University of Groningen
Quaternary ammonium compounds to prevent oral biofilm formation
Miura Sugii, Mari
IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it. Please check the document version below.
Document Version
Publisher's PDF, also known as Version of record
Publication date: 2019
Link to publication in University of Groningen/UMCG research database
Citation for published version (APA):
Miura Sugii, M. (2019). Quaternary ammonium compounds to prevent oral biofilm formation.
Copyright
Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons).
Take-down policy
If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.
Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum.
123
General
Discussion
Chapter 6
GENERAL DISCUSSION
From human health point of view many deleterious and financially demanding effects are associated with bacterial colonization [1–6]. Furthermore, the pressure of bacterial resistance is increasing, which makes the need for new alternatives to combat bacterial adhesion and biofilm formation necessary. For dental clinicians, caries is a common disease related to biofilm formation and its prevention or treatment remains a challenge. Biofilm formation surrounding cemented orthodontic devices is a highly prevalent clinical situation. The difficulty in cleaning the complex structure of orthodontic devices associated with the lack of compliance to the brushing routine can promptly lead to biofilm formation hindering treatment prognosis and depleting patient’s oral health. Therefore, the development of antimicrobial materials to address these complications is necessary.
Quaternary ammonium compounds (QAC) are a class of antimicrobials claimed to be effective against Streptococcus mutans, a cariogenic species. QAC can be included in dental materials either to modify a surface, making it contact-killing or to be leached out and being active in the environment [7–9]. The review on QAC for biomedical applications (chapter 2) evidences that QAC antimicrobial activity can be influenced by the presence and length of alkyl chains attached to the molecule. Additionally, the polymeric backbone and the method used for grafting QAC to materials can play an important role on the antimicrobial mechanism of action (leaching or contact-killing) and durability of the antimicrobial feature. Chapter 2 enlightens the choice of QAC as an antimicrobial compound for the studies in chapters 3 and 4 in view of the antimicrobial action against a broad spectrum of microorganisms. A QAC molecule with an alkyl chain of 12 carbons and methacrylate groups was designed aiming for better interactions with methacrylate based dental composite.
In chapter 3, iodide quaternary ammonium methacryloxy silicate (IQAMS)
IQAMS was applied as a coating and exposed on the surface of the composite the biofilm formation decreased significantly. Thus, it was stated that IQAMS should be applied only after polymerization and also reapplication was suggested. Cytotoxicity was not investigated in this study which is of extreme importance when new materials are developed for the clinic and needs to be done before clinical test can be performed. Based on the above findings a different approach was adopted to continue the studies on QAC compounds. To ensure the antimicrobial activity exerted by QAC on the surface without altering material bulk properties, QAC was used as a coating. Also the substrate chosen to be coated was stainless steel considering that metallic orthodontic brackets, wires and bands would represent bigger surface area than cementation line in a clinical setting.
Therefore, in chapter 4, two polyhydroxyurethanes - PHU and PHU* - containing QAC were developed to coat stainless steel surfaces. PHU or PHU* were coupled to QAC with silanes resulting in coatings with different properties. For all coatings we observed an antimicrobial activity and a cytotoxic response on fibroblasts due to leaching of QAC. Washing the PHU-QAC coating in demineralized water led to decreased cytotoxicity, which did not impair the antimicrobial potential of the coating against S. mutans. Ion exchange did show an effect on the chemical and nanostructure scale but was not able to decrease the cytotoxicity. Increasing the time or concentration of the ion exchange process could improve PO43- diffusion. Ensuring more covalent bonds between QAC and
PHU/PHU* under humidity free conditions (e.g. synthesis inside a glove-box) or coupling QAC by means of urethane bonds (more stable than siloxane bonds) could diminish leaching and also help amending cytotoxicity problems. In addition, the cytotoxicity observed in vitro was related to the area/media volume ratio. In vivo, in the oral cavity for instance, there will be a continuous flow of saliva which will constantly dilute the leachate and therewith decreasing the cytotoxicity problem. Investigating the amount of released QAC by mass spectrometry and a whole range of concentrations on cytotoxicity is an important
125
Chapter 6
115
GENERAL DISCUSSION
From human health point of view many deleterious and financially demanding effects are associated with bacterial colonization [1–6]. Furthermore, the pressure of bacterial resistance is increasing, which makes the need for new alternatives to combat bacterial adhesion and biofilm formation necessary. For dental clinicians, caries is a common disease related to biofilm formation and its prevention or treatment remains a challenge. Biofilm formation surrounding cemented orthodontic devices is a highly prevalent clinical situation. The difficulty in cleaning the complex structure of orthodontic devices associated with the lack of compliance to the brushing routine can promptly lead to biofilm formation hindering treatment prognosis and depleting patient’s oral health. Therefore, the development of antimicrobial materials to address these complications is necessary.
Quaternary ammonium compounds (QAC) are a class of antimicrobials claimed to be effective against Streptococcus mutans, a cariogenic species. QAC can be included in dental materials either to modify a surface, making it contact-killing or to be leached out and being active in the environment [7–9]. The review on QAC for biomedical applications (chapter 2) evidences that QAC antimicrobial activity can be influenced by the presence and length of alkyl chains attached to the molecule. Additionally, the polymeric backbone and the method used for grafting QAC to materials can play an important role on the antimicrobial mechanism of action (leaching or contact-killing) and durability of the antimicrobial feature. Chapter 2 enlightens the choice of QAC as an antimicrobial compound for the studies in chapters 3 and 4 in view of the antimicrobial action against a broad spectrum of microorganisms. A QAC molecule with an alkyl chain of 12 carbons and methacrylate groups was designed aiming for better interactions with methacrylate based dental composite.
In chapter 3, iodide quaternary ammonium methacryloxy silicate (IQAMS) molecule was synthesized containing QAC moieties and one methacrylate group. Although IQAMS was uniformly dispersed in the composite and the degree of conversion of the composite was not impaired, some of the bulk properties of the Transbond XT were affected. The cohesive and adhesive strength decreased and IQAMS incorporated in the Transbond XT was not capable of exerting antimicrobial activity against S. mutans probably because it was entrapped inside the polymerized matrix and not available on the composite surface. Only when
116
IQAMS was applied as a coating and exposed on the surface of the composite the biofilm formation decreased significantly. Thus, it was stated that IQAMS should be applied only after polymerization and also reapplication was suggested. Cytotoxicity was not investigated in this study which is of extreme importance when new materials are developed for the clinic and needs to be done before clinical test can be performed. Based on the above findings a different approach was adopted to continue the studies on QAC compounds. To ensure the antimicrobial activity exerted by QAC on the surface without altering material bulk properties, QAC was used as a coating. Also the substrate chosen to be coated was stainless steel considering that metallic orthodontic brackets, wires and bands would represent bigger surface area than cementation line in a clinical setting.
Therefore, in chapter 4, two polyhydroxyurethanes - PHU and PHU* - containing QAC were developed to coat stainless steel surfaces. PHU or PHU* were coupled to QAC with silanes resulting in coatings with different properties. For all coatings we observed an antimicrobial activity and a cytotoxic response on fibroblasts due to leaching of QAC. Washing the PHU-QAC coating in demineralized water led to decreased cytotoxicity, which did not impair the antimicrobial potential of the coating against S. mutans. Ion exchange did show an effect on the chemical and nanostructure scale but was not able to decrease the cytotoxicity. Increasing the time or concentration of the ion exchange process could improve PO43- diffusion. Ensuring more covalent bonds between QAC and
PHU/PHU* under humidity free conditions (e.g. synthesis inside a glove-box) or coupling QAC by means of urethane bonds (more stable than siloxane bonds) could diminish leaching and also help amending cytotoxicity problems. In addition, the cytotoxicity observed in vitro was related to the area/media volume ratio. In vivo, in the oral cavity for instance, there will be a continuous flow of saliva which will constantly dilute the leachate and therewith decreasing the cytotoxicity problem. Investigating the amount of released QAC by mass spectrometry and a whole range of concentrations on cytotoxicity is an important addition which needs to be done in future research. Assessment of antimicrobial activity against different microorganisms is necessary to broaden the potential applications in clinical settings.
Surface sealants are coatings applied over composite resin restorations to occlude micropores and smoothen the surface. These surface sealants will lead to diminished roughness [10] thus promoting less bacterial adhesion and staining [11]
and thus prolonging life-time for the composite resin restoration [12]. In chapter 5 we combined the antimicrobial properties of QAC with a commercial dental surface sealant, Biscover (Bisco). Even after light curing the methacrylate based material, QAC exhibited antimicrobial properties by diminishing S. mutans biofilm formation, differing from the findings on chapter 3, where the IQAMS compound was inserted in the composite resin for brackets cementation which did not exert antimicrobial properties in any concentration tested. Abrasion resistance against tooth brushing is also a requirement when it comes to coatings for dental application. In order to evaluate a protective effect of the sealant with and without the QAC, surfaces were submitted to a mechanical brushing cycling equivalent to 2.5 months of patients’ brushing routine [10,11,13–15]. The protective effect of the surface sealant was enhanced when QAC was added. Roughness measurements by means of atomic force microscopy or profilometer need to be done to obtain more quantitative parameters instead of only the visual observation with SEM. From SEM is looks like the surface is kept smooth which will probably decrease bacterial adhesion.
In the oral environment, surfaces are embedded in saliva which is a fluid heterogeneously composed by proteins, enzymes, immunoglobulins, mucins and electrolytes [16]. The saliva will promptly interact with all materials that are inserted in the oral cavity and therefore it is of great importance to use salivary coatings on newly developed biomaterials [17,18]. Previous investigations with antimicrobial QAC have shown though that the salivary pellicle do not impede QAC of killing bacteria [19–22]. The studies presented in this thesis were conducted without a salivary conditioning film, since new materials were being developed. To assess antimicrobial properties it is easier to study it without a protein film, but it would be appropriate to investigate the antimicrobial performance of the developed materials in the presence of a saliva pellicle. Also all materials developed still need some optimization such as adequate and more controllable drying time for coatings, leaching levels adjustment and then in situ or
REFERENCES
[1] WHO. Antimicrobial resistance: Global health report on surveillance. Bull World Health Organ 2014:1–256.
[2] Edwardson S, Cairns C. Nosocomial infections in the ICU. Anaesth Intensive Care Med 2019;20:14–8.
[3] Riga EK, Vöhringer M, Widyaya VT, Lienkamp K. Polymer-based surfaces designed to reduce biofilm formation: From antimicrobial polymers to strategies for long-term applications. Macromol Rapid Commun 2017;38:1700216.
[4] Bowen WH, Burne RA, Wu H, Koo H. Oral Biofilms: Pathogens, matrix, and polymicrobial interactions in microenvironments. Trends Microbiol 2018;26:229–42.
[5] Rode S de M, Gimenez X, Montoya VC, Gómez M, Blanc SL de, Medina M, et al. Daily biofilm control and oral health: Consensus on the epidemiological challenge - Latin American Advisory Panel. Braz Oral Res 2012;26:133–43.
[6] Kriebel K, Hieke C, Müller-Hilke B, Nakata M, Kreikemeyer B. Oral biofilms from symbiotic to pathogenic interactions and associated disease - connection of periodontitis and rheumatic arthritis by peptidylarginine deiminase. Front Microbiol 2018;9:1–14.
[7] Ge Y, Wang S, Zhou X, Wang H, Xu H, Cheng L. The use of quaternary ammonium to combat dental caries. Materials (Basel) 2015;8:3532–49. [8] Imazato S, Chen J hua, Ma S, Izutani N, Li F. Antibacterial resin monomers
based on quaternary ammonium and their benefits in restorative dentistry. Jpn Dent Sci Rev 2012;48:115–25. doi:10.1016/j.jdsr.2012.02.003.
[9] Makvandi P, Jamaledin R, Jabbari M, Nikfarjam N, Borzacchiello A. Antibacterial quaternary ammonium compounds in dental materials: A systematic review. Dent Mater 2018;34:851–67.
127
Chapter 6
117
and thus prolonging life-time for the composite resin restoration [12]. In chapter 5 we combined the antimicrobial properties of QAC with a commercial dental surface sealant, Biscover (Bisco). Even after light curing the methacrylate based material, QAC exhibited antimicrobial properties by diminishing S. mutans biofilm formation, differing from the findings on chapter 3, where the IQAMS compound was inserted in the composite resin for brackets cementation which did not exert antimicrobial properties in any concentration tested. Abrasion resistance against tooth brushing is also a requirement when it comes to coatings for dental application. In order to evaluate a protective effect of the sealant with and without the QAC, surfaces were submitted to a mechanical brushing cycling equivalent to 2.5 months of patients’ brushing routine [10,11,13–15]. The protective effect of the surface sealant was enhanced when QAC was added. Roughness measurements by means of atomic force microscopy or profilometer need to be done to obtain more quantitative parameters instead of only the visual observation with SEM. From SEM is looks like the surface is kept smooth which will probably decrease bacterial adhesion.
In the oral environment, surfaces are embedded in saliva which is a fluid heterogeneously composed by proteins, enzymes, immunoglobulins, mucins and electrolytes [16]. The saliva will promptly interact with all materials that are inserted in the oral cavity and therefore it is of great importance to use salivary coatings on newly developed biomaterials [17,18]. Previous investigations with antimicrobial QAC have shown though that the salivary pellicle do not impede QAC of killing bacteria [19–22]. The studies presented in this thesis were conducted without a salivary conditioning film, since new materials were being developed. To assess antimicrobial properties it is easier to study it without a protein film, but it would be appropriate to investigate the antimicrobial performance of the developed materials in the presence of a saliva pellicle. Also all materials developed still need some optimization such as adequate and more controllable drying time for coatings, leaching levels adjustment and then in situ or
in vivo models of testing. Overall, addition of QAC in dental materials is feasible
and brings beneficial effects for oral health maintenance. Many of the dentistry areas could profit from antimicrobial properties of QAC. Still final application required features should be thought through for new material development. Aspects such as cytotoxicity, weakening of the bulk material and bacterial resistance to QAC still need thorough investigations giving room to future researches in the broad field of antimicrobial materials that QAC enabled.
118
REFERENCES
[1] WHO. Antimicrobial resistance: Global health report on surveillance. Bull World Health Organ 2014:1–256.
[2] Edwardson S, Cairns C. Nosocomial infections in the ICU. Anaesth Intensive Care Med 2019;20:14–8.
[3] Riga EK, Vöhringer M, Widyaya VT, Lienkamp K. Polymer-based surfaces designed to reduce biofilm formation: From antimicrobial polymers to strategies for long-term applications. Macromol Rapid Commun 2017;38:1700216.
[4] Bowen WH, Burne RA, Wu H, Koo H. Oral Biofilms: Pathogens, matrix, and polymicrobial interactions in microenvironments. Trends Microbiol 2018;26:229–42.
[5] Rode S de M, Gimenez X, Montoya VC, Gómez M, Blanc SL de, Medina M, et al. Daily biofilm control and oral health: Consensus on the epidemiological challenge - Latin American Advisory Panel. Braz Oral Res 2012;26:133–43.
[6] Kriebel K, Hieke C, Müller-Hilke B, Nakata M, Kreikemeyer B. Oral biofilms from symbiotic to pathogenic interactions and associated disease - connection of periodontitis and rheumatic arthritis by peptidylarginine deiminase. Front Microbiol 2018;9:1–14.
[7] Ge Y, Wang S, Zhou X, Wang H, Xu H, Cheng L. The use of quaternary ammonium to combat dental caries. Materials (Basel) 2015;8:3532–49. [8] Imazato S, Chen J hua, Ma S, Izutani N, Li F. Antibacterial resin monomers
based on quaternary ammonium and their benefits in restorative dentistry. Jpn Dent Sci Rev 2012;48:115–25. doi:10.1016/j.jdsr.2012.02.003.
[9] Makvandi P, Jamaledin R, Jabbari M, Nikfarjam N, Borzacchiello A. Antibacterial quaternary ammonium compounds in dental materials: A systematic review. Dent Mater 2018;34:851–67.
[10] Khalaj K, Soudi A, Tayefi-Nasrabadi M, Keshvad MA. The evaluation of surface sealants’ effect on the color stability of Nano-hybrid composite after polishing with One-Step system (in-vitro). J Clin Exp Dent 2018;10:e927–32.
[11] Dede DÖ, Şahin O, Koroglu A, Yilmaz B. Effect of sealant agents on the color stability and surface roughness of nanohybrid composite resins. J Prosthet Dent 2016;116:119–28.
[12] dos Santos PH, Pavan S, Suzuki TYU, Briso ALF, Assunção WG, Sinhoreti MAC, et al. Effect of fluid resins on the surface roughness and topography of resin composite restorations analyzed by atomic force microscope. J Mech Behav Biomed Mater 2011;4:433–9.
[13] Mara da Silva T, Barbosa Dantas DC, Franco TT, Franco LT, Rocha Lima Huhtala MF. Surface degradation of composite resins under staining and brushing challenges. J Dent Sci 2018;14:87–92.
[14] Korbmacher-Steiner HM, Schilling AF, Huck LG, Kahl-Nieke B, Amling M. Laboratory evaluation of toothbrush/toothpaste abrasion resistance after smooth enamel surface sealing. Clin Oral Investig 2013;17:765–74.
[15] Tsujimoto A, Barkmeier WW, Fischer NG, Nojiri K, Nagura Y, Takamizawa T, et al. Wear of resin composites: Current insights into underlying mechanisms, evaluation methods and influential factors. Jpn Dent Sci Rev 2018;54:76–87.
[16] Humphrey SP, Williamson RT. A review of saliva: Normal composition, flow, and function. J Prosthet Dent 2001;85:162–9.
[17] Fournier A, Payant L, Bouclin R. Adherence of Streptococcus mutans to orthodontic brackets. Am J Orthod Dentofac Orthop 1998;114:414–7. [18] Papaioannou W, Gizani S, Nassika M, Kontou E, Nakou M. Adhesion of
Streptococcus mutans to different types of brackets. Angle Orthod
2007;77:1090–5.
[19] Yue J, Zhao P, Gerasimov JY, Van de Lagemaat M, Grotenhuis A, Rustema-Abbing M, et al. 3D-Printable Antimicrobial Composite Resins. Adv Funct Mater 2015;25:6756–67.
[20] Gong S, Epasinghe J, Rueggeberg FA, Niu L, Mettenberg D, Yiu CKYY, et al. An ormosil-containing orthodontic acrylic resin with concomitant improvements in antimicrobial and fracture toughness properties. PLoS One 2012;7:e42355.
[21] Gong SQ, Epasinghe DJ, Zhou B, Niu LN, Kimmerling KA, Rueggeberg FA, et al. Effect of water-aging on the antimicrobial activities of an
ormosil-129
Chapter 6
119
[12] dos Santos PH, Pavan S, Suzuki TYU, Briso ALF, Assunção WG, Sinhoreti MAC, et al. Effect of fluid resins on the surface roughness and topography of resin composite restorations analyzed by atomic force microscope. J Mech Behav Biomed Mater 2011;4:433–9.
[13] Mara da Silva T, Barbosa Dantas DC, Franco TT, Franco LT, Rocha Lima Huhtala MF. Surface degradation of composite resins under staining and brushing challenges. J Dent Sci 2018;14:87–92.
[14] Korbmacher-Steiner HM, Schilling AF, Huck LG, Kahl-Nieke B, Amling M. Laboratory evaluation of toothbrush/toothpaste abrasion resistance after smooth enamel surface sealing. Clin Oral Investig 2013;17:765–74.
[15] Tsujimoto A, Barkmeier WW, Fischer NG, Nojiri K, Nagura Y, Takamizawa T, et al. Wear of resin composites: Current insights into underlying mechanisms, evaluation methods and influential factors. Jpn Dent Sci Rev 2018;54:76–87.
[16] Humphrey SP, Williamson RT. A review of saliva: Normal composition, flow, and function. J Prosthet Dent 2001;85:162–9.
[17] Fournier A, Payant L, Bouclin R. Adherence of Streptococcus mutans to orthodontic brackets. Am J Orthod Dentofac Orthop 1998;114:414–7. [18] Papaioannou W, Gizani S, Nassika M, Kontou E, Nakou M. Adhesion of
Streptococcus mutans to different types of brackets. Angle Orthod
2007;77:1090–5.
[19] Yue J, Zhao P, Gerasimov JY, Van de Lagemaat M, Grotenhuis A, Rustema-Abbing M, et al. 3D-Printable Antimicrobial Composite Resins. Adv Funct Mater 2015;25:6756–67.
[20] Gong S, Epasinghe J, Rueggeberg FA, Niu L, Mettenberg D, Yiu CKYY, et al. An ormosil-containing orthodontic acrylic resin with concomitant improvements in antimicrobial and fracture toughness properties. PLoS One 2012;7:e42355.
[21] Gong SQ, Epasinghe DJ, Zhou B, Niu LN, Kimmerling KA, Rueggeberg FA, et al. Effect of water-aging on the antimicrobial activities of an ormosil-containing orthodontic acrylic resin. Acta Biomater 2013;9:6964–73.
[22] Van de Lagemaat M, Grotenhuis A, Van de Belt-Gritter B, Roest S, Loontjens TJA, Busscher HJ, et al. Comparison of methods to evaluate bacterial contact-killing materials. Acta Biomater 2017;59:139–47.
