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Journal of Food, Agriculture & Environment Vol.6(2) : 134-139. 2008

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Antimicrobial activity of selected Jordanian medicinal plant extracts against pathogenic microorganisms
Basem F. Dababneh
Department of Nutrition and Food Processing, Faculty of Agricultural Technology, Al-Balqa’a Applied University, P.O.Box 7474, Al-Salt 19117, Jordan. *e-mail: bdababneh@bau.edu.jo, basemdababneh@yahoo.com

Received 14 December 2007, accepted 25 March 2008.

Abstract
Antimicrobial activity of crude extracts from five commonly used medicinal plants in Jordan, Teucrium polium, Dianthus caryophyllus, Carthamus tinctorium, Ammi visnaga and Artemisia herba–alba were evaluated against four pathogenic microorganisms over a wide range of concentrations (50-5000 ppm). Minimum inhibition concentration (MIC) and the diameter of inhibition zone (DIZ) were determined by in vitro bioassays using hole-plate diffusion method against Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli and Candida albicans. All tested crude plant extracts significantly exhibited antimicrobial activity and inhibited the growth of Gram-positive and Gram-negative bacteria as well as C. albicans. The only exception was T. polium which showed no activity against the tested fungus at all tested concentrations. Antimicrobial activity was directly proportional to the tested concentrations. The most active antimicrobial effect was recorded for D. caryophyllus extract against C. albicans at MIC 800 ppm (DIZ = 25 mm), S. aureus at MIC 2000 ppm (DIZ = 20 mm) and P. aeruginosa at MIC 2000 ppm (DIZ = 10 mm). A. visnaga extract possessed the highest inhibitory effect on E. coli at MIC 800 ppm (DIZ = 9 mm). This study shed the light on the ability of extracts from Jordanian medicinal plants to combat pathogens which will help as natural antimicrobial agents as well as can be used in pharmaceutical . and food preservation systems. Key words: Natural antimicrobial, Jordanian medicinal plants, antibacterial activity, antifungal activity, MIC, DIZ.

Introduction Medicinal plants have been prescribed and used with a strong belief in their ability to cure diseases for centuries 48. Over the past 20 years, there has been a lot of interest in the investigation of natural materials as sources of new antibacterial agents 6, 50, insecticidal, acaricidal and cytotoxic activity 16. Plants used in traditional medicine contain wide range of substances to treat chronic as well as acute diseases. The substances that can either inhibit the growth of microorganisms or kill them are commonly considered for developing new drugs for treatment of various infectious diseases 25. Many plant species are considered as potential resource for treating diabetes and various diseases in skin, liver, digestive and the urinary system 44. Herbal medicine in the developing countries has evolved as an alternative solution to health problems as a cheap source 25, 36. Therefore, medicinal plants are intensely screened and tested for a wide range of applications including pharmacology, pharmaceutical botany, medical and clinical microbiology, phytopathology and food preservation 12. Research on plants used as remedies in traditional folk medicine can lead to identification of several biologically active molecules from the 250,000 documented higher plant species 33. The success achieved using medicinal plants and herbal formulations therapeutically based on ethnomedicinal and traditional use against a number of bacterial infections, raises optimism about the future of phytoantibiotics. Based on the indigenous and local knowledge, plants represent a rewarding
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untapped source with a significant potential for developing antimicrobial agents 43. Jordan is very rich in botanical diversity with more than 2000 wild plant species belonging to about 700 genera. As many as 485 species from approximately 99 plant families are categorized as medicinal plants 37. Biologically active compounds and extracts isolated from many plants species used in traditional herbal medicine in Jordan have been the center of interest 2. However, few studies on the antimicrobial activities of the Jordanian medicinal plants were carried out. Therefore, there is still a potential need to screen their effects on various pathogenic microorganisms. The aim of this study was to investigate the antimicrobial inhibitory effect of ethanolic crude extracts obtained from five medicinal plants commonly used in traditional medicine in Jordan. Extracts of Teucrium polium , Dianthus caryophyllus , Carthamus tinctorius, Ammi visnaga and Artemisia herba–alba were used against four microorganisms ( Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli and Candida albicans). Materials and Methods Plant materials: Five medicinal plants namely Teucrium polium, Dianthus caryophyllus, Carthamus tinctorius, Ammi visnaga and Artemisia herba–alba were either collected from the field or purchased from local market. Scientific name, English common
Journal of Food, Agriculture & Environment, Vol.6 (2), April 2008

Sesquiterpene lactones 7. Al-Gabbeish. The values reported for DIZs were the average of three replicates. A. 400. aureus (Table 2). Preparation of plant extracts: Plant material was dried in the shade at room temperature and powdered using an electric mill. Phytotoxic and antimicrobial 53. Dianthramides 39. 15 Active constituents Diterpenoids9. The microorganisms were maintained on slants of nutrient agar (NA) at 4°C.25-1. 600. digestive function stimulant. Ihab H. antifungal 11. Journal of Food. Anti-inflammatory 22. wormwood. C. bishops weed. tooth pain. 100 and 50 ppm. albicans was spread on the surface of potato dextrose agar (PDA). Al-Salt College-Head Department of Biological Sciences. coli (Table 4) and C. antibacterial and anti-inflammatory 33. 2000. The mixture was shaken daily for regular infusion. and anti-allergic 18. Monoterpenes 21. Agriculture & Environment. false saffron. P. bastard. antispasmodic. Vasodilator and Coumarins 13. Carthamine 29. Common medicinal uses Diuretic. Antiretroviral proteins 31 and phenols 11. Antipyretic and anti-bacterial 34.name and parts used regarding each plant are summarized in Table 1. Vol. April 2008 135 . treatment of hyberlipemia. Family: Compositae (Asteraceae) Scientific name: Artemisia herba-alba Used part: Aerial parts English name: Herba-alba.6 (2). The dried plant extracts were dissolved in dimethylsulfoxide (DMSO) to provide a stock solution with the final concentration of 5000 ppm. caryophyllus. saffron thistle. Skin toner 38. tinctorium. LSD analysis was used to compare means. Minimum inhibition concentrations (MICs) were taken as the lowest concentration at which observable growth was inhibited with no significant differences at higher concentrations used. annexin 19 and placenta protein 4 42. Ethnobotanical data and active constituents of studied plants. Screening for antimicrobial activity: Hole-plate diffusion method was used for studying the antimicrobial activity and determining the minimum inhibition concentrations (MICs) 8. Hypoglycemic 51. Flavonoid glycosides 54. bioactivity was determined by the measurement of the diameter of inhibition zone (DIZ) around each hole in mm. diaphoretic. eye diseases. relief of acute dermatitis. antimicrobial immunomodulating. Anti-nociceptive 4. night sweat and dizziness 17. tonic. anti ulcerogenic. Hypolipidemic 40. Each hole (diameter 6 mm) was filled with 50 µl from each dilution of plant extract. fever. Control plates received only DMSO in NA and PDA without plant extracts and were run following the same procedure as above. aeruginosa (Table 3). santonica. Significant differences were defined at p≤0. Antioxidant 10. On the sixth day. albicans (Table 5). Three holes were made on the media using 6 mm diameter sterile cork-borer. Two hundred and fifty grams of each plant powder were soaked in 1. anti-pyretic. tooth pick. The inoculated agar plates were incubated at 37°C for 24 hr. 200. Bronchodilator muscle relaxant 13. 1000. the extract was filtered using Whatman filter paper No. The inoculums were incubated overnight in nutrient broth at 37°C to produce dense microbial suspension of approximately 106cfu/ml. polium. Results and Discussion Extracts obtained from T. crisiol 26. A range of serial dilutions were prepared from the stock solution to provide 4000. sesquiterpenoids 24. 3000. white mugwort. Each inoculum from dense bacterial suspension containing 106 bacterial cells/ml was spread on the surface of nutrient agar medium (NA) while the C. The common medicinal uses and active constituents of the plants under this study are reviewed and well documented (Table 1). wormseeds. Anti-inflammatory and anti-rheumatic 49. 1. Statistical analysis: Data of DIZ are presented as means of three replicates and analyzed using factorial arrangement in complete random design (CRD) with SAS version 9 software package 46. E. hair pomades. arteriosclerosis. antihelminthic 7 Falconoid 45. Treatment of neurological and cardiac disorders and sexual weakness 45. Alkaloids 33. anti-spasmodic and cholagogic 20 . kellin. D. Botanical source Family: Labiatae (Lamiaceae) Scientific name: Teucrium polium Used part: Aerial parts English name: Mountain germander Family: Caryophyllaceae Scientific name: Dianthus caryophyllus Used part: Petals English name: carnation. fatty acids 27. Flavonoids 11. anthocyanins 35. Al-Balqa’ Applied University. anti-tumor. osteoporoses and bone resorption. herba-alba were tested against S. Microbial test suspensions: Test organisms used in this study are Gram-positive bacteria (Staphylococcus aureus). Hypoglycemic 14. Hypoglycemic 1 and in the treatment of jaundice 32. vomiting and gasteritis. shih. The inhibition zone was recognized as the area surrounding the hole with no growth of the tested pathogens. visnaga and A. iridoids. Antispasmodic. . Dry crude extract was produced by evaporating ethanol under low pressure using a rotary vacuum evaporator at 60°C. 150. Family: Umbelliferae (Apiaceae) Scientific name: Ammi visnaga Used part: Fruits English name: Khella.05. Haemostatic agent. clove pink Family: Compositae (Asteraceae) Scientific name: Carthamus tinctorius Used part: Flowers English name: Safflower. Flavonoids. Treatment of yin deficiency of liver and kidney. After the incubation period. 800. diuretic 33. neoclerodane diterpenes 5. Gramnegative bacteria (Pseudomonas aeruginosa and Escherichia coli) and yeast (Candida albicans). Antimicrobial activity was determined by measuring the diameter of inhibition zones (DIZ in mm) and the minimum inhibitory Table 1. The identified clinically resistant strains were obtained from the Jordan University Hospital.5 L of 95% ethanol for 5 days at room temperature. Authentication and identification of plant material was confirmed by Dr. cardiac tonic and diuretic 28. promoting blood coagulation. The final crude extracts were stored in labeled sterile glass vials at -20°C until used for the antimicrobial activity test 23.

plants and extract concentrations were highly significant at p≤0. All tested plant extracts demonstrated broad spectrum antimicrobial activity against tested microorganisms with variable inhibitory effect. polium: E. albicans. herba-alba at MIC 4000 ppm (DIZ = 13 mm). tinctorium 0f 0f 0f 0f 0f 0f 11 fghij 11 fghij 13 defgh 14 cdefg 15 bcdef 15 bcdef 15 bcdef 1.05) at plant extract concentrations used. 100 and 50 ppm. visnaga : E. Antimicrobial activity (DIZ mean in mm) of medicinal plants at various concentrations against Escherichia coli*.25 D. D. 150. Table 4. except T. herba. This could be related to the variations in the quality and quantity of active compounds in the plant extracts (Table 1). visnaga 0f 0f 0f 0f 0f 0f 0f 10 bcde 10 bcde 10 bcde 11 abcd 11 abcd 12 abcd 4.53 A. polium had no antifungal effect at all tested concentrations. herba. polium and A. polium 0f 0f 0f 0f 7e 7e 8 de 8 de 8 de 8 de 9 cde 10 bcde 10 bcde 4. f i li di • Means with different superscript letters are significantly different (P≤0. caryophyllus had a remarkable inhibitory effect at MIC 1000 ppm with DIZ 15 mm. All tested plant extracts were active against tested bacteria with variable inhibitory effects. D. The interaction effect of microorganisms. visnaga 0f 0f 0f 0f 0f 7e 7e 9 cde 9 cde 9 cde 9 cde 9 cde 9 cde 10. visnaga inhibited P.53 D. 600.65 * Values were mean of triplicate readings. tinctorium and A. aeruginosa at MIC 3000 ppm with DIZ 11 mm. coli > S. aeruginosa > S. 4000. visnaga 0f 0f 0f 0f 7j 7j 7j 8 ij 12 efghi 12 efghi 13 defgh 15 bcdef 15 bcdef 4. tinctorium at MIC 1000 ppm (DIZ = 9 mm). tinctorium 0f 0f 0f 0 f 0f 7e 8 de 8 de 10 bcde 12 abcde 12 abcde 13 abcd 13 abcd 1. 2000. aureus.95 C. caryophyllus 0f 0f 8 ij 8 ij 9 hij 13 defgh 14 cdefg 17 abcd 18 abc 20 a 20 a 20 a 20 a 5. P. respectively.48 A. Concentration of plant extract (ppm) 0 50 100 150 200 400 600 800 1000 2000 3000 4000 5000 LSD T. aureus. herba-alba caused inhibitory effect at MIC 3000 ppm with DIZ ranging from 14-16 mm. 3000.61 A. herba-alba: S. aureus > C.alba 0f 0f 0f 0f 0f 0f 0f 0f 0f 8 ij 16 abcde 16 abcde 16 abcde 3. The extract of A. • Means with different superscript letters are significantly different (P≤0. Agriculture & Environment. The growth of C. albicans. coli > C. A. DIZ 9 mm) against E. D. visnaga required higher MIC (4000 ppm) to reach similar inhibition effect. Concentration of plant extract (ppm) 0 50 100 150 200 400 600 800 1000 2000 3000 4000 5000 LSD T. A. polium 0f 0f 0f 0f 0f 0f 8 de 9 cde 9 cde 10 bcde 10 bcde 10 bcde 10 bcde 7. aureus > E.7 * Values were mean of triplicate readings. coli. 400. polium. coli > P. albicans > P.91 A. coli.37 A.aureus. Concentration of plant extract (ppm) 0 50 100 150 200 400 600 800 1000 2000 3000 4000 5000 LSD T. C. tinctorium 0f 0f 0f 7e 7e 7e 7e 8 de 8 de 13 abc 13 abc 13 abc 13 abc 5. aeruginosa > C. herba-alba both had the lowest inhibition activity in comparison to plant extracts tested on E. aeruginosa and S.05) at plant extract concentrations used. A. caryophyllus exhibited significantly the highest antibacterial activity at MIC 2000 ppm with DIZ 20 mm. 1000.54 * Values were mean of triplicate readings. tinctorium: C. followed by C. polium 0 f• 0f 0f 0f 7j 7j 9 hij 10 ghij 11 fghij 13 defgh 14 cdefg 14 cdefg 14 cdefg 5. Overall MICs of active plant extracts and diameter of inhibition zones are shown in Table 6.05) at plant extract concentrations used. C. April 2008 . A. Vol.14 A. herba. coli (Tables 4 and 6). polium had no anticandidal effect at all tested concentrations (Tables 5 and 6). caryophyllus 0f 0f 0f 7e 7e 8 de 8 de 9 cde 9 cde 10 bcde 10 bcde 10 bcde 10 bcde 5. albicans > E. aureus > P. aeruginosa > S . caryophyllus 0f 0f 9 cde 10 bcde 10 bcde 13 abcd 14 abc 14 abc 15 ab 15 ab 15 ab 15 ab 15 ab 4.1 D. while T. C. coli. 200. tinctorium and A.6 (2). albicans was considerably inhibited by D. caryophyllus extract at MIC 800 ppm (DIZ = 25 mm). Table 3. caryophyllus and C.alba 0f 0f 0f 0f 0f 0f 0f 0f 0f 8 de 14 ab 16 a 16 a 5.05. T. visnaga significantly revealed the highest activity (MIC 800 ppm. The inhibition activity of plant extracts on test strains was in decreasing order according to the minimum inhibition concentration as follows: T. Antimicrobial activity (DIZ mean in mm) of medicinal plants at various concentrations against Pseudomonas aeruginosa. 800. polium possessed moderate activity at 2000 ppm with DIZ 10 mm. aureus and E. Plant extracts showed different antimicrobial activity against the tested pathogens and the diameter of inhibition zone was directly proportional to the increase in plant extract concentration reaching a plateau. Previous papers indicated that antimicrobial activity of botanical extracts is related to the presence of 136 Journal of Food. herba-alba required higher concentration (MIC 4000 ppm) to cause similar effect (Tables 3 and 6). visnaga at MIC 2000 ppm (DIZ = 25 mm) and A. • Means with different superscript letters are significantly different (P≤0. T. caryophyllus: C. T. concentration (MIC in ppm) over a range of concentrations 5000.63 C. Table 2 shows the screening test of plant extracts on S.Table 2.76 C. Antimicrobial activity (DIZ mean in mm) of medicinal plants at various concentrations against Staphylococcus aureus*. tinctorium which inhibited at MIC 2000 ppm with DIZ 10 and 13 mm. albicans > P. P. aeruginosa (Table 3) had a marked significant sensitivity towards both D.alba 0f 0f 0f 0f 0f 0f 0f 0f 0f 0f 13 abcd 17 a 17 a 3. A.

Antimicrobial activity (DIZ mean in mm) of medicinal plants at various concentrations against Candida albicans*. Earlier results 30.. 5 Bedir. N. tinctorium D. II. Calis. Agriculture & Environment. 1990. MIC=Minimum inhibition concentration (ppm). R. coli C. R. From the above findings it could be concluded that the tested plant extracts exhibit a broad spectrum of activity against various microorganisms. Mohsen for technical assistance and Ahmad Al-Gabbiesh for his continuous support. 1996. bacteriostatic. Hypoglycaemic effect of Artemisia herba-alba. as we believe is of great importance. M. 1 A. Table 6. S. Vol. Further investigations to determine bactericidal. K.. 3 Bai. 47 support our findings that the composition of essential oil depends on the plant species. S. tinctorium A. albicans MIC 3000 4000 2000 N. fungicidal or fungistatic effects are recommended with emphasis on the identification of the active antimicrobial chemical constituents of these commonly used Jordanian medicinal plants. Ghasemzadeh.A=Not active. and Sticher. G.. H. • Means with different superscript letters are significantly different (P≤0. Mayyada B. 1994.6 (2)... These differences could be attributed to structural nature of the microorganisms 52 and plant constituents.herba-alba Plant species C. A. H. and Twaij. Minimum inhibitory concentration and diameter inhibition zone of crude plant extracts against pathogenic microorganisms. Fitoterapia LXVII(5):435-442. The absence of antifungal effect against C. Zerbe. Traditional Chinese material: A respect and prospect. M.alba Table 5. Antinociceptive effect of Teucrium polium leaf extract in the diabetic rat formalin test.visnaga D. albicans. visnaga C.. * Values were mean of triplicate readings. M. Phytochemistry 51:921925. Fungitoxic property of D. flavonoids. T. N.05) at plant extract concentrations used. anthocyanins and terpenoids in addition to other compounds of phenolic nature or containing free hydroxyl groups 41. D. which lead to variation of their antimicrobial activities. E. Heydari. Shehadeh for insightful discussion and revising the manuscript..67 f 0 0f 0f 0f 0f 0f 0f 0f 0f 0f 0f 13 d 13 d 2 f 137 . A. A. and Najib. Reem M. April 2008 Concentration of plant extract (ppm) 0 50 100 150 200 400 600 800 1000 2000 3000 4000 5000 LSD 0 0f 0f 0f 0f 0f 0f 0f 0f 0f 0f 0f 0f - f 0 0f 0f 9 de 11 de 11 de 21 bc 25 b 25 b 25 b 25 b 26 b 26 b 8. T. 2005. L. H. The optimal effectiveness of medicinal plants may not be due to one active constituent. However. Neoclerodane diterpenoids from Teucrium polium. Nik. F.different chemical agents including essential oils. Effect of a valuable extract on some blood parameters in diabetic animals. A. caryophyllus MIC 2000 2000 1000 800 DIZ=Diameter of inhibition zone (mean in mm). Results from this study indicate that tested extracts possessed variable antimicrobial effects against both Gram-positive and Gramnegative bacteria as well as C. A. O. Al-Khazraji. O. 6 Bonjar. Batshoun. and Roghani-Dehkordi. caryophyllus against Fusarium oxysporum was reported to be due to a group of phenolic constituents such as kaempferide triglycoside 11. aureus P. S.76 f 0 0f 0f 0f 0f 7e 7e 7e 9 de 9 de 9 de 10 de 10 de 2.. polium Journal of Food.43 f 0 0f 0f 0f 0f 10 de 10 de 11 de 18 c 25 b 25 b 25 b 25 b 6. Tasdemir. aeruginosa E. 1999. Results of the present study should be considered for the possible application of plant extracts as natural bacteriostatic and bactericidal component in various products and as natural preservatives extending the pharmaceutical and dietary products shelf life. 4 Baluchnejadmojarad. Owais. D. polium ethanolic extract might be related to absence of active constituents or needed to use higher concentrations. albicans by the range of tested concentrations of T. caryophyllus T. Special thanks to Dr. I. but to the combined actions of different plant constituents 3. the chemotypes and the climatic conditions. Khalaf for her laborious work. Biological activity of some Jordanian medicinal plant extracts. Journal of Ethnopharmacology 97:207-210. the plants differed significantly in the activity against tested microorganisms. Moreover. Such postulation requires further investigations. References Al-Shamaony. polium Microorganism A.A DIZ 20 10 15 25 MIC 3000 2000 4000 1000 DIZ 15 13 13 9 MIC 4000 3000 800 2000 DIZ 15 11 9 25 MIC 3000 4000 4000 4000 DIZ 16 14 17 13 Acknowledgements The author wishes to express his appreciation and thanks to Etekal Z. Planta Medica 56:502. Journal of Ethnopharmacology 243(3):167-171. M.A DIZ 14 10 10 N. the differences observed in antimicrobial activities of the investigated plant extracts suggest the susceptibility variations of microorganisms to various chemical components.. W. Roghani. 2 Alkofahi... herba.

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