse mundo que possa mudar meu amor e admiração 

por você! Obrigada por tantos momentos maravilhosos, por cada palavra, 

carinho, por apoiar minhas decisões e me ajudar a conquistar meus sonhos!  

Meu amor e admiração por você são eternos! 

 



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

Agradecimentos especiais 
___________________________________



Agradecimentos Especiais 

Letícia Cabrera Capalbo 

 

A Deus 
Pela minha vida, saúde e por todas as pessoas maravilhosas colocou em 

meu caminho. Sinto-me abençoada por ter tantas pessoas boas ao meu lado e 

por conseguir realizar meus sonhos. Agradeço por me iluminar nas tomadas de 

decisões e orientar meus caminhos. 

 

À minha família 
Por serem meus maiores incentivadores e motivadores. Meus exemplos de 

pessoas. Obrigada por estarem sempre ao meu lado e fazerem com que eu 

nunca desistisse dos meus sonhos! Cada conquista em minha vida é por 

vocês, sem seu apoio nada seria possível. 

 

Ao meu orientador prof. Juliano Pelim Pessan 
Pela oportunidade em realizar meu sonho e fazer pós graduação em 

Odontopediatria. Por ser um grande orientador e incentivador. Por compartilhar 

seus ensinamentos com sabedoria e paciência. Pela confiança, suporte e ajuda 

oferecidos a mim para que a pesquisa fosse concluída com sucesso. Obrigada 

por sua disponibilidade, atenção e carinho e com todos seus alunos! A cada dia 

aprendo mais com você e me orgulho em fazer parte do seu time! 

  



Agradecimentos Especiais 

Letícia Cabrera Capalbo 
 

 

 

Ao meu noivo Renan e à sua família 
Aos meus sogros, Reinaldo e Cristina, obrigada por todo carinho e torcida 

por mim! À minha cunhada Rafaela, obrigada pela amizade e tantos bons 

momentos vividos juntas. Aos avós de Renan, Cecília e Dante, obrigada por 

tanta generosidade e fazerem nossos dias mais alegres. Vocês moram em meu 

coração! 

Ao Renan, obrigada mais uma vez por ser meu porto seguro e estar 

sempre disposto a me ajudar. Seu apoio e incentivo são muito importantes para 

mim. Ao seu lado tudo fica melhor, meu companheiro para a vida! 

 

Ao professor Alberto Carlos Botazzo Delbem 
Por toda sua ajuda, paciência e ensinamentos passados. Sua sabedoria, 

experiência e dedicação foram essenciais no desenvolvimento desse projeto. 

Agradeço a disponibilidade, boa vontade e atenção com todos os alunos, 

garantindo o bom funcionamento do laboratório.  

 

Aos professores Robson F. Cunha e Cristiane Duque 
Por todos ensinamentos passados em aulas e clínicas e pela boa 

convivência. Sou grata por ter a oportunidade de aprender diariamente com 

professores e pessoas excepcionais como vocês.   

  



Agradecimentos Especiais 

Letícia Cabrera Capalbo 
 

 

Às minhas amigas Carol Barros, Lara Cervantes, 

Mariana Pereira, Bruna Egumi, Thayane Businari, Isa 

Catanoze, Betina Commar e Luara Colombo  
Por se fazerem presentes mesmo distantes, por serem grandes amigas e 

pessoas incríveis. Obrigada pelo carinho, apoio e desejos de sucesso!  Nossa 

amizade se iniciou na graduação, mas continua para toda a vida, pois nosso 

amor e admiração umas pelas outras são sinceros! Obrigada pela boa 

companhia e momentos inesquecíveis vividos juntos! Amo cada uma de vocês! 

 

Ao meu primo Rodrigo e aos amigos Hiskell e Leopoldo 

Meus grandes amigos desde antes do início da pós graduação. Com vocês 

dividi meus medos, frustrações e também as alegrias da vida pessoal e 

acadêmica. Vocês sempre estiveram dispostos a ajudar e aconselhar, além de  

torcerem verdadeiramente por mim! Sou grata por ter pessoas como vocês em 

minha vida! 

 

Ao Gabriel Pereira Nunes 
Pelo grande companheirismo e aprendizado que tivemos juntos durante 

estes dois anos. Pela paciência, disposição e dedicação para me ensinar e 

ajudar durante todas as fases desse projeto. Com você dividi preocupações, 

dificuldades e alegrias da pós graduação. Agradeço a amizade, parceria e por 

me ajudar a enfrentar cada obstáculo dessa caminhada! 

 

À Marcelle Danelon 
 Pela ajuda, cooperação e assistência prestadas. Agradeço pela 

generosidade em compartilhar sua experiência e conhecimento, tornando esse 

trabalho mais completo. 

 



Agradecimentos Especiais 

Letícia Cabrera Capalbo 
 

 

À Mayra Frasson, Liliana Báez, Igor Zen, Caio 

Sampaio, Tamires Passadori, Isabela Ferreira e 

Rodrigo Sakuma 
Caio, Mayra e Igor: vocês foram essenciais durante esse período, pois 

além de serem companheiros e amigos maravilhosas, também são pessoas 

nas quais me inspiro! Desde o começo, antes de prestar a prova para o 

Mestrado, vocês me ajudaram e por isso sou eternamente grata! 

Liliana, Tamires, Rodrigo e Isa: a convivência com vocês é maravilhosa. 

Dividimos muitos momentos bons e também preocupações, mas sempre 

percebemos que juntos somos mais fortes! 

A amizade de todos vocês é muito importante para mim, sou grata por tê-

los conhecido e ter o prazer de conviver com vocês! 

 
Aos amigos do laboratório e pós graduação: Thayse 
Hosida, Leonardo Morais, Heitor Ceolin, Renan 
Ceolin, Amanda Andolfatto, Priscila Toninato, Lais 
Arias, Nayara Gonçalves, Francyenne Castro, Ana 
Paula, Jesse Augusto, Rafaela Laruzo, Juliana 
Morabito, Karina Caiaffa, Marcela Macedo, Vanessa 
Rodrigues, José Antônio Souza, Viviane Zequini, Ana 
Carolina Lisboa 

Obrigada por trazerem leveza e diversão aos meus dias. O convívio com 

vocês durante esses dois anos foi muito bom! Obrigada por terem me recebido 

tão bem no laboratório e me ajudado quando precisei! Dentro ou fora do 

laboratório, a amizade e companheirismo de vocês fez muita diferença na 

minha caminhada. 



 

   

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Agradecimentos 
__________________________________________



Agradecimentos 

Letícia Cabrera Capalbo 
 

 

À Universidade Estadual Paulista “Júlio de Mesquita Filho”, na pessoa 

do diretor da Faculdade de Odontologia de Araçatuba, Prof. Tit. Glauco 
Issamu Miyahara, e do vice-diretor, Prof. Tit. Alberto Carlos Botazzo 
Delbem. 

 

Ao programa de Pós-Graduação em Ciência Odontológica da Faculdade 

de Odontologia de Araçatuba – UNESP, representado pelo seu coordenador 

Prof. Assoc. Luciano Tavares Ângelo Cintra, pela competência e qualidade 

na condução do programa de pós-graduação. 
 

Aos funcionários da Seção Técnica de Graduação e Pós-Graduação da 

Faculdade de Odontologia de Araçatuba - UNESP, Valéria Queiroz 
Marcondes Zagatto, Lilian Sayuri Mada e Cristiane Regina Lui Matos, pela 

eficiência e profissionalismo. 

 

Aos funcionários da área de Odontopediatria da Faculdade de Odontologia 

de Araçatuba - UNESP, Luiz, Mário e Ricardo, pela ajuda, atenção e suporte 

prestados a mim. 

 

À Coordenação de Aperfeiçoamento de Pessoal de Nível Superior 
(CAPES) pelo apoio financeiro nesses dois anos. 

 

Ao Frigorífico Olhos D’Água e ao sr. Amâncio de Oliveira, que 

disponibilizaram os dentes bovinos. A ajuda e generosidade de vocês foi 

essencial para a realização desse trabalho! 

 

A todos que direta ou indiretamente contribuíram para a concretização 

deste trabalho, 

 

Minha eterna gratidão!



 

 

 

 
 
 
 
 
 
 
 
 
 
 

Epígrafe 
_______________________



Epígrafe 

Letícia Cabrera Capalbo 
 

 
 
 
 
 
 
 
 

“Feliz aquele que transfere o que sabe e aprende o que ensina.” 
 

Cora Coralina 
 
 
 
 
 

“Por vezes sentimos que aquilo que fazemos não é senão uma 
gota de água no mar. Mas o mar seria menor se lhe faltasse 

uma gota.” 
 

Madre Teresa de Calcutá



 

 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

  
 
 

Resumo 
___________________



Resumo 

 
Letícia Cabrera Capalbo 

Capalbo, LC. Análise in vitro da capacidade de soluções contendo fluoreto 
e/ou hexametafosfato na remineralização da dentina. 2020. Dissertação 

(Mestrado em Ciência Odontológica, área de Saúde Bucal da Criança) - 

Faculdade de Odontologia de Araçatuba, Universidade Estadual Paulista, 

Araçatuba 2020. 
 

O objetivo do presente estudo foi avaliar a capacidade de soluções contendo 

HMP e F, sozinhos ou em associação, em induzir a remineralização dentinária 

em um protocolo in vitro. Blocos de dentina radicular bovina (4 × 6 cm, n = 100) 

foram preparados e submetidos à indução de lesões de cárie artificiais em dois 

terços da superfície; cada bloco foi utilizado como seu próprio controle. Em 

seguida, os blocos foram divididos em 10 grupos experimentais (n=10/grupo), 

de acordo com as soluções a serem testadas: (1) Placebo (sem F ou HMP); (2) 

0,5% HMP; (3) 0,75% HMP; (4) 1% HMP; (5) 250 ppm F; (6) 500 ppm F; (7) 

1100 ppm F; (8) 250 ppm F + 0,5% HMP; (9) 500 ppm F + 0,75% HMP e (10) 

1100 ppm F + 1% HMP. Os blocos foram tratados por um minuto, duas vezes 

ao dia com as respectivas soluções, e submetidos a uma ciclagem de pH 

durante 7 dias. Em seguida, foram determinadas a porcentagem de 

recuperação da dureza de superfície (%RDS) e a área integrada da lesão de 

subsuperfície (ΔKHN). Os dados foram submetidos a ANOVA e teste de Fisher 

LSD (p<0.05). Uma relação dose-reposta foi observada entre as concentrações 

de F nas soluções sem HMP e as variáveis %RDS e ΔKHN; quanto às 

soluções contendo apenas HMP, uma relação dose-resposta foi observada 

somente para ΔKHN. Em relação à %RDS, os grupos placebo e 0,5% HMP, e 

os grupos 0,75% e 1% HMP não apresentaram diferenças significativas entre 

si. Quando associado ao F, o HMP aumentou a capacidade de remineralização 

da superfície e subsuperfície dentinária, visto que os grupos contendo F + HMP 

apresentaram resultados significativamente melhores em relação aos grupos 

contendo F sozinho. Em acréscimo, a solução contendo 250 ppm F + 0,5% 

HMP promoveu um efeito remineralizador semelhante à solução contendo 500 

ppm F.  Já em relação à ∆KHN, diferenças estatísticas foram observadas entre 

todos os grupos na área tratada, sem diferenças significativas quanto às áreas 

controle e desmineralizada. Os resultados permitem concluir que a adição de 

HMP às soluções fluoretadas potencializou o efeito destas sobre a 



Resumo 

 
Letícia Cabrera Capalbo 

remineralização das lesões artificiais de cárie em dentina, tanto na superfície 

quanto em profundidade.  

 

Palavras–chave: Dentina. Flúor. Fosfato. Remineralização Dentária. 



 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 Abstract 
_______________________



Abstract 

Letícia Cabrera Capalbo 

 

Capalbo, LC. In vitro analysis of the capacity of solutions containing 
fluoride and/or hexametaphosphate on the remineralization of dentin. 
2020. Dissertação (Mestrado em Ciência Odontológica, área de Saúde Bucal 

da Criança) - Faculdade de Odontologia de Araçatuba, Universidade Estadual 

Paulista, Araçatuba 2020. 

 

The present study aimed to investigate the ability of solutions containing HMP 

and F, alone or in association, in promoting dentin remineralization in an in vitro 

protocol. Bovine root dentin blocks (4 × 6 cm, n = 100) were prepared, and 

caries-like lesions were induced in two thirds of the surface; each block served 

as its own control. Then, blocks were divided into 10 experimental groups (n = 

10 / group), according to the solutions to be tested: (1) Placebo (without F or 

HMP); (2) 0.5% HMP; (3) 0.75% HMP; (4) 1% HMP; (5) 250 ppm F; (6) 500 

ppm F; (7) 1100 ppm F; (8) 250 ppm F + 0.5% HMP; (9) 500 ppm F + 0.75% 

HMP and (10) 1100 ppm F + 1% HMP. Specimens were treated for one minute, 

twice a day with the respective solutions, and subjected to a pH-cycling regime 

for 7 days. Next, the percentage of the superficial hardness recovery (%SHR) 

and integrated loss of subsurface hardness (ΔKHN) were determined. Data 

were submitted to ANOVA and Fisher LSD’s test (p<0.05). A dose-response 

relationship was observed between F concentrations in solutions without HMP 

and the variables %SHR and ΔKHN; as for the solutions containing HMP alone, 

a dose-response relationship was only observed for ΔKHN. Regarding %SHR, 

no significant differences were observed Placebo and 0.5% HMP groups, nor 

between 0.75% and 1% HMP groups. When associated with F, HMP was 

shown to increase the remineralizing capacity of the solutions both at the 

surface and the subsurface of dentin specimens, since the groups containing F 

+ HMP showed significantly superior results compared to groups containing F 

alone. In addition, the solution containing 250 ppm F + 0.5% HMP promoted a 

remineralizing effect similar to that containing 500 ppm F. Regarding ∆KHN, 

significant differences were observed among all groups in the treated area, 

while no significant differences were observed among the groups in the control 

and demineralized areas. The results allowed the conclusion that the addition of 



Abstract 

Letícia Cabrera Capalbo 

HMP to fluoridated solutions significantly enhanced their remineralizing potential 

on dentin artificial caries lesions, both at the surface and in depth. 

 
Keywords: Dentin. Fluoride. Phosphate. Tooth Remineralization. 



 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 
 

Lista de abreviaturas e 
símbolos 

__________________________________________________________



Listas de abreviaturas e símbolos 

 
Letícia Cabrera Capalbo 

 

LISTA DE ABREVIATURAS E SÍMBOLOS 
 

ANOVA Analysis of Variance/Análise de Variância  

Ca+2 Calcium/Cálcio 

CaCl2 Calcium Chloride/Cloreto de cálcio 

CaF2 Calcium Fluoride/Fluoreto de cálcio 

Ca(OH)2 Calcium Hydroxide/Hidróxido de cálcio  

Ca(NO3)2 Calcium Nitrate/Nitrato de cálcio 

°C Degrees Celsius/Graus Celsius 

F Fluoride/Fluoreto 

g  Gram/Grama 

H2O Water/Água 

HMP Sodium hexametaphosphate/Hexametafosfato de sódio 

KCL Potassium Chloride/Cloreto de potássio 

KHN Knoop Hardness Number/Número de Dureza Knoop 

KH2PO4 Potassium dihydrogenphosphate/Fosfato Monopotássico 

L Liter/Litro 

Log10 Logarithm, base 10/Logaritmo na base 10 

mL Milliliter/Mililitro 

M  Molar 

Mm Millimolar/Mili Molar 

mm Millimeter/Milímetro 

mg Milligram/Miligrama 

mmol Milimol 

MMPs Matrix Metalloproteinases/Metaloproteinases da matriz 

NaF Sodium Fluoride/Fluoreto de sódio 

NaH2-PO4 Sodium dihydrogen phosphate/Fosfato Monossódico 

NaN3 Sodium azide/Azida de sódio 

μm  Micrometer/Micrometro 

μM  Micromolar/Micro molar 

p  Probability/Probabilidade 

pH Hydrogenionic Potential /Potencial Hidrogeniônico 

PO4
-3

 Phosphate/Fosfato 

Ppm Parts per million/Partes por milhão 



Listas de abreviaturas e símbolos 

 
Letícia Cabrera Capalbo 

SD Standard Deviation/Desvio padrão 

SH  Surface hardness 

s Seconds/segundos 

TISAB 
Total Ionic Strenght Adjustment Buffer/Tampão de Ajuste da 

Força Iônica Total 

TMP Sodium Trimetaphosphate/Trimetafosfato de sódio 

UNESP São Paulo State University/Universidade Estadual Paulista  

ΔKHN 
Integrated subsurface hardness/Dureza integrada de 

subsuperfície 

%SHR Percentage of surface hardness recovery 

%RDS Porcentagem de recuperação de Dureza de superfície 

  

  

  

  

  

  

  

  

  

  

  

	  

  

 



 

 

 

 

 

 

 

 
 
 
 
 
 
 
 
 

Sumário 
_____________________



Sumário 

Letícia Cabrera Capalbo 

SUMÁRIO 

 

1. ABSTRACT .................................................................................................... 23 
2. INTRODUCTION …………………………………………………………………… 24 
3. MATERIALS AND METHODS ……………………………………………………... 26 
4. RESULTS …………………………………………………………………………. 29 
5. DISCUSSION ……………………………………………………………………… 30 
6. REFERENCES ................................................................................................ 35 
  
7. ANEXOS  
7.1 ANEXO A .................................................................................................... 38 
7.2 ANEXO B .................................................................................................... 53 
7.3 ANEXO C .................................................................................................... 54 
7.4 ANEXO D .................................................................................................... 55 
7.5 ANEXO E .................................................................................................... 56 
7.6 ANEXO F .................................................................................................... 57 
7.7 ANEXO G .................................................................................................... 58 
7.8 ANEXO H .................................................................................................... 59 

  
  
  
  
  
  
  
  
  
  
  
  



 

Letícia Cabrera Capalbo 
 

In vitro analysis of the capacity of solutions containing fluoride and/or 
hexametaphosphate on the remineralization of dentin 

  
LC Capalbo1, ACB Delbem1, GP Nunes1, MAR Buzalaf2, JP Pessan1 

 

1Department of Preventive and Restorative Dentistry, School of Dentistry, 

Araçatuba, São Paulo State University (UNESP), Araçatuba, SP, Brazil  
2Department of Biological Sciences, Bauru School of Dentistry, University of 

São Paulo, Bauru, SP, Brazil 

Short title: Solutions containing F and/or HMP on dentin remineralization. 

 

Keywords: Dentin. Fluoride. Phosphate. Tooth Remineralization. Dental caries.  

 

Corresponding author: 
Juliano Pelim Pessan 

School of Dentistry, Araçatuba, São Paulo State University (UNESP) 

Department of Preventive and Restorative Dentistry 

Rua José Bonifácio 1193 

16015-050 Araçatuba - SP - Brazil 

Tel: (+55) 18 3636 3314              

Email: juliano.pessan@unesp.br 

 

Declaration of interest 

The authors declare no conflict of interest that may affect the manuscript 

judgment. 

  



 

Letícia Cabrera Capalbo 
 

In vitro analysis of the capacity of solutions containing fluoride and/or 
hexametaphosphate on the remineralization of dentin 

 

Abstract 

Objective: to investigate the ability of solutions containing HMP and F, alone or 

in association, in inducing dentin remineralization in vitro. Methods: Root dentin 

blocks (4 × 6 cm, n=100) were prepared, and caries-like lesions were induced in 

2/3 of the surface; each block served as its own control. Blocks were divided 

into 10 experimental groups (n = 10 / group), according to the solutions to be 

tested: Placebo (without F or HMP); 0.5% HMP; 0.75% HMP; 1% HMP; 250 

ppm F; 500 ppm F; 1100 ppm F; 250 ppm F + 0.5% HMP; 500 ppm F + 0.75% 

HMP; and 1100 ppm F + 1% HMP. Specimens were treated for one minute, 

twice a day, with the respective solutions, and submitted to a pH-cycling 

regimen for 7 days. Next, the percentage of the surface hardness recovery 

(%SHR) and integrated area of subsurface hardness (ΔKHN) were determined. 

Data were submitted to ANOVA and Fisher LSD’s test (p<0.05). Results: A 

dose-response relationship was observed between F concentrations in 

solutions without HMP and %SHR and ∆KHN. For both response variables, 

solutions containing F + HMP promoted a significantly higher remineralizing 

effect compared to groups containing F alone. Also, Groups treated with 500 

ppm F or 250 ppm F + 0.5% HMP were not significantly different regarding 

%SHR. Conclusion: the addition of HMP to fluoridated solutions significantly 

enhanced their remineralizing potential on dentine artificial caries lesions in 

vitro, both at the surface and in depth. 

 

Clinical Significance: The simultaneous use of fluoride and HMP may be a 

promising alternative for remineralizing dentine caries lesions in individuals of 

all ages. 

 

 



 

 
Letícia Cabrera Capalbo 

 

24 

1. Introduction  

 Dental caries is a complex, multifactorial disease, being biofilm-sucrose-

dependent [1,2]. It is caused by bacterial acid production through the 

metabolisms of fermentable carbohydrates in the dental biofilm [3], and results 

from successive demineralization and remineralization cycles at the dental hard 

tissues [4]. When demineralization prevails, mineral loss leads to the formation 

of a subsurface lesion. As this process evolves, the lesion may progress, 

compromising the structural integrity of the enamel (chemical process) until it 

reaches the underlying dentin (chemical and biological process), which requires 

detailed knowledge of this tissue’s capacity to resist to mineral losses. In 

addition to carious lesion at coronal dentin (a more advanced process involving 

tooth cavitation), root dentin may also be afflicted by dental caries starting from 

a subsurface lesion, similar to enamel. Dentin has smaller hydroxyapatite 

crystals compared to enamel, is formed in a collagen matrix, and has a higher 

carbonate content, which explains its greater solubility. Thus, during the 

formation of a dentin caries lesion, both processes of mineral dissolution and 

protein degradation take place [5]. 

 The prevention and treatment of initial dental caries lesions, especially in 

high-risk patients, is a constant challenge in the clinical practice. Efforts have 

been directed towards the search for technological advances that promote the 

remineralization of carious lesions, as well as reverse the caries process at the 

earliest possible stage [6]. The administration of fluoride (F) in community-

based strategies, as well as in vehicles for self- and professional application, is 

a well-established strategy for the control of dental caries, since it acts both in 

reducing demineralization, and promoting tissue remineralization [7,8]. 

 In the dentin, this process becomes even more complex ought to tissue 

heterogeneity. Thereby, two main strategies (focused on the de-/re-

mineralization and the enzymatic inhibition) have been extensively investigated. 

The increase in the therapeutic effects of fluoridated products on mineral 

dynamics can potentially improve its effects on the control of dental caries, and 

the addition of organic or inorganic phosphate salts to fluoridated products is an 

effective alternative to increase their effectiveness on the progression of enamel 



 

 
Letícia Cabrera Capalbo 

 

25 

caries lesions, according to in vitro and in situ studies [9–11]. Simultaneously, 

new strategies aimed at the preservation of collagen fibrils and the promotion of 

dentin remineralization have been proposed, in a process known as 

biomimetics [12]. This approach is mediated by specific bioactive agents that 

improve and reinforce dentin through localized changes in biochemical and 

biomechanical properties [13]. The viability of an agent that could reconcile 

these two properties could optimize both the desired results, enabling its use in 

clinical practice.  

 Sodium trimetaphosphate (TMP) and sodium hexametaphosphate (HMP) 

are cyclic inorganic phosphates that have been added to fluoridated products 

such as toothpastes, gels, varnishes, and mouthwash solutions [10,11,14–17] 

to increase their ability in reducing enamel demineralization and enhancing its 

remineralization. It has recently been shown that treatment of dentin artificial 

caries lesions with 1,5% TMP in association with Ca(OH)2 was successful in 

promoting remineralization [18]. However, contradictorily to the evidence of the 

role of TMP and F on enamel caries (when co-administered), this association 

has been shown to be ineffective in the remineralization of dentin lesions [19], 

which may be associated with use of an unfavorable F:TMP molar ratio.  

 As for HMP, it presents a high capacity to adsorb to enamel and to 

reduce its solubility [10], in addition to having a strong tendency to form 

complexes with cations [20,21]. This leads to its precipitation on rich electron-

donor sites on the enamel surface, which promotes greater adsorption of Ca2+ 

and PO43- ions [22]. Considering the promising results of the association of 

HMP with F on enamel de- and re-mineralization, it would be interesting to 

evaluate the effect of this association on dentine caries lesions. Based on the 

similarity of enamel and dentin structures (i.e., mineralized dental tissues), it is 

possible that the enhanced remineralizing potential of F and HMP observed for 

enamel might also be verified for dentin. Such effects and their extent, however, 

remain unknown. 

 Thus, this study aimed to investigate the capacity of solutions containing 

HMP and F, alone or in association, to induce dentin remineralization in an in 

vitro protocol. The study’s null hypothesis was that the use of the test solutions 



 

 
Letícia Cabrera Capalbo 

 

26 

(containing F and HMP) would not lead to different remineralization rates 

compared with their respective controls (containing F alone). 

 

2. Material and Methods 

2.1. Experimental Design  

 Sample size was based on a previous pilot study, considering the 

primary outcome from surface and cross-sectional hardness analysis, in terms 

of the mean difference between groups (10 and 163.5, respectively), standard 

deviation (5.2 and 85.3, respectively), an α-error of 5% and a β-error of 20%. 

Root dentine blocks (6 × 4 × 2 mm, n = 100) were obtained from bovine incisors 

and kept in thymol solution 0.1% for 30 days prior to experimental procedures. 

The surface of the blocks was serially polished, followed by induction of 

subsurface lesions, and the blocks were randomly assigned into 10 groups (n = 

10/group): placebo (without F/HMP); 250 ppm F (250F); 500 ppm F (500F); 

1100 ppm F (1100F); 0.5% HMP; 0.75% HMP; 1% HMP; 250F+0.5% HMP; 

500F+0.75% HMP; 1100F+1% HMP. Blocks were subjected to pH cycling and 

treatment with the solutions. Following, surface and cross-sectional hardness 

were assessed. Blocks remained under moist conditions (deionized water) at 

4ºC prior to hardness analyzes. 

 

2.2. Blocks preparation and caries-like lesion 

 The teeth were cut using an IsoMet Low Speed Saw (Buehler Ltd., Lake 

Bluff, Ill., USA) under water cooling and then serially polished with a grinder 

polisher (Buehler, Lake Bluff, Illinois, USA). The blocks were divided into three 

areas: 

1 – Sound area (positive control area): One-third of the surface was 

covered with acid-resistant varnish to prevent contact with the 

demineralizing solution (caries-like lesions induction) and with the 

treatment solutions; 



 

 
Letícia Cabrera Capalbo 

 

27 

2 – Demineralized area (negative control area): The specimens were 

immersed in demineralizing solution to form a subsurface dentin lesion 

(in the two thirds not covered by acid-resistance varnish). Thereafter, one 

third of this area was also covered with acid-resistant varnish to prevent 

contact with the treatment solutions; 

3 – Treated area (experimental area): After demineralization, the dentin 

surface (one third) was submitted to one of the experimental treatment 

solutions and pH-cycling. 

 To produce dentin subsurface lesions, the blocks were immersed in a 

demineralizing solution (50 mM acetic acid, 2.2 mM CaCl2, 2.2 mM KH2PO4, 

47.6 µM NaF; pH = 5,0 [23]) for 30 minutes, followed by a remineralizing 

solution (1.5 mmol/l Ca(NO3)2 × H2O; 0.9 mmol/l NaH2-PO4 × H2O; 150 mmol/l 

KCl in 0.02 mol/l cacodylic buffer, pH 7.0; 0.05 mg F/ml as NaF [23]) for two 

hours, three times-day, during two days. The blocks were kept in remineralizing 

solution overnight. After this period, the specimens were immersed in the 

demineralizing solution for five additional days, totaling seven days of the 

experimental procedure.  

 

2.3. Treatments and pH-cycling  

 One hundred dentin blocks were randomly divided into 10 experimental 

groups (n = 10/group) according to the treatment solutions, and submitted to a 

7-day pH-cycling regimen at 37°C. The specimens were treated for one minute 

and then immersed in demineralizing solution (1.5 mM CaCl2, 0.9 mM KH2PO4, 

and 50 mM lactic buffer; pH 5.0) for 8 hours. Next, the blocks were treated 

again, followed by immersion in remineralizing solution (5 mM CaCl2, 0.9 mM 

KH2PO4, 130 mM KCl, 20 mM HEPES, and 5 mM NaN3; pH 7.0) for 16 hours 

[18,19,24]. Between all steps, the dentin specimens were rinsed with deionized 

water.  

 

 



 

 
Letícia Cabrera Capalbo 

 

28 

2.4. Dentin hardness analysis 

 After the 7-day pH cycling, surface hardness (SH) was measured using a 

Micromet 5114 hardness tester (Buehler, Lake Bluff, IL, USA) and a Buehler 

OmniMet software (Buehler). A Knoop diamond indenter was used under a 10 g 

load, for 10 seconds [19]. Five indentations were produced in each of the three 

areas with a distance of 100 µm among them, and positioned in the center of 

each area. 

 For cross-sectional hardness (CSH), the dentin blocks were longitudinally 

sectioned, included in acrylic resin with the cut face on the top and the serially 

polished. 14 indentations were performed in each area of the sample at different 

depths (5, 10, 15, 20, 25, 30, 40, 50, 70, 90, 110, 130, 220 and 330 µm) under 

a 2 g load applied for 10 s.  

 Following, the integrated area (cross-sectional profiles of hardness into 

dentine) was calculated using the trapezoidal rule (GraphPad Prism, version 

3.02) in each depth, from the outer surface up to 300 µm in depth, for the three 

areas (sound, demineralized and treated dentine). These values were used for 

the calculation of the integrated recovery of subsurface hardness (∆KHN; KHN 

× µm) [10,25]. 

 

2.5. Statistical Analysis 

 SigmaPlot 12.0 software was used for statistical analysis, and a 5% level 

of significance was set. Data analysis considered the values of %SHR and 

ΔKHN. %SHR (raw) and ∆KHN (log10-transformed) data passed normality 

(Shapiro-Wilk) and homogeneity (Barlett) tests, and were submitted to one-way 

ANOVA and Fisher LSD’s test. Regarding the sound (control) area, ∆KHN data 

were submitted to one-way ANOVA; for the demineralized area, ∆KHN data 

were analyzed by Kruskal-Wallis test.  

 

 

 



 

 
Letícia Cabrera Capalbo 

 

29 

3. Results 

 The present in vitro protocol induced subsurface caries-like lesions in 

dentin with an average depth of 130 µm. At greater depths, hardness values 

approached those of the control area. At the control area (sound dentine), mean 

(SD) hardness values were 72.7 (4.5) and 30.9 (0.5), respectively for SH and 

∆KHN. At the demineralized, the corresponding values were 27.7 (2.3) and 16.8 

(0.4). No significant differences were observed among the groups regarding 

%SHR and ∆KHN, both at the sound (control) and demineralized areas.  

 Table 1 shows %SHR and ∆KHN according to the groups. A dose-

response relationship was observed between F concentrations in the 

experimental solutions without HMP and %SHR and ∆KHN (Placebo < 250 ppm 

F < 500 ppm F < 1100 ppm F). The lowest %SHR values were observed for 

Placebo and all groups treated with HMP alone, which were significantly 

different from the other groups. On the other hand, 500F+0.75% HMP, 1100F, 

and 1100F+1% HMP groups had the highest %SHR values. No significant 

differences were observed between Placebo and 0.5% HMP, 0.75% HMP and 

1% HMP, or 250F+0.5% HMP and 500F.  

 Regarding ∆KHN, significant differences were observed among all 

groups. Placebo and groups containing HMP alone presented the lowest ∆KHN, 

following a dose-response trend between HMP concentrations and the 

remineralizing effects. As for the other groups, a pattern similar to %SHR was 

observed.  

  



 

 
Letícia Cabrera Capalbo 

 

30 

Table 1.  Mean (SD) percentage of surface hardness recovery (%SHR) and integrated 

recovery of subsurface hardness (ΔKNH) according to the groups 

Groups %SHR ΔKHN 

Placebo 9.5 (2.4)a 56.5 (4.1)a 

0.5% HMP 11.9 (3.3)a 87.6 (6.8)b 

0.75% HMP 18.9 (3.8)b 101.2 (7.4)c 

1% HMP 17.8 (3.5)b 133.2 (9.8)d 

250 ppm F 26.2 (2.4)c 370.0 (27.5)e 

500 ppm F 41.2 (2.2)d 533.5 (85.3)f 

1100 ppm F 56.2 (5.2)e 782.1 (58.6)g 

250 ppm F + 0.5% HMP 38.7 (4.7)d 493.4 (38.1)h 

500 ppm F + 0.75% HMP  48.8 (5.0)f 655.4 (42.8)i 

1100 ppm F + 1% HMP 66.6 (5.0)g 859.0 (36.4)j 

Different superscript letters indicate significant differences among the groups within 

each column. One-way ANOVA on the natural outcomes (%SHR) or log10-transformed 

data (ΔKNH), and Fisher LSD’s test (p<0.05, n=10/group). 

 

4. Discussion 

 Approaches to increase the benefits of F, while minimizing its side-

effects, have been intensively investigated, yet with much to be explored. Those 

strategies include the use of polyphosphate salts, nano-hydroxyapatite 

particles, calcium glycerophosphate, amorphous calcium phosphate, and 

functionalized β-tricalcium phosphate [26]. This is the first study evaluating the 

remineralizing potential of F solutions supplemented with HMP in dentin lesions. 

Given that the addition of HMP to F solutions resulted in a higher remineralizing 

potential than their counterparts without HMP, both in surface and in depth, the 

study’s null hypothesis was rejected. 



 

 
Letícia Cabrera Capalbo 

 

31 

 Fluoride is the most widely used agent for controlling root caries, 

especially when applied at high concentrations [27]. It is considered as the gold 

standard for dental caries prevention regarding its ability to prevent dental 

demineralization as well as to promote remineralizing. It has also been shown to 

have the capacity to completely inhibit MMPs activity [28].   Although most of 

the studies on the effects of TMP and HMP were carried out with tooth enamel, 

there is evidence that these salts also have therapeutic properties on dentin. 

Gonçalves et al. [18] demonstrated that 1.5% TMP can act as an effective 

inhibitor of collagen degradation mediated by MMP-2 and MMP-9, as well as 

proteases extracted from healthy dentin. In addition, the treatment of artificial 

dentin caries lesions with 1.5% TMP supplemented with Ca(OH)2 induced their 

remineralization [19]. Surprisingly, however, the association between TMP and 

F was proven ineffective in promoting the remineralization of dentin lesions [19], 

possibly due to the use of an unsuitable F:TMP molar ratio. Therefore, in the 

present study, HMP concentrations of 0.5% and 1% were established based on 

previous studies in which HMP was added to the 250 ppm F and 1100 ppm F 

dentifrices, respectively [10,11,16]. As for the concentration of 0.75% HMP, 

since no study has evaluated the association between HMP and 500 ppm F, it 

was extrapolated from the aforementioned data.  

 In spite of containing phosphate in its structure, cyclic HMP cannot be 

regarded as a source of free phosphate to interact with dental hard tissues, 

considering that the hydrolyzation of its molecule does not spontaneously occur 

under physiological conditions [20,29,30]. In fact, the effect of HMP is 

associated with its capacity to interact with the tooth surfaces. When adsorbed, 

HMP becomes negatively charged, allowing the retention of Ca2+ and CaF+ 

ions, which leads to the formation of a protective layer that restricts acid 

diffusion and increases calcium and fluoride penetration into tooth hard 

tissues[10,16]. In this regard, it has been suggested that this protective layer is 

capable to retain fluoride compounds that can be released when cariogenic 

challenges occur, promoting the formation of more reactive compounds [31]. 

This mechanism has a direct impact on the incorporation of fluoride and calcium 

by enamel and dentine, intensifying remineralization at the subsurface. Previous 



 

 
Letícia Cabrera Capalbo 

 

32 

studies have shown that TMP adsorption and CaF2 deposition on dentin 

produce a synergistic effect and reduce acid diffusion. In turn, these processes 

increase calcium phosphate precipitation and, consequently, protect dentin 

against erosion [32], and promote obliteration of dentine tubules [33]. Although 

no study has investigated the association between F and HMP on the 

deposition of CaF2 in dentin so far, our findings on the remineralizing effects of 

such association might be related to CaF2 deposition. 

 The present study allowed the observation of a clear dose-response 

relationship between F concentrations in the treatment solutions and their 

resulting remineralizing effects; the solution containing 1100 ppm F was 46,6% 

and 112% more effective than those containing 500 and 250 ppm F, 

respectively. Similar rates (51,6% and 102%, respectively) were observed in an 

in vitro study on enamel demineralization [10], thus validating the model 

employed in the present investigation. Nonetheless, the additional benefit of 

HMP on dentine remineralization was much lower than those reported on 

enamel demineralization. While Camara et al. [10] demonstrated that dentifrices 

containing 250 ppm F + 0,5% HMP were 189% and 43% greater than 

dentifrices containing 250 ppm F or 1100 ppm F, respectively, treatment with 

250 ppm F + 0,5% HMP in the present study promoted a remineralizing effects 

33.3% higher than 250 ppm F and 59% lower than that observed for 1100 ppm 

F. Although the reasons for such differences are not apparent, they might be 

due to the factors related to the structural differences between both tissues 

(enamel and dentin), so that further studies are needed to better understand the 

dynamics linking HMP and dentin. 

 Despite significant differences were observed among all groups 

concerning their remineralizing potential at the subsurface, it was noteworthy 

that the addition of HMP to the 250 ppm F solution promoted the greatest 

improvement (33.3%) in relation to its counterpart without HMP; the 

corresponding percentages for solutions containing HMP and fluoride at 500 

ppm F and 1100 ppm F were 22.5% and 9.8%, respectively, compared with 

their counterparts without HMP. Although the causes for such a trend were not 

investigated in the present study, it is possible that the sequestration of Ca2+ 



 

 
Letícia Cabrera Capalbo 

 

33 

and CaF+ by HMP at higher concentrations (negatively influencing the uptake of 

these ions by dentine) might have played a key role on the results observed. 

This reinforces the need for an appropriate HMP/F ratio in order to achieve 

optimum effects on de- and re-mineralization [10].  

 The formation of caries-like lesions in bovine dentin is an important 

approach for the study of new strategies to prevent or treat dentin carious 

lesions [34–38]. Considering the differences between enamel and dentin in 

relation to its matrix, hydroxyapatite crystals size and solubility, the latter is 

especially important from a clinical perspective, as a smaller drop in oral pH 

(around 6.2 – 6.3) is capable of dissolving dentin mineral, while the estimated 

value for enamel solubility is 5.5 [5]. Therefore, dentin is more susceptible to 

demineralizing events than enamel, what hinders direct extrapolation of the 

results obtained for enamel caries to dentine caries. In addition, the degradation 

of the collagen matrix makes the dentin more porous, which allows a greater 

penetration of H+ ions, leading to a more severe mineral loss [32,39]. Another 

important aspect observed was the pattern of the caries-like lesions formed in 

dentin. Despite the hardness averages in longitudinal section at greater depths 

were close to the those in the sound (control) area, even at 330 µm depth it did 

not reach those values, as in enamel models. Once again, this trend might also 

be related to the structural differences between enamel and dentin mentioned 

above, especially the higher porosity of dentine. 

 Despite all precautions to mimic the conditions found in the oral 

environment were taken, the in vitro nature of the present study has some 

inherent limitations related to biological events, including collagen degradation 

and effects of oral microorganisms [40,41]. Also, the high organic content, and 

consequently, the elastic properties of the dentine [38] are factors that can 

influence hardness measurement. Marshall et al. [42] showed that dentin tested 

under hydrated conditions provides more realistic data, which are closer to in 

vivo conditions. Therefore, this approach was adopted in the present 

investigation, in order to better standardize the method of hardness analysis, 

thus providing more reliable data.  



 

 
Letícia Cabrera Capalbo 

 

34 

 Finally, considering the benefits of the association of F and HMP on 

dentin remineralization observed in the present study, such association could 

be assessed in different topical formulations. For daily use, toothpastes and 

mouthwashes must be considered to enhance the remineralization in patients at 

high risk. For clinical practice, formulations with high concentration of F, such as 

gels, foams and varnishes, can also be considered as alternative treatment 

options for initial dentin caries. 

 To sum up, it can be concluded that the addition of HMP to fluorided 

solutions led to enhanced remineralization of artificial caries lesions in dentin, 

both at the surface and in depth. Therefore, considering the promising results of 

the present study in dentin, alongside previous investigations regarding the 

protective effect of these compounds against enamel erosive wear and enamel 

de- and re-mineralization, the association of F and HMP must be considered as 

a valuable strategy not only for high-risk patients, but also for people in general, 

considering its benefits on both dental caries and enamel erosive wear. 

 

Acknowledgments 

 The study was supported by São Paulo Research Foundation (FAPESP, 

Grant #2019/02354-0) and Coordination for the Improvement of Higher 

Education Personnel (CAPES, scholarship to Letícia Cabrera Capalbo - 

Finance Code 001). 

 

  



 

 
Letícia Cabrera Capalbo 

 

35 

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[27]       L.G. Petersson, The role of fluoride in the preventive management of dentin 
hypersensitivity and root caries, Clinical Oral Investigations. (2013). 
https://doi.org/10.1007/s00784-012-0916-9. 

[28]       M.T. Kato, A. Bolanho, B.L. Zarella, T. Salo, L. Tjäderhane, M.A.R. Buzalaf, Sodium 
fluoride inhibits MMP-2 and MMP-9, Journal of Dental Research. (2014). 
https://doi.org/10.1177/0022034513511820. 

[29]       L. Changgen, L. Yongxin, Selective flotation of scheelite from calcium minerals with 
sodium oleate as a collector and phosphates as modifiers. II. The mechanism of the 
interaction between phosphate modifiers and minerals, International Journal of 
Mineral Processing. (1983). https://doi.org/10.1016/0301-7516(83)90012-1. 

[30]       I.K. Choi, W.W. Wen, R.W. Smith, The effect of a long chain phosphate on the 
adsorption of collectors on kaolinite, Minerals Engineering. (1993). 
https://doi.org/10.1016/0892-6875(93)90096-6. 

[31]       M.M. Manarelli, A.C.B. Delbem, T.D.R. Binhardi, J.P. Pessan, In situ remineralizing 
effect of fluoride varnishes containing sodium trimetaphosphate, Clinical Oral 
Investigations. (2015). https://doi.org/10.1007/s00784-015-1492-6. 

[32]       M. Danelon, J.P. Pessan, K.M. Prado, J.P. Ramos, N.G. Emerenciano, M.J. Moretto, 
C.C.R. Martinhon, A.C.B. Delbem, Protective Effect of Fluoride Varnish Containing 
Trimetaphosphate against Dentin Erosion and Erosion/Abrasion: An in vitro Study, 
Caries Research. 54 (2020) 292–296. https://doi.org/10.1159/000505179. 

[33]       C.O. Favretto, A.C.B. Delbem, J.C.S. Moraes, E.R. Camargo, P.T.A. de Toledo, D. Pedrini, 
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microparticles or nanoparticles, Clinical Oral Investigations. (2018). 
https://doi.org/10.1007/s00784-018-2384-3. 

[34]       B.M. Moron, L.P. Comar, A. Wiegand, W. Buchalla, H. Yu, M.A.R. Buzalaf, A.C. 
Magalhães, Different protocols to produce artificial dentine carious lesions in vitro and 
in situ: Hardness and mineral content correlation, Caries Research. 47 (2013) 162–170. 
https://doi.org/10.1159/000345362. 

[35]       S.H. Okuyama K, Nakata T, Pereira PN, Kawamoto C, Komatsu H, Prevention of artificial 
caries: effect of bonding agent, resin composite and topic fluoride application, 
Operative Dentistry. 31 (2006) 135–142. 

[36]       E. Zaura, M.J. Buijs, J.M. ten Cate, Effects of ozone and sodium hypochlorite on caries-
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[37]       S. Pavan, Q. Xie, A.T. Hara, A.K. Bedran-Russo, Biomimetic approach for root caries 
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Dentine: A Systematic Evaluation of the Method, Caries Research. (2016). 
https://doi.org/10.1159/000448147. 

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[41]       M. Marquezan, F.N.P. Corrêa, M.E. Sanabe, L.E. Rodrigues Filho, J. Hebling, A.C. 
Guedes-Pinto, F.M. Mendes, Artificial methods of dentine caries induction: A hardness 
and morphological comparative study, Archives of Oral Biology. (2009). 
https://doi.org/10.1016/j.archoralbio.2009.09.007. 

[42]       G.W. Marshall, S. Habelitz, R. Gallagher, M. Balooch, G. Balooch, S.J. Marshall, 
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Anexos 
____________________



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5. ANEXOS 
ANEXO A 

JOURNAL OF DENTISTRY 

INSTRUÇÕES AOS AUTORES  

JOURNAL OF DENTISTRY 
 
IMPACT FACTOR 
2016: 3.456 © Thomson Reuters Journal Citation Reports 2017  
INTRODUCTION 
The Journal of Dentistry is the leading international dental journal within the field of 
Restorative Dentistry. Placing an emphasis on publishing novel and high-quality 
research papers, the Journal aims to influence the practice of dentistry at clinician, 
research, industry and policy-maker level on an international basis. 
Topics covered include the management of dental disease, periodontology, 
endodontology, operative dentistry, fixed and removable prosthodontics, and dental 
biomaterials science, long-term clinical trials including epidemiology and oral health, 
dental education, technology transfer of new scientific instrumentation or procedures, 
as well clinically relevant oral biology and translational research. 
Submissions are welcomed from other clinically relevant areas, however, the Journal 
places an emphasis on publishing high-quality and novel research. 
Queries in relation to manuscript content should be directed to the Journal Editorial 
Office in the first instance. 
Submissions 
Authors are requested to submit their original manuscript and figures via the online 
submission and editorial system for Journal of Dentistry. Using this online system, 
authors may submit manuscripts and track their progress through the system to 
publication. Reviewers can download manuscripts and submit their opinions to the 
editor. Editors can manage the whole submission/review/revise/publish process. 
Please register at: http://ees.elsevier.com/jjod 
Types of paper 
Contributions falling into the following categories will be considered for publication:- 
Original Research 
Reports: maximum length 6 printed pages approximately 20 typescript pages, including 
illustrations and tables. 
- Review articles: maximum length 10 printed pages, approximately 33 typescript 
pages, including illustrations and tables. 
- Short communication for rapid publication: maximum length 2 printed pages, 
approximately 7 typescript pages, including illustrations. 
- Letters providing informed comment and constructive criticism of material previously 
published in the Journal. 
All typescripts must be accompanied by a Permission Note. This is a letter signed by 
each author (not just the corresponding author), affirming that the paper has been 
submitted solely to Journal of Dentistry and that it is not concurrently under 
consideration for publication in another journal. 
Prospective authors should confirm that the submitted work, including images, are 
original. Authors are reminded that if included images (e.g. Tables and Figures) have 
been previously published may require copyright permission. 
Please note the Journal of Dentistry does not accept Case Reports and these will be 
removed from the system if submitted. 
Authorship 



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Only those persons who have made a significant contribution to the manuscript 
submitted should be listed as authors. The Editor-in-Chief expects that a manuscript 
should normally have no more than 6 authors, unless a case is made by the 
corresponding author within the article cover letter to include other authors. All of the 
named authors should have been involved in the work leading to the publication of the 
paper and should have read the paper before it is submitted for publication. 
Submission checklist 
You can use this list to carry out a final check of your submission before you send it to 
the journal for review. Please check the relevant section in this Guide for Authors for 
more details. 
Ensure that the following items are present: One author has been designated as the 
corresponding author with contact details: 
• E-mail address 
• Full postal address 
All necessary files have been uploaded: 
Manuscript: 
• Include keywords 
• All figures (include relevant captions) 
• All tables (including titles, description, footnotes) 
• Ensure all figure and table citations in the text match the files provided 
• Indicate clearly if color should be used for any figures in print 
Graphical Abstracts / Highlights files (where applicable) 
Supplemental files (where applicable) 
Further considerations 
• Manuscript has been 'spell checked' and 'grammar checked' 
• All references mentioned in the Reference List are cited in the text, and vice versa 
• Permission has been obtained for use of copyrighted material from other sources 
(including the Internet) 
• A competing interests statement is provided, even if the authors have no competing 
interests to declare 
• Journal policies detailed in this guide have been reviewed 
• Referee suggestions and contact details provided, based on journal requirements. 
For further information, visit our Support Center. 
Ethics in publishing 
Please see our information pages on Ethics in publishing and Ethical guidelines for 
journal publication. 
Human and animal rights 
If the work involves the use of human subjects, the author should ensure that the work 
described has been carried out in accordance with The Code of Ethics of the World 
Medical Association (Declaration of Helsinki) for experiments involving humans; 
Uniform Requirements for manuscripts submitted to Biomedical journals. Authors 
should include a statement in the manuscript that informed consent was obtained for 
experimentation with human subjects. The privacy rights of human subjects must 
always be observed. 
All animal experiments should comply with the ARRIVE guidelines and should be 
carried out in accordance with the U.K. Animals (Scientific Procedures) Act, 1986 and 
associated guidelines, EU Directive 2010/63/EU for animal experiments, or the 
National Institutes of Health guide for the care and use of Laboratory animals (NIH 
Publications No. 8023, revised 1978) and the authors should clearly indicate in the 
manuscript that such guidelines have been followed. 
Declaration of interest 
All authors must disclose any financial and personal relationships with other people or 
organizations that could inappropriately influence (bias) their work. Examples of 



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potential conflicts of interest include employment, consultancies, stock ownership, 
honoraria, paid expert testimony, patent applications/registrations, and grants or other 
funding. If there are no conflicts of interest then please state this: 'Conflicts of interest: 
none'. More information. 
Submission declaration and verification 
Submission of an article implies that the work described has not been published 
previously (except in the form of an abstract or as part of a published lecture or 
academic thesis or as an electronic preprint, see 'Multiple, redundant or concurrent 
publication' section of our ethics policy for more information), that it is not under 
consideration for publication elsewhere, that its publication is approved by all authors 
and tacitly or explicitly by the responsible authorities where the work was carried out, 
and that, if accepted, it will not be published elsewhere in the same form, in English or 
in any other language, including electronically without the written consent of the 
copyright-holder. To verify originality, your article may be checked by the originality 
detection service Cross Check. 
Changes to authorship 
Authors are expected to consider carefully the list and order of authors before 
submitting their manuscript and provide the definitive list of authors at the time of the 
original submission. Any addition, deletion or rearrangement of author names in the 
authorship list should be made only before the manuscript has been accepted and only 
if approved by the journal Editor. To request such a change, the Editor must receive 
the following from the corresponding author: (a) the reason for the change in author list 
and (b) written confirmation (e-mail, letter) from all authors that they agree with the 
addition, removal or rearrangement. In the case of addition or removal of authors, this 
includes confirmation from the author being added or removed. Only in exceptional 
circumstances will the Editor consider the addition, deletion or rearrangement of 
authors after the manuscript has been accepted. While the Editor considers the 
request, publication of the manuscript will be suspended. If the manuscript has already 
been published in an online issue, any requests approved by the Editor will result in a 
corrigendum. 
Clinical trial results 
In line with the position of the International Committee of Medical Journal Editors, the 
journal will not consider results posted in the same clinical trials registry in which 
primary registration resides to be prior publication if the results posted are presented in 
the form of a brief structured (less than 500 words) abstract or table. However, 
divulging results in other circumstances (e.g., investors' meetings) is discouraged and 
may jeopardise consideration of the manuscript. Authors should fully disclose all 
posting in registries of results of the same or closely related work. 
Reporting clinical trials Randomized controlled trials should be presented according to 
the CONSORT guidelines. At manuscript submission, authors must provide the 
CONSORT checklist accompanied by a flow diagram that illustrates the progress of 
patients through the trial, including recruitment, enrollment, randomization, withdrawal 
and completion, and a detailed description of the randomization procedure. The 
CONSORT checklist and template flow diagram are available online. 
Registration of clinical trials 
Registration in a public trials registry is a condition for publication of clinical trials in this 
journal in accordance with International Committee of Medical Journal Editors 
recommendations. Trials must register at or before the onset of patient enrolment. The 
clinical trial registration number should be included at the end of the abstract of the 
article. A clinical trial is defined as any research study that prospectively assigns 
human participants or groups of humans to one or more health-related interventions to 
evaluate the effects of health outcomes. Health-related interventions include any 
intervention used to modify a biomedical or health-related outcome (for example drugs, 



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surgical procedures, devices, behavioural treatments, dietary interventions, and 
process-of-care changes). Health outcomes include any biomedical or health-related 
measures obtained in patients or participants, including pharmacokinetic measures and 
adverse events. Purely observational studies (those in which the assignment of the 
medical intervention is not at the discretion of the investigator) will not require 
registration. 
Article transfer service This journal is part of our Article Transfer Service. This means 
that if the Editor feels your article is more suitable in one of our other participating 
journals, then you may be asked to consider transferring the article to one of those. If 
you agree, your article will be transferred automatically on your behalf with no need to 
reformat. Please note that your article will be reviewed again by the new journal. 
More information. 
Upon acceptance of an article, authors will be asked to complete a 'Journal Publishing 
Agreement' (see more information on this). An e-mail will be sent to the corresponding 
author confirming receipt of the manuscript together with a 'Journal Publishing 
Agreement' form or a link to the online version of this agreement. 
Subscribers may reproduce tables of contents or prepare lists of articles including 
abstracts for internal circulation within their institutions. Permission of the Publisher is 
required for resale or distribution outside the institution and for all other derivative 
works, including compilations and translations. If excerpts from other copyrighted works 
are included, the author(s) must obtain written permission from the copyright owners 
and credit the source(s) in the article. Elsevier has preprinted forms for use by authors 
in these cases. 
For open access articles: Upon acceptance of an article, authors will be asked to 
complete an 'Exclusive License Agreement' (more information). Permitted third party 
reuse of open access articles is determined by the author's choice of user license. 
Author rights 
As an author you (or your employer or institution) have certain rights to reuse your 
work. More information. 
Elsevier supports responsible sharing  
Find out how you can share your research published in Elsevier journals. 
Role of the funding source 
You are requested to identify who provided financial support for the conduct of the 
research and/or preparation of the article and to briefly describe the role of the 
sponsor(s), if any, in study design; in the collection, analysis and interpretation of data; 
in the writing of the report; and in the decision to submit the article for publication. If the 
funding source(s) had no such involvement then this should be stated. 
Funding body agreements and policies Elsevier has established a number of 
agreements with funding bodies which allow authors to comply with their funder's open 
access policies. Some funding bodies will reimburse the author for the Open Access 
Publication Fee. Details of existing agreements are available online. 
After acceptance, open access papers will be published under a noncommercial 
license. For authors requiring a commercial CC BY license, you can apply after your 
manuscript is accepted for publication. 
Open access 
This journal offers authors a choice in publishing their research: 
Subscription 
• Articles are made available to subscribers as well as developing countries and patient 
groups through our universal access programs. 
• No open access publication fee payable by authors. 
Open access 
• Articles are freely available to both subscribers and the wider public with permitted 
reuse. 



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• An open access publication fee is payable by authors or on their behalf, e.g. by their 
research funder or institution. 
Regardless of how you choose to publish your article, the journal will apply the same 
peer review criteria and acceptance standards. 
For open access articles, per permitted third party (re)use is defined by the following 
Creative Commons user licenses: 
Creative Commons Attribution-NonCommercial-NoDerivs (CC BY-NC-ND) 
For non-commercial purposes, lets others distribute and copy the article, and to include 
in a collective work (such as an anthology), as long as they credit the author(s) and 
provided they do not alter or modify the article. The open access publication fee for this 
journal is USD 2200, excluding taxes. Learn more about Elsevier's pricing policy: 
http://www.elsevier.com/openaccesspricing. 
Green open access 
Authors can share their research in a variety of different ways and Elsevier has a 
number of green open access options available. We recommend authors see our green 
open access page for further information. Authors can also self-archive their 
manuscripts immediately and enable public access from their institution's repository 
after an embargo period. This is the version that has been accepted for publication and 
which typically includes author-incorporated changes suggested during submission, 
peer review and in editor-author communications. Embargo period: For subscription 
articles, an appropriate amount of time is needed for journals to deliver value to 
subscribing customers before an article becomes freely available to the public. This is 
the embargo period and it begins from the date the article is formally published online 
in its final and fully citable form. Find out more. This journal has an embargo period of 
12 months.  
Elsevier Publishing Campus 
The Elsevier Publishing Campus (www.publishingcampus.com) is an online platform 
offering free lectures, interactive training and professional advice to support you in 
publishing your research. The College of Skills training offers modules on how to 
prepare, write and structure your article and explains how editors will look at your paper 
when it is submitted for publication. Use these resources, and more, to ensure that 
your submission will be the best that you can make it. 
Language (usage and editing services) 
Please write your text in good English (American or British usage is accepted, but not a 
mixture of these). Authors who feel their English language manuscript may require 
editing to eliminate possible grammatical or spelling errors and to conform to correct 
scientific English may wish to use the English Language Editing service available from 
Elsevier's WebShop. 
Submission 
Our online submission system guides you stepwise through the process of entering 
your article details and uploading your files. The system converts your article files to a 
single PDF file used in the peer-review process. Editable files (e.g., Word, LaTeX) are 
required to typeset your article for final publication. All correspondence, including 
notification of the Editor's decision and requests for revision, is sent by e-mail. 
Submit your article 
Please submit your article via http://ees.elsevier.com/jjod. 
Referees 
Please submit the names and institutional e-mail addresses of several potential 
referees. For more details, visit our Support site. Note that the editor retains the sole 
right to decide whether or not the suggested reviewers are used. 
PREPARATION 
Peer review 



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This journal operates a double blind review process. All contributions will be initially 
assessed by the editor for suitability for the journal. Papers deemed suitable are then 
typically sent to a minimum of two independent expert reviewers to assess the scientific 
quality of the paper. The Editor is responsible for the final decision regarding 
acceptance or rejection of articles. The Editor's decision is final. More information on 
types of peer review. 
Use of word processing software 
It is important that the file be saved in the native format of the word processor used. 
The text should be in single-column format. Keep the layout of the text as simple as 
possible. Most formatting codes will be removed and replaced on processing the 
article. In particular, do not use the word processor's options to justify text or to 
hyphenate words. However, do use bold face, italics, subscripts, superscripts etc. 
When preparing tables, if you are using a table grid, use only one grid for each 
individual table and not a grid for each row. If no grid is used, use tabs, not spaces, to 
align columns. The electronic text should be prepared in a way very similar to that of 
conventional manuscripts (see also the Guide to Publishing with Elsevier). Note that 
source files of figures, tables and text graphics will be required whether or not you 
embed your figures in the text. See also the section on Electronic artwork. 
To avoid unnecessary errors you are strongly advised to use the 'spell-check' and 
'grammar-check' functions of your word processor. 
Introduction 
The introduction must be presented in a structured format, covering the following 
subjects, although not under subheadings: succinct statements of the issue in question, 
and the essence of existing knowledge and understanding pertinent to the issue. In 
keeping with the house style of Journal of Dentistry, the final paragraph of the 
introduction should clearly state the aims and/or objective of the work being reported. 
Prospective authors may find the following form of words to be helpful: "The aim of this 
paper is to ..." Where appropriate, a hypothesis (e.g. null or a priori) should then be 
stated. Essential title page information 
• Title. Concise and informative. Titles are often used in information-retrieval systems. 
Avoid abbreviations and formulae where possible. 
• Author names and affiliations. Please clearly indicate the given name(s) and family 
name(s) of each author and check that all names are accurately spelled. Present the 
authors' affiliation addresses (where the actual work was done) below the names. 
Indicate all affiliations with a lowercase superscript letter immediately after the author's 
name and in front of the appropriate address. Provide the full postal address of each 
affiliation, including the country name and, if available, the 
e-mail address of each author. 
• Corresponding author. Clearly indicate who will handle correspondence at all stages 
of refereeing and publication, also post-publication. Ensure that the e-mail address is 
given and that contact details are kept up to date by the corresponding author. 
• Present/permanent address. If an author has moved since the work described in the 
article was done, or was visiting at the time, a 'Present address' (or 'Permanent 
address') may be indicated as a footnote to that author's name. The address at which 
the author actually did the work must be retained as the main, affiliation address. 
Superscript Arabic numerals are used for such footnotes. 
The title page should contain the following information: 
- Title of paper 
- Short title 
- Name(s), job titles and address(es) of author(s) (no academic degrees necessary) 
- Name, address, telephone, fax and e-mail of the corresponding author 
- Up to 6 keywords 
Spelling: International English. 



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Authors are urged to write as concisely as possible. 
The house style of Journal of Dentistry requires that articles should be arranged in the 
following order: Title, Abstract, Introduction, Materials and Methods, Results, 
Discussion, Conclusions, Acknowledgements, References, Tables, Figures. A cover 
letter should accompany the new manuscript submission, within which the authors 
should indicate the significance of the work being submitted in a statement no more 
than 100 words. A signed permission note (details below) must also be included. 
Abstract: should not exceed 250 words and should be presented under the following 
subheadings: 
Objectives, Methods; Results; Conclusions (For Reviews: Objectives; Data; Sources; 
Study selection; Conclusions). A 50 word 'Clinical Significance' statement should 
appear at the end of the abstract advising readers of the clinical importance and 
relevance of their work. These subheadings should appear in the text of the abstract. 
Please repeat the title of the article at the top of the abstract page. 
Introduction: must be presented in a structured format, covering the following subjects, 
although not under subheadings: succinct statements of the issue in question, and the 
essence of existing knowledge and understanding pertinent to the issue. In keeping 
with the house style of Journal of Dentistry, the final paragraph of the introduction 
should clearly state the aims and/or objective of the work being reported. Prospective 
authors may find the following form of words to be helpful: "The aim of this paper is to 
..." Where appropriate, a hypothesis (e.g. null or a priori) should then be stated. 
Keywords: up to 6 keywords should be supplied. 
Abbreviations and acronyms: terms and names to be referred to in the form of 
abbreviations or acronyms must be given in full when first mentioned. Units: SI units 
should be used throughout. If non-SI units must be quoted, the SI equivalent must 
immediately follow in parentheses. The complete names of individual teeth must be 
given in the test. In tables and legends for illustrations individual teeth should be 
identified using the FDI two-digit system. 
Statistics 
Statistical methods should be described with enough detail to enable a knowledgeable 
reader with access to the original data to verify the reported results. When possible, 
findings should be quantified and appropriate measures of error or uncertainty (such as 
confidence intervals) given. Details about eligibility criteria for subjects, randomization 
and the number of observations should be included. The computer software and the 
statistical method(s) used should be specified with references to standard works when 
possible (with pages specified). See http://www.icmje.org/manuscript_1prepare.html for 
more detailed guidelines. 
Illustrations: should be submitted electronically using appropriate commercial software. 
Prospective authors should follow the relevant guidelines (available from: 
http://www.elsevier.com/ artworkinstructions). In addition, it is noted that while authors 
sometimes need to manipulate images for clarity, manipulation for purposes of 
deception or fraud will be seen as scientific ethical abuse and will be dealt with 
accordingly. For graphical images, journals published by Elsevier apply the following 
policy: no specific feature within an image may be enhanced, obscured, moved, 
removed, or introduced. Adjustments of brightness, contrast, or color balance are 
acceptable if and as long as they do not obscure or eliminate any information present 
in the original. Nonlinear adjustments (e.g. changes to gamma settings) must be 
disclosed in the figure legend. 
Abstract 
The Abstract should not exceed 250 words and should be presented under the 
following subheadings: 
Objectives, Methods; Results; Conclusions. A 50 word 'Clinical Significance' statement 
should appear at the end of the abstract advising readers of the clinical importance and 



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relevance of their work. These subheadings should appear in the text of the abstract. 
Please repeat the title of the article at the top of the abstract page. For Review Articles 
the abstract should be presented under the following subheadings: Objectives; Data; 
Sources; Study selection; Conclusions. 
Graphical abstract 
Although a graphical abstract is optional, its use is encouraged as it draws more 
attention to the online article. The graphical abstract should summarize the contents of 
the article in a concise, pictorial form designed to capture the attention of a wide 
readership. Graphical abstracts should be submitted as a separate file in the online 
submission system. Image size: Please provide an image with a minimum of 531 × 
1328 pixels (h × w) or proportionally more. The image should be readable at a size of 5 
× 13 cm using a regular screen resolution of 96 dpi. Preferred file types: TIFF, EPS, 
PDF or MS Office files. You can view Example Graphical Abstracts on our information 
site. Authors can make use of Elsevier's Illustration Services to ensure the best 
presentation of their images and in accordance with all technical requirements. 
Keywords 
Provide a maximum of 6 keywords, using British spelling and avoiding general and 
plural terms and multiple concepts (avoid, for example, 'and', 'of'). Be sparing with 
abbreviations: only abbreviations firmly established in the field may be eligible. These 
keywords will be used for indexing purposes. 
Formatting of funding sources 
List funding sources in this standard way to facilitate compliance to funder's 
requirements: 
Funding: This work was supported by the National Institutes of Health [grant numbers 
xxxx, yyyy]; the Bill & Melinda Gates Foundation, Seattle, WA [grant number zzzz]; and 
the United States Institutes of Peace [grant number aaaa]. It is not necessary to include 
detailed descriptions on the program or type of grants and awards. When funding is 
from a block grant or other resources available to a university, college, or other 
research institution, submit the name of the institute or organization that provided the 
funding. If no funding has been provided for the research, please include the following 
sentence: 
This research did not receive any specific grant from funding agencies in the public, 
commercial, or not-for-profit sectors. 
Artwork 
Image manipulation 
Whilst it is accepted that authors sometimes need to manipulate images for clarity, 
manipulation for purposes of deception or fraud will be seen as scientific ethical abuse 
and will be dealt with accordingly. 
For graphical images, this journal is applying the following policy: no specific feature 
within an image may be enhanced, obscured, moved, removed, or introduced. 
Adjustments of brightness, contrast, or color balance are acceptable if and as long as 
they do not obscure or eliminate any information present in the original. Nonlinear 
adjustments (e.g. changes to gamma settings) must be disclosed in the figure legend. 
Electronic artwork 
General points 
• Make sure you use uniform lettering and sizing of your original artwork. 
• Embed the used fonts if the application provides that option. 
• Aim to use the following fonts in your illustrations: Arial, Courier, Times New Roman, 
Symbol, or use fonts that look similar. 
• Number the illustrations according to their sequence in the text. 
• Use a logical naming convention for your artwork files. 
• Provide captions to illustrations separately. 
• Size the illustrations close to the desired dimensions of the published version. 



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• Submit each illustration as a separate file. 
A detailed guide on electronic artwork is available. You are urged to visit this site; some 
excerpts from the detailed information are given here. 
Formats 
If your electronic artwork is created in a Microsoft Office application (Word, PowerPoint, 
Excel) then please supply 'as is' in the native document format. Regardless of the 
application used other than Microsoft Office, when your electronic artwork is finalized, 
please 'Save as' or convert the images to one of the following formats (note the 
resolution requirements for line drawings, halftones, and line/halftone combinations 
given below): 
EPS (or PDF): Vector drawings, embed all used fonts. 
TIFF (or JPEG): Color or grayscale photographs (halftones), keep to a minimum of 300 
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minimum of 1000 dpi. 
TIFF (or JPEG): Combinations bitmapped line/half-tone (color or grayscale), keep to a 
minimum of 500 dpi. 
Please do not: 
• Supply files that are optimized for screen use (e.g., GIF, BMP, PICT, WPG); these 
typically have a low number of pixels and limited set of colors; 
• Supply files that are too low in resolution; 
• Submit graphics that are disproportionately large for the content. 
Color artwork 
Please make sure that artwork files are in an acceptable format (TIFF (or JPEG), EPS 
(or PDF), or MS Office files) and with the correct resolution. If, together with your 
accepted article, you submit usable color figures then Elsevier will ensure, at no 
additional charge, that these figures will appear in color online (e.g., ScienceDirect and 
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regarding the costs from Elsevier after receipt of your accepted article. Please indicate 
your preference for color: in print or online only. Further information on the preparation 
of electronic artwork. 
Illustration services 
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Please visit the website to find out more. 
Figure captions 
Ensure that each illustration has a caption. Supply captions separately, not attached to 
the figure. A caption should comprise a brief title (not on the figure itself) and a 
description of the illustration. Keep text in the illustrations themselves to a minimum but 
explain all symbols and abbreviations used. 
Tables 
Please submit tables as editable text and not as images. Tables can be placed either 
next to the relevant text in the article, or on separate page(s) at the end. Number tables 
consecutively in accordance with their appearance in the text and place any table notes 
below the table body. Be sparing in the use of tables and ensure that the data 
presented in them do not duplicate results described elsewhere in the article. Please 
avoid using vertical rules and shading in table cells. 
References 
Citation in text 



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47 

Please ensure that every reference cited in the text is also present in the reference list 
(and vice versa). Any references cited in the abstract must be given in full. Unpublished 
results and personal communications are not recommended in the reference list, but 
may be mentioned in the text. If these references are included in the reference list they 
should follow the standard reference style of the journal and should include a 
substitution of the publication date with either 'Unpublished results' or 'Personal 
communication'. Citation of a reference as 'in press' implies that the item has been 
accepted for publication. 
Reference links 
Increased discoverability of research and high quality peer review are ensured by 
online links to the sources cited. In order to allow us to create links to abstracting and 
indexing services, such as Scopus, CrossRef and PubMed, please ensure that data 
provided in the references are correct. Please note that incorrect surnames, 
journal/book titles, publication year and pagination may prevent link creation. When 
copying references, please be careful as they may already contain errors. Use of the 
DOI is encouraged. A DOI can be used to cite and link to electronic articles where an 
article is in-press and full citation details are not yet known, but the article is available 
online. A DOI is guaranteed never to change, so you can use it as a permanent link to 
any electronic article. An example of a citation using DOI for an article not yet in an 
issue is: VanDecar J.C., Russo R.M., James D.E., Ambeh W.B., Franke M. (2003). 
Aseismic continuation of the Lesser Antilles slab beneath northeastern Venezuela. 
Journal of Geophysical Research, https://doi.org/10.1029/2001JB000884. Please note 
the format of such citations should be in the same style as all other references in the 
paper. 
Web references 
As a minimum, the full URL should be given and the date when the reference was last 
accessed. Any further information, if known (DOI, author names, dates, reference to a 
source publication, etc.), should also be given. Web references can be listed separately 
(e.g., after the reference list) under a different heading if desired, or can be included in 
the reference list. 
Data references 
This journal encourages you to cite underlying or relevant datasets in your manuscript 
by citing them in your text and including a data reference in your Reference List. Data 
references should include the following elements: author name(s), dataset title, data 
repository, version (where available), year, and global persistent identifier. Add 
[dataset] immediately before the reference so we can properly identify it as a data 
reference. The [dataset] identifier will not appear in your published article. 
References in a special issue 
Please ensure that the words 'this issue' are added to any references in the list (and 
any citations in the text) to other articles in the same Special Issue. 
Citation in text 
Please ensure that every reference cited in the text is also present in the reference list 
(and vice versa). Any references cited in the abstract must be given in full. Unpublished 
results and personal communications are not recommended in the reference list, but 
may be mentioned in the text. If these references are included in the reference list they 
should follow the standard reference style of the journal and should include a 
substitution of the publication date with either 'Unpublished results' or 'Personal 
communication'. Citation of a reference as 'in press' implies that the item has been 
accepted for publication. 
Reference links 
Increased discoverability of research and high quality peer review are ensured by 
online links to the sources cited. In order to allow us to create links to abstracting and 
indexing services, such as Scopus, CrossRef and PubMed, please ensure that data 



Anexos  

 
Letícia Cabrera Capalbo 

 

48 

provided in the references are correct. Please note that incorrect surnames, 
journal/book titles, publication year and pagination may prevent link creation. When 
copying references, please be careful as they may already contain errors. Use of the 
DOI is encouraged. 
A DOI can be used to cite and link to electronic articles where an article is in-press and 
full citation details are not yet known, but the article is available online. A DOI is 
guaranteed never to change, so you can use it as a permanent link to any electronic 
article. An example of a citation using DOI for an article not yet in an issue is: 
VanDecar J.C., Russo R.M., James D.E., Ambeh W.B., Franke M. 
(2003). Aseismic continuation of the Lesser Antilles slab beneath northeastern 
Venezuela. Journal of Geophysical Research, https://doi.org/10.1029/2001JB000884. 
Please note the format of such citations should be in the same style as all other 
references in the paper. 
Web references 
As a minimum, the full URL should be given and the date when the reference was last 
accessed. Any further information, if known (DOI, author names, dates, reference to a 
source publication, etc.), should also be given. Web references can be listed separately 
(e.g., after the reference list) under a different heading if desired, or can be included in 
the reference list. 
Data references 
This journal encourages you to cite underlying or relevant datasets in your manuscript 
by citing them in your text and including a data reference in your Reference List. Data 
references should include the following elements: author name(s), dataset title, data 
repository, version (where available), year, and global persistent identifier. Add 
[dataset] immediately before the reference so we can properly identify it as a data 
reference. The [dataset] identifier will not appear in your published article. 
References in a special issue 
Please ensure that the words 'this issue' are added to any references in the list (and 
any citations in the text) to other articles in the same Special Issue. 
References 
Citation in text 
Please ensure that every reference cited in the text is also present in the reference list 
(and vice versa). Any references cited in the abstract must be given in full. Unpublished 
results and personal communications are not recommended in the reference list, but 
may be mentioned in the text. If these references are included in the reference list they 
should follow the standard reference style of the journal and should include a 
substitution of the publication date with either 'Unpublished results' or 'Personal 
communication'. Citation of a reference as 'in press' implies that the item has been 
accepted for publication. 
Reference links 
Increased discoverability of research and high quality peer review are ensured by 
online links to the sources cited. In order to allow us to create links to abstracting and 
indexing services, such as Scopus, CrossRef and PubMed, please ensure that data 
provided in the references are correct. Please note that incorrect surnames, 
journal/book titles, publication year and pagination may prevent link creation. When 
copying references, please be careful as they may already contain errors. Use of the 
DOI is encouraged. 
A DOI can be used to cite and link to electronic articles where an article is in-press and 
full citation details are not yet known, but the article is available online. A DOI is 
guaranteed never to change, so you can use it as a permanent link to any electronic 
article. An example of a citation using DOI for an article not yet in an issue is: 
VanDecar J.C., Russo R.M., James D.E., Ambeh W.B., Franke M. (2003). Aseismic 
continuation of the Lesser Antilles slab beneath northeastern Venezuela. Journal of 



Anexos  

 
Letícia Cabrera Capalbo 

 

49 

Geophysical Research, https://doi.org/10.1029/2001JB000884. Please note the format 
of such citations should be in the same style as all other references in the paper. 
Web references 
As a minimum, the full URL should be given and the date when the reference was last 
accessed. Any further information, if known (DOI, author names, dates, reference to a 
source publication, etc.), should also be given. Web references can be listed separately 
(e.g., after the reference list) under a different heading if desired, or can be included in 
the reference list. 
Data references 
This journal encourages you to cite underlying or relevant datasets in your manuscript 
by citing them in your text and including a data reference in your Reference List. Data 
references should include the following elements: author name(s), dataset title, data 
repository, version (where available), year, and global persistent identifier. Add 
[dataset] immediately before the reference so we can properly identify it as a data 
reference. The [dataset] identifier will not appear in your published article. 
References in a special issue 
Please ensure that the words 'this issue' are added to any references in the list (and 
any citations in the text) to other articles in the same Special Issue Reference 
management software Most Elsevier journals have their reference template available in 
many of the most popular reference management software products. These include all 
products that support Citation Style Language styles, such as Mendeley and Zotero, as 
well as EndNote. Using the word processor plug-ins from these products, authors only 
need to select the appropriate journal template when preparing their article, after which 
citations and bibliographies will be automatically formatted in the journal's style. If no 
template is yet available for this journal, please follow the format of the sample 
references and citations as shown in this Guide. Users of Mendeley Desktop can easily 
install the reference style for this journal by clicking the following link: 
http://open.mendeley.com/use-citation-style/journal-of-dentistry When preparing your 
manuscript, you will then be able to select this style using th