© Borgis - Postępy Fitoterapii 4/2011, s. 227-231
*Paweł Krzyściak1, Mirosław Krośniak2, Maciej Gąstoł3, Dorota Ochońska4, Wirginia Krzyściak5
Przeciwdrobnoustrojowe działanie derenia
Antimicrobial activity of Cornelian cherry (Cornus mas L.)
1Department of Mycology, Chair of Microbiology, Jagiellonian University, Medical College Cracow, Poland
Head of Department: prof. dr hab. med. Anna B. Macura
2Department of Food Chemistry and Nutrition, Jagiellonian University, Medical College, Pharmacy Faculty, Cracow, Poland
Head of Department: dr hab. Paweł Zagrodzki
3Department of Pomology and Apiculture, Agricultural University, Cracow, Poland
Head of Department: prof. dr hab. inż. Marek Grabowski
4Department of Epidemiology of Infections, Chair of Microbiology, Jagiellonian University, Medical College Cracow, Poland
Head of Department: dr hab. med. Małgorzata Bulanda, prof. UJ
5Department of Medical Diagnostics, Jagiellonian University, Medical College, Pharmacy Faculty, Cracow, Poland
Head of Department: dr hab. Ryszard Drożdż
A lot of Cornaceae family plants are used in a traditional medicine. Few studies described its antimicrobial activity, but only bark and fruit extracts were investigated so far. In our study ethanol or methanol extracts from different part of cornelian cherry: bark, fruits, leaves and seeds were used to evaluate their activity against 4 species of bacteria: Gram-positive – Staphylococcus aureus and Streptococcus pyogenes, Gram-negative – Escherichia coli and Pseudomonas aeruginosa and 3 species of fungi – Candida albicans, Aspergillus fumigatus, Trichophyton mentagrophytes. It was found that the highest antimicrobial activity obtained with disc-diffusion method revealed leaves and seeds extracts against S. aureus and C. albicans with diameters of inhibition zones between 10-15 mm. S. pyogenes and T. mentagrophytes were resistant to all extracts. In case of P. aeruginosa and E. coli the most effective was ethanol seed extracts.Our results show cornelian cherry as one from small know plants with interesting properties. This effect can be useful as new potential method of food protection before biological damage and also as new type of antibacterial and antifungal drugs.
Men by whole life meet from bacteria and fungi. One of them can be useful for peoples, another are unimportant and some of them are dangerous. They spoil food and may cause human and animal diseases. Therefore continually are searched new biological products which can brake or destroy their development, being simultaneously harmless for men. In the recent years have increased interest in research of antimicrobial activity of medicinal plants (1). But there are only a few studies on antimicrobial activity of cornelian cherry (Cornus mas L.) one from Cornaceae family plant focused only on bark and fruits of this plant (2, 3).
Cornelian cherry is a shrub or small tree of height from 3 to 8 m; it blooms very early – from February to March, fruits usually are ripened in September. The fruit is of reasonable size, up to 25 mm long, with a single large seed (endocarp). Fruit are used fresh, dried or in preserves. They are juicy, with a nice acid flavour. Roasted seeds are ground into a powder and used as coffee substitute. They are also suitable for oil extraction. The leaves are a good source of tannin. Cornelian cherry is widely used in Chinese and American Indian traditional medicine. The bark and fruits are used as astringent, febrifuge and nutritive. Fruits are good for bowel complaints and fevers, some authors pointed out their curative effect to cholera. The flowers are used against diarrhoea (4). Fruit extracts are also used for cosmetic purposes, replacing synthetic astringent substances, so they are claimed to exert a favorable action on the human complexion (5).
The food is the second target/place for antimicrobial substance. In the present moment with regard on demand on food with prolonged durability complies different methods of protection before growing of bacteria or fungi. Often applied synthetic agents are harmful for human especially they cause allergies (7, 8). From this reason seeks different additions to food which can eradicate bacteria and fungi being simultaneously harmless for men. Maybe substances isolated from cornelian cherry can be applied to food protection in future.
The aim of this study was to evaluate the antimicrobial potential of cornelian cherry.
Material and Methods
Biological materials and extracts preparation
All plant materials: fruits, seeds (September – October 2008), leaves (June 2008) and bark (February 2009) used in this experiment were obtained from experimental orchard in Garlica Murowana near the Cracow, Poland. This orchard is a property of Agriculture University of Cracow. Fruits were collected at the end of September and frozen at -20°C. Pulp was separated from seeds by manual crushing of defrosted fruits. After this, pulp was shaken with methanol or ethanol for 24 h, then centrifuged for 15 min at 5000 rpm and freeze drying until dry mass.
Seeds, leaves and bark were frozen with fluid nitrogen and powdered. Powder was shaken with methanol or ethanol for 24 h, then centrifuged for 15 min at 5000 rpm and freeze drying until dry mass.
Microorganisms and media
In this study four bacteria species were used, 2 Gram-positive and 2 Gram-negative: Staphylococcus aureus ATCC 25923, Streptococcus pyogenes and Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853 respectively and 3 species of fungi: yeast – Candida albicans ATCC 90028, mould – Aspergillus fumigatus and dermatophyte – Trichophyton mentagrophytes ATCC 18748.
Each microorganism prior to the antimicrobial testing was cultured on suitable media: Blood Agar for bacteria and Sabouraud Glucose Agar for fungi. The disk diffusion test was made on Mueller Hinton Agar (Biocorp) supplied with 2% glucose and Mueller Hinton Agar supplied with 2% glucose and 5% sheep blood for S. pyogenes.
Agar disk diffusion method
Blank paper disks (6 mm in diameter, EMPOL, Poland) were impregnated with 20 μl of individual extracts in three concentrations of each extract: 10 mg/ml, 1 mg/ml and 0.1 mg/ml to obtain 200 μg, 20 μg and 2 μg of extract respectively per disk. The disks were allowed to dry at room temperature. The strain to be tested was initially suspended in a sterile distilled water to a turbidity matching 0.5 McFarland standard with densitometer DEN-1 (BioSan). The prepared inocula were swabbed onto the surfaces of Mueller Hinton Agar plates with the sterile swab, and left to dry at room temperature. Disks with extracts were placed manually onto inoculated agar plate. The plates were incubated in an inverted position under conditions appropriate for the organisms tested. The plates with bacteria were incubated at 37°C for 24 h, while for Candida, Aspergillus and Trichophyton at 27°C for 48 h, and in case of moulds for the moment in which fungal growth could be observed with naked eye: 5 and 7 days, respectively. The inhibition zones were recorded.
Among examined microorganisms the most susceptible was Staphylococcus aureus strain. The six from eight investigated extracts were effective against it. Activity have shown, both methanol and ethanol extracts of seeds and leaves (zones 12 mm) and ethanol extract of bark (zone 9) and methanol extract of fruits (zone 8 mm). Activity against P. aeruginosa have shown five extracts (ethanol extract of seeds, leaves and bark – zone 10, 9, 7 mm respectively, and both methanol and ethanol extract of fruits – zones 7 and 8 mm respectively). The half of investigated extracts were active against E. coli and C. albicans. In the case of E. coli were ethanol extracts of seeds and leaves (zones 10, 7 mm respectively) and methanol and ethanol extracts of fruits (both 8 mm zones) and in the case of C. albicans effective were both methanol and ethanol extracts of seeds and leaves (zones 12, 10, 15, 12 mm respectively). A. fumigatus was susceptible only to methanol extracts of seeds. Resistant to all extracts were S. pyogenes and T. mentagrophytes.
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