Psidium cattleianum (Myrtaceae) as a Natural Antimicrobial Source Against Oral Bacteria

Investigation of natural compounds from popular medicinal plants have advanced in the last decades in the attempt to characterize scientifically the possible beneficial effects and side effects, as well as the determination of a safe dose of use [1]. Among the families of medicinal plants, the Myrtaceae family stands out, whose genus Psidium adds fruit species such as Psidium guajava, the popular guava. However, the species Psidium cattleianum and its variants, known as strawberry guava, has been studied in the last decades because of its medicinal property such as analgesic effects [2], antioxidant [3,4], anti-proliferative action [4], anticancer [4] and antimicrobial agent [5]. In popular medicine, P. cattleianum is known for its action against diabetes, tummy aches, urinary tract diseases and diarrhea, where it is recommended to use shoots, leaves and bark of the trunk [6].


Introduction
Investigation of natural compounds from popular medicinal plants have advanced in the last decades in the attempt to characterize scientifically the possible beneficial effects and side effects, as well as the determination of a safe dose of use [1]. Among the families of medicinal plants, the Myrtaceae family stands out, whose genus Psidium adds fruit species such as Psidium guajava, the popular guava. However, the species Psidium cattleianum and its variants, known as strawberry guava, has been studied in the last decades because of its medicinal property such as analgesic effects [2], antioxidant [3,4], anti-proliferative action [4], anticancer [4] and antimicrobial agent [5]. In popular medicine, P. cattleianum is known for its action against diabetes, tummy aches, urinary tract diseases and diarrhea, where it is recommended to use shoots, leaves and bark of the trunk [6].

P. cattleianum Medicinal Compounds
The medicinal effects are derived from the bioactive compounds in plants that originate from the secondary metabolism, which is not involved in the essential metabolic pathways for plant's life, but play the role of defense in plants against bacteria, fungi, viruses, environmental stress, Herbivorous attack and attraction of pollinating agents [7]. The phenolic compounds are found in plants extracts and showed antimicrobial activity by causing degradation in bacterial cell wall, damage in cytoplasm membrane and consequent cellular material extravasation, cytoplasm coagulation and disruption in the ion/electrons membrane transport system. Any disarrangement on the cellular surface entails in ruptures in cell surface, which leads the output of vital cellular components and entry of harmful in bacterial cell [8].
Terpenoid compounds are found in essential oils. Monoterpenes and sesquiterpenes are the most abundant terpenoids compounds and play an antimicrobial action involving membrane fluidification, destabilization of membrane binding proteins involved in signaling and anchorage, rest of cell cycle and induction of apoptosis in Candida albicans cells [9]. Turina et al. [10] reported that owing the high hydrophobicity of monoterpenes, their toxic effects on the bacterial cell membrane result in its expansion, fluidity, and permeability, in addition to causing inhibition of respiration and change in the process of ions transport, but further studies should be conducted to characterize in detail these mechanisms. In the work of [11], the Brazilian P. cattleianum plant showed by GC/MS 33 components corresponding to 96.9% of the oil, and the main terpenoids of the essential oil were α-thujene (25.2%), followed by 1,8-cineole (16.4%) and β-caryophyllene (10.2%). This results were similar to the African P. cattleianum plant, that were identified by CG/ MS a total of 53 chemical components, accounting for 61% of the essential oil and the major component was caryophyllene

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oxide (12.43%), bicyclo(4.4.0)dec-l-ene (6.61%), 2,3-butanediol diacetate (4.84%) and patchoulene (4.73%). The presence of many terpenic and ester compounds is thought to contribute to the unique flavor of the P. cattleianum leaves [12]. In the aqueous extract of P. cattleianum the phenolic compounds present were: 3 flavonoids (kaempferol, quercetin, and cyanidin) and 1 is a tannin (ellagic acid) [5]. Although, the oil and extract compounds of P. cattleianum leaves may vary in response to different physical, atmospheric, growth region and time of year collections.

Antimicrobial Effects of Essencial Oil and Extracts from P. Cattleianum Leaves
Scur et al. [13], testing the antimicrobial activity of Psidium cattleianum essential oil, (prepared with metanol) showed a high minimum inhibitory concentration (MIC) of 200mg/ml for grampositive microorganisms (E. faecalis, S. epidermidis, S. aureus, B. subtillis and C. albicans) and for gram-negative microorganisms (P. aeruginosa, S. enteritidis, P. mirabilis, K. pneumonia and E.coli). Although, comparative studies involving the essential oil of leaves of P. cattleianum are still in scarce and its antimicrobial effects are undescribed and uncharacterized. The extract of P. cattleianum produced with ethyl acetate showed MIC of 125ug/ml against S.aureus while the methanol extract showed an MIC of 250ug/ mL, but both extracts types had no effect against gram-negative microorganisms [14]. However, Gaetti-Jardim et al. [15] tested the antimicrobial action of aqueous and hydro alcoholic extracts from P. cattleianum leaves against gram-negative microorganisms (F. nucleatum and P. gingivalis) showed MICs of 2mg/mL in both extracts with a reduction of 50% in bacterial growth and 8mg/mL for aqueous and hydro alcoholic extracts with a reduction of 90% in bacterial growth. These different results may be related to the different extractive methods and the solvent used in the process employed by the authors, which may have influenced in isolating bioactive compounds of plant and determination of properties [16].
Brighenti et al. [17] evaluated the leaves aqueous extract of P. cattleianum on the biofilm of Streptococcus, showed that biofilms exposed to the extract effects at 1.6% (v/v) for 2h were shown to decrease in S. mutans proteins from carbohydrate metabolism and a reduction in pH drop after exposure to the extract. The decrease in extracellular polysaccharide (EPS) formation observed after sucrose application may also be related to the inhibition of glicosiltranferases (GTF) activity in dental biofilms, which reduce de biofilm resistance of S. mutans.
Menezes et al. [18] also studied and corroborated that the aqueous extract of P. cattleianum significantly reduced the S. mutans count and decreased the enamel demineralization rate in rats, characterizing an anti-carie activity. Brighenti et al. [5], evaluating the anti-caries effect of P. cattleianum leaf extract using an "in situ" caries model found that P. cattleianum leaf extract might reduce enamel demineralization, acidogenic potential, microorganism viability and extracellular polysaccharide production. Having clearly demonstrated the potential activity of P. cattleianum leaf extract to interfere in "in situ" biofilm pathogenicity, the effect of this plant extract on biofilm formation "in vivo" should be evaluated in the future.

Toxicity of P. cattleianum
The low toxicity of Psidium spp. extract is described in the literature. The LD50 of P. guajava leaf extract is more than 5g kg¬-. Teixeira et al. [19] demonstrated that P. guajava infusion does not alter chromosomes or the cell cycle both in vitro and in vivo. Costa et al. [20] show that P. cattleianum leaf extract does not have any genotoxic or mutagenic effects on some cell types of mice, suggesting that this extract can be used as a safe therapeutic agent. The crude ethanolic extract of the plant was obtained through maceration and fractionated with hexane, dichloromethane, and ethyl acetate for test in microcrustacean Artemia salina for toxicological assessment. Regarding toxicity, hexane and dichloromethane fractions were considered nontoxic, whereas the crude ethanol extract and the ethyl acetate fraction showed low toxicity [4]. There is no data in literature about P. cattleianum essential oil toxicity.

Conclusion
Extracts and essential oil from Psidium cattleianum leaves has antimicrobial properties against oral microorganisms, presenting absence or low toxicity and cytotoxicity in eukaryotic cells. Studies with P. cattleianum essential oils and extracts that characterize medicinal effects and its photo chemical composition should be encouraged for propose future alternatives to combat infections, control microorganisms growth and promote surfaces treatment against biofilms development with the aim that, in the future, can be applied in health area in an economically viable way.