Corresponding Author: Miriam Machado Cunico - Universidade Federal do Paraná Departamento de Farmácia - Laboratório de Fitoquímica - Av. Prefeito Lothário Meissner, 632 - CEP.80210-170 - Curitiba – PR - Brazil - fone: +55- 41- 3360-4066 - e-mail: miriammcunico@gmail.com Revista de Ciências Farmacêuticas Básica e Aplicada Journal of Basic and Applied Pharmaceutical Sciences Rev Ciênc Farm Básica Apl., 2012;33(4):515-519 ISSN 1808-4532 Bioautography to assess antibacterial activity of Ottonia martiana Miq. (Piperaceae) on the human oral microbiota Miriam Machado Cunico 1,4* ; Celso Garcia Auer 2 ; Cristina Peitz de Lima 1 ; Lilian Cristina Côcco 3 ; Carlos Itsuo Yamamoto 3 ; Marilis Dallarmi Miguel 1 ; Obdulio Gomes Miguel 1 ; Carlos Roberto Sanquetta 4 1 Universidade Federal do Paraná, Departamento de Farmácia, Laboratório de Fitoquímica, Curitiba, PR, Brazil. 2 Embrapa Florestas, Laboratório de Fitopatologia, Colombo, PR, Brazil. 3 Universidade Federal do Paraná, Departamento de Engenharia Química, LACAUTets, Curitiba, PR, Brazil. 4 Universidade Federal do Paraná, Departamento de Engenharia Florestal, LIF, Curitiba, PR, Brazil. ABSTRACT Ottonia martiana Miq. (Piperaceae), a plant known popularly in southern Brazil as “anestésia” and used in the treatment of odontalgia for its anesthetic action on the oral mucosa, was investigated for antibacterial activity by paper disc agar diffusion and bioautographic methods, against microorganisms present in the human oral cavity [Streptococcus mutans (ATCC 25175), Streptococcus mitis (ATCC 49456), Streptococcus pyogenes (ATCC 19615), Streptococcus salivarius (ATCC 25975), Escherichia coli (ATCC 11229 and 25922), Pseudomonas aeruginosa (ATCC 27853) and Enterobacter aerogenes (ATCC 27853).The crude extract of O. martiana (32.9 mg mL -1 ) had antibacterial potential against all Gram-positive bacteria tested. Analysis of the bioautograms led to the detection of bioactive substances, among which it was possible to identify piperovatine (Rf 0.35), piperlonguminine (Rf 0.52) and isopiperlonguminine (Rf 0.52). The piperovatine and isopiperlonguminine were isolated from the roots of O. martiana, guided by a bioautographic antibacterial bioassay. Keywords: Amides. Antibacterial activity. Bioautography. Toothache. Ottonia martiana. INTRODUCTION Although antimicrobial activity has already been demonstrated in many plant extracts, few studies on bacteria of clinical relevance in dentistry have been conducted in Brazil. Despite scientiic advances, we are still constantly affected by caries and periodontal disease. Microorganisms of the human oral biota, such as Streptococcus mutans, are associated with the formation of dental plaque and are a determining factor in the development and progression of caries and periodontal disease (Pereira et al., 2009). Gram- negative bacteria, such as Escherichia coli, Enterobacter spp. and Pseudomonas aeruginosa, are opportunistic microorganisms colonizing the oral cavity, and have a role in the formation of oral bioilm and development of nosocomial pneumonia (Amaral et al., 2009). Studies on Salmonella bacteria revealed a family of proteins related to the formation of bioilms, promoting the colonization of new surfaces, and prevention, by oral hygiene and control of the oral bioilm, is of extreme importance (Amaral et al., 2009). Some of these microorganisms have acquired antibiotic resistance and this has led to a search for potential alternative antimicrobial substances in plants (Sarac & Ugur, 2007; Firas & Al-Bayati., 2008; Pereira et al., 2009). Amongst some plant species popularly used to treat diseases of the oral cavity is Ottonia martiana Miq. (Piperaceae), a local shrub of the Brazilian Atlantic Forest (Cunico et al., 2006). In Brazil, O. martiana is known as “anesthesia” among the coastal community of Paraná state. The roots and aerial parts of O. martiana are largely employed to relieve toothache, in ethanolic mouthwashes, due to its anesthetic action on the oral mucosa (Cunico et al., 2005; Cunico et al., 2006). In South and Southeast Brazil, it is popularly known as “jaborandi”, “jaguarandi” and “taburutá” (Cunico et al., 2005). Phytochemical investigations of the roots and aerial parts of O. martiana resulted in the isolation of the isobutylamides piperlonguminine, isopiperlonguminine and piperovatine, bioactive substances of medical interest (Cunico et al., 2006). Seventy-seven compounds have been identiied in three essential oils extracted from the leaves, fruits and roots of this species (Cunico et al., 2007). These chemical substances were previously isolated from a number of closely-related Piperaceae species (Cunico et al., 2005).