Betelvine ( Piper betle L . ) : A potential source for oral care

Piper betle L. (betelvine) is a valuable crop that is widely used as masticatory and with a long past history of varied traditional uses. Betelvine possesses numerous phytochemicals with important pharmacological attributes. Active molecules such as Fluoride, Eugenol, Hydroxylchavicol, Chlorogenic acid etc. present in betelvine with potent antibacterial, antifungal as well as anti-carcinogenic properties signify tremendous prospective of the plant for the formulation of natural product based drugs for maintaining hygiene and cure of diseases in the oral cavity.


INTRODUCTION BIOCHEMICAL PROFILE OF PIPER BETLE L.
Piper betle L. is indeed very popular and also highly investigated plant. Chemical analysis conducted on betelvine revealed presence of diverse phytochemicals, concentration of which are dependent on the prevailing climate and season [23]. Some of the phytochemicals present in betelvine are important biologically active compounds that accounts for its pharmacological and therapeutic potential. The chief phytochemical constituent of betelvine is essential oil which is responsible for its characteristic pungent aroma [24]. The essential oil of betelvine is light yellow to dark yellow in colour, present in the range between 0.1% to 2.0%, depending on the variety and agro-climatic condition of the area of cultivation. A list of important constituents of betelvine oil as reported in different varieties of the plant is shown in Table 2 [25][26][27][28]. Phytochemicals present in Piper betle L. possesses antimicrobial [29], antiseptic [30], antioxidant [31], anti-inflamatory [32], anti-carcinogenic [33], anti-diabetic [34], anti-genotoxic [35], wound healing [36], anti-hemolytic [37], anti-dermatophytic [38], anti-hypercholesterolemic [39], immunomodulatory [40] and anti-asthmatic [41] properties besides several other pharmacological and therapeutic attributes.

TRADITIONS OF ORAL HYGIENE USING BETELVINE
Leaves are the most valued part of the plant with long past history of varied and traditional human use. Mention of use of betel leaves is found in ancient Sanskrit manuscripts that date back to 600-400 BC and references are also found in ancient tales. Betel leaf extract finds frequent use as an adjuvant or is used singly for the preparation of Ayurvedic medicine and such practices are in use since thousands of years for the cure of common disease to hereditary as well as chronic disorders [42,43].
Chewing betel leaves is a traditional culture prevalent in regions from South Asia eastward to the Pacific. Chewing is customarily practiced by the people of countries like Pakistan, Maldives, India, Nepal, Sri Lanka, Bhutan, Bangladesh, Burma (Myanmar), China, Laos, Thailand, Malaysia, Indonesia, Cambodia, Vietnam, Taiwan, the Philippines, Palau, Yap, Guam, Papua   New Guinea, the Solomon Islands, and Vanuatu. There are estimated 600 betel quid chewers found globally [44]. Chewers regard that this habit of chewing helps to maintain wellbeing of the mouth [45]. In the South East Asia, betel leaves are used for the control of dental caries and for the cure of other periodontal disorder [46]. Indonesian people use betel leaves for cure of oral candidiasis [47]. Chewing betel leaves help prevent halitosis or stop bad breadth [48]. Chewing also makes the gums strong and thereby helps to conserve the teeth [49]. There is enhanced antibiotic, peroxidase and lysozyme activity in the mouth due to increase in secretion of saliva as a result of chewing betel quid. This helps in checking growth of bacteria inside the mouth and thereby protects oral tissues. Betel leaf extract possesses broad spectrum antibacterial, anifungal and antiprotozoal activity [50]. Betel leaves are used for cure of inflammations inside the oral cavity in traditional system of medicine [51].

INFLUENCE OF BETELVINE EXTRACT ON ORAL PATHOGENS
Poor cleaning of the mouth, particularly after having food results in deposition of food substances over the teeth and gums that ultimately converts into a sticky colourless film called as plaque.
Dental plaque if not cleaned properly and not taken proper care it may lead to tooth decay and cavity formation, a condition called as dental caries. Plaque consists of high magnitude of microorganisms, particularly bacteria. These micro-organisms come and harbor in the plaque and thrive on the left over food substances. Dental caries gets developed in several steps. Firstly, pioneer micro-organisms adhere to dental plaque where they proliferate and form colonies. During final stages, there is aggregation of filamentous organisms and spirochetes forming a cohesive biofilm over the teeth and gums and causing damage of the both. Microorganisms harbouring dental plaque keep on releasing acid that causes acidic change inside the mouth. High concentration of acid causes erosion of the tooth enamel and lead to cavity formation. The acid then penetrates into the tooth and brings damage from the inside. Such dental erosions are irreversible that leads to a permanent loss of the tooth structure. Caries is often associated with irritations of the gum tissues. Plaque may also grow below the gums which ultimately causes breakage of the bone that supports the teeth. Dental caries is one of the commonest infectious and a painful disorder of the mouth, management of which is very difficult. For the maintenance of proper oral hygiene, often different antibiotics are generally prescribed in order to get rid of dental caries. However, administration of antibiotics for the cure of dental caries create problems like coloration of the tongue, formation of lesions within the oral cavity and may also bring problems of taste perception [52,53].
Streptococcus mutans is one of the most dominate bacterium that contributes to the development of dental caries [54,55]. It is also an early plaque colonizing bacterium. Among the oral fungal pathogens, Candida albicans is found associated with plaque in early stages. Ali et al. [56] studied the influence of betelvine extract on oral bacterial pathogens such as Escherichia coli, Staphylococcus aureus, Streptococcus mutans and Streptococcus salivarius, as well as on oral fungal pathogens such as Candida albicans. They could obtain 840 mg GAE/g phenol content from dried betel leaves with 95% ethanol that was added to commercially available toothpaste in order to study the effect of such conjugated mixtures on oral pathogens. The conjugated mixtures showed significant zone of inhibition for both bacterial and fungal pathogens used in their experiment.
Teanpaisan et al. [57] gave a comparative account of influence of Thai traditional herbs on seven oral micro-organisms including five Gram positive cariogenic bacteria viz. Enterobacter faecalis, Lactobacillus fermentum, L. salivarius, Streptococcus sobrinus and S. mutans as well as two Gram negative oral pathogenic bacteria, Aggregatibacter actinomycetemcomitans and Fusobacterium nucleatum. Among all the traditional herbs used in their experiment, highest zone of inhibition was produced by the betelvine extract against both gram positive and gram negative bacteria. The minimum inhibitory concentration of P. betle L. was found in the range between 1.04 to 5.21 mg/mL and minimum bactericidal concentration ranged between 2.08 to 8.33 mg/Ml. According to Teanpaisan et al., betelvine extract render impediment to the formation of bacterial biofilm on the tooth surface. Phytochemical screening revealed 4-chromanol to be a major constituent present in betelvine extract.
Caries development depends primarily on the adherence capacity of initial plaque forming bacteria such as Streptococcus mutans [58,59]. Deshpande and Kadam [60] were also able to control Streptococcus mutans by the way of treatment with betel leaf extract. They could control this bacterium by the application of methanol, ethyl acetate, petroleum ether and chloroform extracts of betel leaf. Methanol extract produced maximum zone of inhibition for this bacterium. Deshpande and Kadam reported presence of Eugenol and 4 Chromanol in extracts of betelvine. The average contant of Eugenol in extracts of certain varieties of betelvine leaves was found to be in the range of 0.659 to 1.110 % and the concentration was found to vary depending on the season [61]. Eugenol reportedly has antibacterial and antifungal activities [62][63][64].
Cariostatic property of Eugenol was studied specifically by Jing-Shu et al. [65]. In the experiment conducted by Jing-Shu et al., bacterial adherence capacity showed marked reduction due to treatment of bacterial cells with Eugenol. Eugenol treatment also reduced the acid production by Streptococcus mutans grown in culture. They were also able to control carious lesions in experimental rats due to topical application of Eugenol. Eugenol can counter a wide range of agent that is known to be carcinogenic in nature [66]. Euginol comprises of one free hydroxy group attached to the benzene ring that is considered very important for effective bacterial inhibition [67] Habit of keeping betel quid in the mouth lowers incidence of oral pathogenic microflora that is responsible for dental caries and thereby check bad breathe [68]. Antimicrobial activity of the different ingredients of betel quid i.e. kattha, lime, betel leaf, betel nut, cardamom, clove and fennel seeds was tested on oral pathogens causing dental caries by Shrishailappa et al. [69].
A combined effect of all the ingredients of betel quid was also tested on the same micro-organisms. Different ingredients of betel quid showed varied inhibition capacity against the tested oral pathogens.
Streptococcus sanguinis, S. mitis and Actinomyces are other primary colonizers of dental plaque. Fathilah [70] coated glass plates with extracts of Piper betle L. and Psidium guajava L. and studied the adherence capacity of these oral pathogens on the glass plates. In the experiment he could observe reduced incidence of bacteria on the plant extract treated glass plates. According to Fathilah, hydrophobicity of the bacterial cells is affected by betelvine extract that causes reduced adherence and bacterial colony formation. Micrographs taken with Scanning electron microscope (SEM) showed oral pathogenic cells in non-dividing state due to treatment with betelvine extract. Betel extracts used in the experiment conducted by Fathilah showed presence of 9.25 ppm fluoride content. Fluoride is the ionic form of fluorine with cariostatic and anti-bactericidal property even at very low concentration [71][72][73].
According to the World Health Organization (WHO), adequate uses of fluoride can effectively reduce the problem of dental caries [74]. This fluoride is a common constituent of commercially available toothpaste that is seen to considerably bring a decline of dental caries. Fluoride prevents tooth decay by the way of making the teeth resistant to attacks from plaque bacteria, speeds up the process of remineralization and also disrupts acid production in already affected teeth [75].
Nalina and Rahim [76] also made study of oral pathogenic bacteria found in association with dental plaque. They treated Streptococcus mutans with crude aqueous extract of betelvine. According to Nalina and Rahim, betelvine extract brings damage to the bacterial cells and also lower the acid production by pathogenic bacteria. They also observed plasma membrane damage and coagulation of the nucleoid due to treatment of S. mutans with betelvine extract. Chemical analysis conducted by Nalina and Rahim revealed presence of phenolic compound, Hydroxylchavicol in betelvine. Betel leaf extract comprise of about 26% of this phenolic compound [77]. Hydroxylchavicol present in betelvine is an active antimicrobial agent [78]. Efficacy of this phenolic compound to reduce the adherent capacity of cells of S. mutans suggests for its usefulness for the development of antibacterial agents that would be effective against oral pathogens and for treatment of infections of the mouth [79]. Another active phenolic compound widely found in natural food substances including betelvine is Chlorogenic acid. Chlorogenic acid is also endowed with broad spectrum antimicrobial efficacy [80], including those micro-organisms that are found during early dental plaque formation, such as S. mutans [81].

CONCLUSION
Chemical profiling and bioactive properties of betelvine have been well elucidated in number of scientific studies. It is evident that betel leaves contain a number of phytoconstituents with anti-pathogenic efficacy. Betelvine extract is potentially capable of reducing adherence of the pathogenic micro-organisms on the tooth surface. It can deter primary colonizers of dental plaque. The extract of the plant is also capable of inhibiting growth and proliferation of secondary and tertiary microbial colonizers of dental plaque. By limiting pathogenic microbes on the tooth surface and hindering their subsequent growth, oral problems can be restricted. That betelvine can heal wound implies its potentiality as a prospective source for healing of infections inside the buccal cavity. The plant is endowed with number of cariostatic agents such as Fluoride, Eugenol, Hydroxylchavicol, Chlorogenic acid etc. which can reduce occurrence of pathogenic oral micro-flora and can become an important constituent of antiplaque formulation that is plant based. All these makes Piper betle L. a potential source to be used for maintenance of oral hygiene and dental care.