Int.J.Curr.Microbiol.App.Sci (2016) 5(1): 306-317 ISSN: 2319-7706 Volume 5 Number 1(2016) pp. 306-317 Journal homepage: http://www.ijcmas.com Original Research Article http://dx.doi.org/10.20546/ijcmas.2016.501.029 Antimicrobial and Antioxidant Activity of the Leaves, Bark and Stems of Liquidambar styraciflua L. (Altingiaceae) Graziele Francine Franco Mancarz1*, Ana Carolina Pareja Lobo1, Mariah Brandalise Baril1, Francisco de Assis Franco2 and Tomoe Nakashima1 1Pharmaceutical ScienceDepartment, Universidade Federal do Paraná, Curitiba, PR, Brazil 2Coodetec Desenvolvimento, Produção e Comercialização Agrícola Ltda, Cascavel, PR, Brazil *Corresponding author A B S T R A C T K e y w o r d s The genus Liquidambar L. is the best-known genus of the Altingiaceae Horan family, and species of this genus have long been used for the Liquidambar treatment of various diseases. Liquidambar styraciflua L., which is styraciflua, popularly known as sweet gum or alligator tree, is an aromatic deciduous antioxidant tree with leaves with 5-7 acute lobes and branched stems. In the present activity, study, we investigated the antimicrobial and antioxidant activity of aerial antimicrobial parts of L.styraciflua. Antimicrobial activity was evaluated using the activity, microdilution methodology. The DPPH and phosphomolybdenum methods microdilution method, were used to assess the antioxidant capacity of the samples. The extracts DPPH assay showed moderate or weak antimicrobial activity. The essential oil had the lowest MIC values and exhibited bactericidal action against Escherichia Article Info coli, Enterobacter aerogenes and Staphylococcus aureus. The ethyl acetate fraction and the butanol fraction from the bark and stem showed the best Accepted: antioxidant activity. This study revealed that the essential oil of L. 16 December 2015 styraciflua leaves displays antimicrobial activity and can be used as an Available Online: alternative against pathogenic microorganisms. The extracts and fractions of 10 January 2016 this species have a high antioxidant capacity, revealing new sources of antioxidants for the food and pharmaceutical industries. Introduction Infectious diseases are a prominent cause of agents (Silver and Bostian, 1993).Thisability morbidity and mortality worldwide, despite has driven the search for substances with technical and scientific advances. One of the antimicrobial activity in natural products, as main factors responsible for this status is the many secondary metabolites of plants have ability of microorganisms to acquire been proven to have antimicrobial properties mechanisms of resistance to antimicrobial in vitro (Chauhan et al., 2015). 306 Int.J.Curr.Microbiol.App.Sci (2016) 5(1): 306-317 Various degenerative diseases, such as diseases, such as stomach disorders, wounds cancer and Parkinson's disease, can be and cough; however, the essential oil caused by the uncontrolled production of extracted from the leaves is also known to reactive oxygen species. The use of exhibit medicinal properties (Lingbeck et antioxidants to aid in the prevention and al., 2015). treatment of these diseases has been studied due to their potential to reduce oxidative Some studies of species of the genus stress (Sadi et al., 2015). Many medicinal Liquidambar demonstrated antimicrobial plants contain a large number of phenolic potential (Sa diç et al., 2005; Kim et al., compounds, such as flavonoids and tannins, 2008; Lee Y-S et al., 2009); however, no which exhibit antioxidant activity (Araujo et studies have been conducted with the al., 2008), encouraging research in this area. essential oil and extracts of the species L. styraciflua. Some studies demonstrated A little explored family is Altingiaceae antioxidant, antitumor, anticoagulant and Horan, which consists of approximately hepatoprotective activity for the species fifteen species that are divided into three L.orientalis and L.formosona (Wang et al., genera: Liquidambar L., Altingia Noronha 2010; Konno C et al., 1988; Yang et al., and Semiliquidambar HT. Chang (Ickert- 2011). The essential oil of the leaves and Bond and Wen, 2006).The extracts and stem of L. styraciflua has anti-inflammatory resins of plants from this family are used in activity and antioxidant activity with low traditional Chinese and Ayurvedic medicine cytotoxicity (El-Readi et al., 2013), but for the treatment of stomach pain, there are no reports in the literature bronchitis, inflammation, amenorrhea and concerning the antioxidant capacity of skin diseases, such as eczema and scabies extracts and fractions of the aerial parts of (El-Readi et al., 2013). this species. The genus Liquidambar is the best-known The lack of studies related to this species genus of the family and consists of four encourages this work, which aims to intercontinental species in the temperate evaluate the antimicrobial and antioxidant zone of the northern hemisphere: activity of the extracts, fractions and Liquidambar formosana, Liquidambar essential oil of L.styraciflua. acalycina, Liquidambar orientalis and Liquidambar styraciflua (Ickert-Bond et al., Materials and Methods 2005). One of the species of this genus, L.styraciflua L., which is commonly known Plant Material as sweet gum, red gum, alligator tree and Liquidambar, has a wide natural distribution The botanical material was collected from in the southern and southeastern United specimens of L.styraciflua L., States, extending to Mexico and Central ALTINGIACEAE, which were grown in the America. This organism is a deciduous tree experimental field of the Brazilian that is 20-25m high and has leaves with 5-7 Agricultural Research Corporation, acute lobes that are green in the spring and EMBRAPA Forests (Colombo - PR). The summer but turn purple, red, or yellow in the leaves, bark and stems of L. styraciflua were fall (Lorenzi et al., 2003). More than 10,000 collected during the fall and spring of years ago, Native Americans used the storax 2011/2012. of this species for the treatment of various 307 Int.J.Curr.Microbiol.App.Sci (2016) 5(1): 306-317 The hydroalcoholic extract of the leaves and against the selected microorganisms were the hydroalcoholic extract of the bark and determined in sterile 96-wells or holes (TPP) stems of L. styraciflua were prepared using a according to previously described maceration technique at a moderate methodologies (Ayres et al., 2008; temperature. From these extracts, we Hanamanthagouda et al., 2010; Silva et al., prepared fractions into a separatory funnel 2011), with some modifications. The using the liquid-liquid partition method and recommendations ofClinical and Laboratory solvents with increasing polarity (n-hexane, Standards Institute (2005) were applied. chloroform, ethyl acetate, butanol and ethanol 70%). The hydroalcoholic extracts were added to an initial concentration of 2,000 µg/mL, and The essential oil was extracted from dried the serial dilution procedure was then leaves of L. styraciflua via the performed with the first culture medium to hydrodistillation method, using the obtain decreasing concentrations (2,000 Clevenger apparatus. µg/mL to 15.625 µg/mL). For the essential oil, an initial concentration of 1,000 µg/mL Reagents and Chemicals with the solvent 10%Tween 80 was used, and concentrations of 1,000 µg/mL to 3.9 All chemicals and solvents were purchased µg/mL were obtained by serial dilution with from Sigma Aldrich Co. (St. Louis, MO, medium. USA) and Merck Millipore (Darmstadt, Germany). In the last columns of the plate,we added the positive controls (30 µg/mL of erythromycin Antimicrobial Activity for bacteria; 50 µg/mL of ketoconazole and 50 µg/mL of terbinafine for yeast), the For the antibacterial tests, we selected eight negative controls (solvent) and the sterility standard strains of bacteria from the control for the culture medium. company NEWPROV, including four Gram- positive bacteria and four Gram-negative To verify the ability of the sample to inhibit bacteria. In the antifungal tests, we used five the growth of microorganisms or cause the standard yeast strains from the Mycology death of microorganisms, a methodology Laboratory of Clinical Hospital (Curitiba- described by Veljic et al. (2010) was used, PR), which are often isolated from patients. with some modifications. From wells that The culture media used in this study exhibited an MIC, we collected a10 L included Mueller-Hinton broth and aliquot and transferred the aliquot to Petri Sabouraud broth, which were both obtained dishes containing the appropriate culture from HIMEDIA. After growth on Petri medium for each microorganism. plates containing a suitable culture medium, the bacteria and yeasts were inoculated into After the incubation period, samples that sterile saline (0.9%) and the turbidity was showed no visible microbial growth, adjusted to McFarland standard n°0.5 to indicating the death of 99.5% of the obtain a standard concentration of microorganisms in the inoculum, were microorganisms (1 x 108colony-forming considered to have bactericidal/fungicidal unit/mL). activity. The samples with visible microbial growth were considered to have The minimum inhibitory concentration bacteriostatic/fungistatic activity. (MIC) values of the extracts and essential oil 308 Int.J.Curr.Microbiol.App.Sci (2016) 5(1): 306-317 Antioxidant Activity triplicate. These samples were incubated for 90 minutes in a 95°C water bath, and after The DPPH method that was used to cooling, the absorbance of the samples at determine the antioxidant capacity followed 695nm was measured