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Ткаченко Г.М., Буюн Л.И., Осадовский З., Гончаренко В.И., Прокопив А.И.
ANTIMICROBIAL ACTIVITY OF ETHANOLIC EXTRACT
OBTAINED FROM FICUS VASTA FORSSK. (MORACEAE)
Институт биологии и охраны среды, Поморская Академия в Слупске,
Национальный ботанический сад им. Н.Н. Гришко НАН Украины, Ботанический сад Львовского национального университета им. Ивана Франко, Львовский национальный университет им. Ивана Франко
Ключевые слова: Moraceae, Ficus vasta Forssk., антимикробные средства, диско-диффузионный метод
Аннотация. Цель настоящего исследования состояла в изучении in vitro ингибирующего влияния этанольного экстракта, полученного из листьев Ficus vasta Forssk. (Moraceae) на рост грамотрицательных [Pseudomonas aeruginosa (ATCC 27853), локально изолированные штаммы Pseudomonas aeruginosa, продуцирующие металло-ß-лактамазы (MßL), Escherichia coli (ATCC 25922)] и грамположительных микроорганизмов [Staphylococcus aureus (ATCC 25923), локально изолированный штамм метициллин-резистентного Staphylococcus aureus и Streptococcus pneumoniae (ATCC 49619)], а также грибка Candida albicans. Профиль антимикробной активности растения в отношении тестированных штаммов свидетельствует о том, что наиболее восприимчивыми к этанольному экстракту F. vasta оказались S. aureus и E. coli. Эти результаты дают многообещающую исходную информацию для потенциального использования этого неочищенного экстракта, а также некоторых из выделенных соединений для лечения бактериальных и грибковых инфекций.
Keywords: Moraceae, Ficus vasta Forssk., antimicrobial activity, disc diffusion technique
Abstract. In the present study, the inhibitory effects of ethanolic extract obtained from leaves of Ficus vasta Forssk. (Moraceae) on the growth of Gram-negative bacteria [Pseudomonas aeruginosa (ATCC 27853), locally isolated Pseudomonas aeruginosa strains producing metallo-beta-lactamases (MBL) enzyme, and Escherichia coli (ATCC 25922)] and Gram-positive bacteria [Staphylococcus aureus (ATCC 25923), methicillin-resistant Staphylococcus aureus locally isolated and Streptococcus pneumoniae (ATCC 49619)] as well as Candida albicans were investigated in vitro. The antimicrobial activity profile of plant against the tested strains indicated that Staphylococcus aureus and Escherichia coli were the most susceptible bacteria of all the bacterial strains tested. These results provide promising baseline information for the potential use of this crude extract as well as some of the isolated compounds in the treatment of bacterial and fungal infections.
Introduction. The angiosperm family Moraceae represents morphologically quite diverse plant group, including terrestrial and hemi-epiphytic trees, shrubs, lianas, subshrubs and herbs. Its distribution range lies mostly within the tropics and subtropics with several taxa extending to the temperate zone [2, 5, 8]. Among 37 genera of Moraceae comprising 1050-1100 species in total, Ficus L. is the largest one with ca 750 species of tropical and subtropical distribution worldwide. Its characteristic features include the presence of waxy glands on vegetative plant parts, heterostyly, and prolonged protogyny, i.e., the anthesis of staminate flowers in already mature fruits. These features are functionally linked to the unique pollination mode in Ficus, which involves mutualistic relationships with agaonid wasps (order Hymenoptera) and has attracted much research interest over the last two centuries [3, 6].
Ficus trees have widely been used by humans over their history in a variety of industries and fields of activity. Virtually all parts of their body are utilized by local people in various medicinal practices to cure wounds, sores, stomach and eye problems, headaches and toothaches, and even tumours and cancer, etc. A number of species are known helpful in healing disorders of digestive and respiratory systems, parasitic infections, and also as painkillers, tonics, and ecbolics [9].
These species have great medicinal values as it has been reported to have enormous phytochemical constituents including tannins, flavonols and flavonoids, terpenoids, coumarins, glycosides, esters, carbohydrates, serine
protease, etc. Thus, these plants have great medicinal potential for the therapy of infection [10].
Ficus vasta Forssk. is a monoecious deciduous tree reaching up to 25 m in height, terrestrial, hemi-epiphytic or epilithic, with yellowish to whitish or brownish hirsute leafy twigs. The species is native to northeastern Africa and usually grows in relatively dry habitats or in riverine vegetation. Its leaves reach 8-35 cm in length and 4-25 cm in width; they are cordiform to suborbicular or ovate to reniform, coriaceous and densely puberulous to almost glabrous, with shortly acuminate to rounded apex, cordate base, and long petiole. The pedunculate puberulous syconia of subglobose to ellipsoid shape are born in pairs or solitary in the leaf axils or just below the leaves and reach 2-2.5 cm in diameter, at maturity green with paler spots [4].
It is important to investigate scientifically this plant which have been used in traditional medicines as potential sources of novel antimicrobial compounds. So, the present study was conducted to investigate in vitro antimicrobial activity of ethanolic extract obtained from leaves of F. vasta against Gram-positive and Gram-negative bacteria as well as fungus Candida albicans to determine the possible use of this plant in preventing infections.
Рисунок 1 - Leaves of Ficus vasta Forssk
Material and methods. The leaves of Ficus vasta Forssk. were sampled in M.M. Gryshko National Botanical Garden (Kyiv, Ukraine) and Botanical Garden of Ivan Franko Lviv National University (Lviv, Ukraine). The whole collections of tropical and subtropical plants at both Gardens (including Ficus spp. plants) have the status of a National Heritage
Collections of Ukraine. The collected leaves were brought into the laboratory for antimicrobial studies. Freshly leaves were weighted, washed, crushed, and homogenized in 96% ethanol (in ratio 1:10, w/w) at room temperature.
The testing of antibacterial activity of the plant extract was carried out in vitro by Kirby-Bauer disc diffusion technique [1]. Gram-negative bacteria [Pseudomonas aeruginosa (ATCC 27853), locally isolated Pseudomonas aeruginosa strains producing metallo-beta-lactamases (MBL) enzyme, and Escherichia coli (ATCC 25922)] and Gram-positive bacteria [Staphylococcus aureus (ATCC 25923), methicillin-resistant Staphylococcus aureus locally isolated and Streptococcus pneumoniae (ATCC 49619)], as well as fungus Candida albicans were used as test organisms. Zone diameters were determined and averaged. Results of the antimicrobial activities assessment are presented as mean ± standard error of the mean. All statistical calculation was performed on separate data from each bacterial and fungal strains.
Results and discussion. The results of antimicrobial activity of ethanolic extract obtained from leaves of F. vasta are presented in Fig. 2.
Рисунок 2 - Antimicrobial activity of the ethanolic extract obtained from leaves of F. vasta against bacterial and fungal strains measured as inhibition zone diameter (M±m, n=8)
In the present study, the inhibitory effects of ethanolic extract obtained from leaves of F. vasta on the growth of Gram-negative and Gram-positive bacteria, as well as fungus Candida albicans were investigated in vitro. The antimicrobial activity profile of plant extract against the tested strains indicated that Staphylococcus aureus and Escherichia coli were the most susceptible bacterium of all the bacterial test strains. Some species of plants are still of special interest for further investigations in this regard as in the
case of F. vasta, which showed exceptionally high activity against E. coli and S. aureus (Fig. 2). Remarkably, the less susceptible microorganisms to the antimicrobial activity of F. vasta were P. aeruginosa and locally isolated Pseudomonas aeruginosa strains producing metallo-beta-lactamases (MBL) enzyme (Fig. 3), as well as methicillin-resistant S. aureus locally isolated, Streptococcus pneumoniae, and fungus C. albicans. Among other bacterial strains tested, there were no significant differences in susceptibility to plant extract screened between the Gram-positive and Gram-negative bacteria.
Рисунок 3 - Antimicrobial activity of the ethanolic extract obtained from leaves of F. vasta against Pseudomonas aeruginosa (ATCC 27853) (A) and locally isolated Pseudomonas aeruginosa strains producing metallo-beta-lactamases (MBL) enzyme (B) measured as inhibition zone diameter
Our results are comparable with many other investigations with various Ficus species [10, 11]. In addition, these data fully corroborated our previous studies which reported that various Ficus species have noticeable antibacterial activity against bacterial strains [12-17]. It is generally assumed, that antibacterial activity of various Ficus species can be explain due to the presence of secondary metabolites [10]. The known antimicrobial mechanisms associated to groups of chemicals (tannins, flavonols and flavonoids, terpenoids, coumarins, glycosides, esters, carbohydrates, serine protease), the isolated compounds from Ficus plants belong may explain the antimicrobial potency of the crude extract obtained from F. vasta. Membrane disruption could be suggested as one of the likely mechanisms of action of triterpens [7]. The activity of flavonoids might be due to their ability to complex with bacterial cell wall [7] and therefore, inhibiting the microbial growth. The fact that F. vasta naturally contains a wide range of active principles could also explain the important antimicrobial spectrum of the different parts used in this study.
A
B
Conclusions. The results of the present study provide an important basis for the use of ethanolic extract from the leaves of Ficus vasta for the treatment of infections associated to the microorganisms used in this study. The crude extracts as well as the isolated compounds in future could be useful for the development of new antimicrobial drug. However, further pharmacological and toxicity studies currently going on in our laboratory will be necessary to confirm this hypothesis.
Список литературы
1. Bauer A.W., Kirby W.M., Sherris J.C., Turck M. 1966. Antibiotic susceptibility testing by a standardized single disk method. Am. J. Clin. Pathol., 45(4): 493-496.
2. Berg C.C. 2001. Moreae, Artocarpeae, and Dorstenia (Moraceae), with introductions to the family and Ficus and with additions and corrections to Flora Neotropica Monograph 7. Flora Neotropica Monograph 83. The New York Botanical Garden, New York, pp. 1-346.
3. Berg C.C., Corner E.J.H. 2005. Moraceae (Ficus). In: Noteboom H.P. (ed.) Flora Malesiana, Ser. 1, Vol. 17, Part 2. National Herbarium Nederland, Leiden, pp. 1-730.
4. Berg C.C., Wiebes J.T. 1992. African fig trees and fig wasps. Koninklijke Nederlandse Akademie van Wetenschappen, Verhandelingen Afdeling Natuurkunde, 2de reeks, deel 89. North-Holland, Amsterdam, 298 p.
5. Tkachenko G., Buyun L., Osadovskyy Z., Truhan M., Sosnowski E., Prokopiv A., Goncharenko V. 2016. In vitro screening of antimicrobial activity of ethanolic extract obtained from Ficus lyrata Warb. (Moraceae) leaves. Agroecological Journal, 2: 155-160.
6. Tkachenko H., Buyun L., Osadowski Z., Honcharenko V., Prokopiv A. 2016. Studies on antibacterial activity of Ficus natalensis Hochst. subsp. leprieurii (Miq.) C.C. Berg (Moraceae) leaf extract. Лжарське рослинництво: ввд досвщу минулого до новггшх технологш: матерiали п'ято! Мiжнародноi науково-практично! iнтернет-конференцii. -Полтава, 27-28 грудня 2016 р. - Полтава: РВВ ПДАА, С. 256-259.
