Propolis - based chitosan varnish: drug delivery, controlled release and antimicrobial activity against oral pathogen bacteria

Franca et al. BMC Complementary and Alternative Medicine 2014, 14:478 http://www.biomedcentral.com/1472-6882/14/478 RESEARCH ARTICLE Open Access Propolis - based chitosan varnish: drug delivery, controlled release and antimicrobial activity against oral pathogen bacteria Juçara R Franca1, Mariana P De Luca2, Tatiana G Ribeiro1, Rachel O Castilho1, Allyson N Moreira2, Vagner R Santos3,4* and André AG Faraco1 Abstract Background: Dental caries is the most prevalent oral disease in several Asian and Latin American countries. It is an infectious disease and different types of bacteria are involved in the process. Synthetic antimicrobials are used against this disease; however, many of these substances cause unwarranted undesirable effects like vomiting, diarrhea and tooth staining. Propolis, a resinous substance collected by honeybees, has been used to control the oral microbiota. So, the objective of this study was to develop and characterize sustained-release propolis-based chitosan varnish useful on dental cariogenic biofilm prevention, besides the in vitro antimicrobial activity. Methods: Three formulations of propolis - based chitosan varnish (PCV) containing different concentrations (5%, 10% and 15%) were produced by dissolution of propolis with chitosan on hydro-alcoholic vehicle. Bovine teeth were used for testing adhesion of coatings and to observe the controlled release of propolis associated with varnish. It was characterized by infrared spectroscopy, scanning electron microscopy, casting time, diffusion test in vitro antimicrobial activity and controlled release. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were tested for the main microorganisms involved in the cariogenic biofilm through the microdilution test in 96-well plates. Results: The formulations presented a tooth surface adherence and were able to form films very fast on bovine tooth surface. Also, propolis-based chitosan varnishes have shown antimicrobial activity similar to or better than chlorhexidine varnish against all oral pathogen bacteria. All microorganisms were sensitive to propolis varnish and chitosan. MIC and MBC for microorganisms of cariogenic biofilme showed better results than chlorhexidine. Propolis active components were released for more than one week. Conclusion: All developed formulations turn them, 5%, 10% and 15% propolis content varnish, into products suitable for clinical application on dental caries prevention field, deserving clinical studies to confirm its in vivo activity. Keywords: Propolis, Chitosan, Varnish, Drug delivery, Cariogenic biofilm, Prevention, Oral microorganisms Background Dental caries is the most prevalent oral disease in several Asian and Latin American countries, despite the significant decline in many developed countries over recent decades [1-4]. Moreover, it is expected to increase in many developing countries in Africa [5-7]. Although its * Correspondence: 3 Department of Oral Clinical, Oral Pathology and Oral Surgery, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil 4 Faculdade de Odontologia UFMG, Campus Pampulha, Av. Pres. Antonio Carlos, 6627, Pampulha, Belo Horizonte, MG 31270-901, Brasil Full list of author information is available at the end of the article incidence is particularly high during childhood [3,8-10], people are susceptible throughout their lifetime [11,12]. Dental caries is an infectious disease, the development of which is a dynamic process involving alternating demineralization and remineralization, rather than unidirectional demineralization [13]. Demineralization of tooth structure is caused by the organic acids produced in dental plaque biofilm by the metabolic action of the cariogenic microorganisms on fermentable carbohydrates [14,15]. Formation of biofilm is a biological process associated with the attachment, detachment © 2014 Franca et al.; licensee BioMed Central. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Franca et al. BMC Complementary and Alternative Medicine 2014, 14:478 http://www.biomedcentral.com/1472-6882/14/478 Page 2 of 11 and proliferation of oral bacteria on the tooth surfaces. The dental biofilm is formed via adhesion of planktonic bacteria to a protein pellicle coating the tooth surfaces [16,17]. Many types of bacteria participate in the formation of the dental biofilm [18]. Dental caries is caused by the interaction, over the time, of bacteria that produce acids with a substrate that the bacteria can metabolize and many host factors that include teeth and saliva [19]. There are only a few groups of microorganisms that can adhere to the teeth, which includes mutans Strepcococci group, Lactobacillus and some Actinomyces species [20,21]. More than five Streptococcus species and Actinomyces viscosus are regarded as early colonizers of tooth surfaces, while mutans Streptococci such as S. sobrinus, S. salivarius, S. sanguinis and S. mutans are considered middle important colonizers of the dental biofilm [19]. The inhibition of plaque biofilm formation is the key to successful control and prevention of dental caries. Previous antibacterial mouth rinses, which generally contain fluorides, alcohols, detergents and other antimicrobial substances, effectively reduce plaque formation. Synthetic antimicrobials used in tooth pastes and mouth rinses include povidone iodine products, chlorhexidine, cetylpyridinium chloride, triclosan and zinc citrate [22,23]. However, many of these substances cause unwarranted undesirable effects like vomiting, diarrhea and tooth staining. Prevention, based on common risk factors, plays an important role on caries management [24,25] and recent studies have shown the anti-caries activity of different natural products such as propolis [26,27]. Propolis is a resinous substance collected by honeybees from buds and exudates of certain trees and plants and stored inside their hives. It has been used in folk medicine from ancient times to treat various ailments [28]. Propolis has been used for centuries by world population due to its pharmacological properties such as anti-inflammatory, healing, antimicrobial and antioxidant [29]. In dentistry, propolis has been used to control the oral microbiota [30]. The antibacterial activity of propolis is reported due to flavonoids, aromatic acids, and esters present in resins. Galangin, pinocembrin, and pinostrobin are known as the most effective flavonoids agents against bacteria. Ferulic acid and caffeic acid also contribute to the bactericidal action of propolis [31]. (...truncated)