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African Journal of Traditional, Complementary and Alternative Medicines
African Ethnomedicines Network
ISSN: 0189-6016
Vol. 8, Num. 2, 2011, pp. 159-164

African Journal of Traditional, Complementary and Alternative Medicines, Vol. 8, No. 2, 2011, pp. 159-164

DETECTION OF THE ANTIBACTERIAL EFFECT OF NIGELLA SATIVA GROUND SEEDS WITH WATER

Hussein Ahmed Bakathir and Nageeb Ahmed Abbas

Department of Pharmacology – Faculty of Pharmacy University of Aden *E-mail: bakathirhussin@yahoo.co

Code Number: tc11022

Abstract

Nigella Sativa (NS) seeds have been used for medicinal purposes for centuries both as herbs and its oil. In Islam it is regarded as one of the greatest forms of healing medicine included in the medicine of prophet Mohammed. Huge number of studies have been carried out in recent years on the pharmacological effects of these seeds and also the possible relationship with their constituents. A number of these investigations emphasized the antimicrobial effect of them by using different extracts. In our study we have tried to use the normal human mechanism in digestion by using the ground seeds. A modified paper disc diffusion method was used to test the antibacterial effect of NS seeds. Clear inhibition of the growth of Staphylococcus aureus was observed by concentration of 300mg/ml with distilled water (D.W.) as control , this inhibition was confirmed by using the positive control Azithromycin. The inhibition obtained was higher with Nigella sativa ground seeds from Hadramout (HNSGS) than with Nigella sativa ground seeds from Ethiopia (ENSGS). No inhibition was found in the growth of E.Coli and Enterobacter. This was emphasized by using the positive control Ciprofloxacin. The positive inhibition may be attributed to the two important active ingredients of NS, Thymoquinone and melanin.

Key words: Nigella Sativa , antibacterial effect , Staphylococcus aureus, paper disc-diffusion method.

Introduction

The seeds of Nigella Sativa Linn. (Ranunculaceae ), commonly known as black seed or black cumin has been used for medicinal purposes for centuries both as herb and pressed into oil in Asia, Middle East, and Africa (Batheeb; Zohary and Hopf, 2000). It has been traditionally used for a variety of conditions and treatments related to respiratory health, stomach and intestinal health, kidney and liver function, circulatory and immune system support and general well-being (Khan et al., 2003). In Islam, it is regarded as one of the greatest forms of healing medicine available and included in the medicine of the Prophet Mohammed (Aljawezjjah, 2001). Its oil has been used in treatment of dermatological diseases e.g. eczema and boils (Zohary and Hopf, 2000). These many therapeutic uses earned Nigella Sativa the Arabic approbation (habbatul barakah the seed of blessing ).

Huge number of studies and researches have been carried out in recent years on the pharmacological effects of these seeds covered both fields of experimental and clinical pharmacology; not only in the use of these plants but also in the investigation of the constituents and active ingredients that may explain or interpret these pharmacological actions. One of these active ingredients is Thymoquinone (volatile oil of these seeds) and Melanine (fixed oil) (Roy et al. 2006; Adel, 2006). Some investigations performed to show the possibility of antimicrobial and antibacterial activities of these seeds using their extracts or oil (Ali and Blunden, 2003; El.Fatatry, 1975; Hanafy and Hatem, 1991; Morsi, 2000; Roy et al., 2006). Modern studies have been done to investigate immuno-modulatory, immunosuppressive and anticancer properties of these black seeds (Islam et al., 2004; Mbarek et al., 2007; Mohammed, 2005; Adel, 2006).

The antioxidant action of thymoquinone and its 5-lipoxygenase inhibition may explain the different anti-inflammatory effect of these seeds. Interestingly, it was found that the fixed oil of N.Sativa had both antioxidant and anti-eicosanoid effect greater than thymoquinone which is its active constituent (Ali and Blunden, 2003).

The aim of our study is to find out a method for testing the antibacterial effect of these seeds by using the ground seeds the way it used in the local human traditional diet.

Materials and methods

Method

A modified disc diffusion method was used for determination of the antibacterial activities of the Nigella Sativa ground seeds with water(NSGSW ), and the following steps were carried out:

  1. The equipment used were cleaned and sterilized by immersing them in ethanol for 10 min then washed with distill water(D.W.) and dried adequately by using sterile bandage.
  2. Nigella sativa HNS and ENS were ground by the grinding machine and then put in two separate bottles, one for HNS and the other for ENS.
  3. 0.75gm of both type of Nigella sativa from the two separated bottles was measured by using sensitive balance, and transferred with aid of spatula into separate test tubes covered with foil. 2.5ml D.W. was added to each test tube in microbiology laboratory..
  4. 2 agar plates were taken and divided to equal 4 regions or 3 regions as required.
  5. The nutrient broth was inoculated with a loopful of the selected bacteria and the plain agar was also inoculated with the broth containing this bacteria.
  6. The test tubes were shaken vigorously to form homogenous mixture and in first plate ENSGSW in region (1) 5 µl of mixture was added, in region (2) 10 µl mixture and in region (3)20 µl of mixture was added, in region (4) 10ml of D.W used as a negative control and in the second plate, by using 20 micropipette, 20 µl of HNSGSW was placed to region (1) and 20µl of ENSGSW to region (2) in region (3) positive control was added (Antimicrobial standard Azithromycin and Ciprofloxacin).
  7. Finally, the tested plates were placed in the incubator at 37ºC for 24 hrs, and for the assessment of the results, pictures of the Petri plates were taken at the end of each experiment and the inhibition zones around each paper disc (5mm diameter, filter paper) were measured and recorded.

Results and discussion

The identification of the antibacterial effect was achieved by taking different concentration of N.S. from the two preparations (HNSGSW and ENSGSW ) and applying it directly or by direct mixing with D.W. The concentration which produced best effect was 300mg/ml (Figures 1, 2, 3). Figure 3 showed the comparison of the inhibition caused by the different concentrations of both preparations. The largest inhibition was observed by the HNSGSW that reached 24 mm. No inhibition was observed by the negative control (D.W) (Figures 1 and 2). These positive effects of both preparations were emphasized by using the positive control antibiotic Azithromycin (Figure 4 and Table 3). These positive effects are in agreement with previous findings by other authors (El.Fatatry, 1975; Hanafy andHatem, 1991; Mashhadan and Rakhshandeh, 2005; Morsi, 2000; Roy et al., 2006; Adel, 2006).

Mashhadan found that the aqueous extract did not show any effect but other extracts (methanol, chloroform) showed high inhibitory effect against all the tested microorganisms including Staphylococcus, where concentration used ranged from 62.5mg/ml to 1g/ml (Mashhadan and Rakhshandeh,2005). Morsi had proven that both the crude alkaloid extract and the water extract of the seeds were effective against some tested microorganisms like staphylococcus despite their resistance to other antibiotics (Ali and Blunden, 2003; Morsi, 2000). Probably these variations could be explained by the different extraction methods, and also the amount of ingredients of the same plant can be affected by the area and season of collection. A combination of garlic with NS was tested individually and in combination for their antimicrobial activities against Staphylococcus where both showed modest antimicrobial effects (Roy et al., 2006). NS of both origins caused no inhibitory effect on the growth of E.Coli using D.W as control (Figures 5 and 6). The extract had no effect also on the growth of Enterobacter (Figure 7 and Table 4 ). Zuridah H. and et al found also negative effect of N.S. on the growth of E. coli by using methanolic extract and concentration 25mg/ml (Zuridah et al., 2008). Although the mechanism of the antimicrobial effect of these seeds has not been reported, its antimicrobial action could be attributed to the active ingredients especially thymoquinon and melanin (Roy et al., 2006; Adel, 2006).

Tables 1 & 2

Conclusion

According to the method used, a clear and undeniable antibacterial effect caused by the N.S. ground seeds on the growth of Staphylococcus was obtained. The inhibition of the Staphylococcus growth is higher with NSGSW from Hadhramout than with NSGSW from Ethiopia.

References
  1. Abn Alqaijem Aljawezjjah (2001). Altteb alnabawi, study and comment: Ammar Zaki Albaroodi, first edition, Alttawfeeqiah library, 291 – 293, Eygypt.
  2. Adel Majthoob Hassieb (2006). Melanin is the secret of Nigella Sativa Al-Faisal Scientific Journal, 4(3): 110 – 143.
  3. Ali, B.H. and Blunden, G. (2003). Pharmacological and toxicological properties of Nagila Sativa Phytother. Res.17:299– 305.
  4. Ali Salem Batheeb (2007). medicinal plants in Yemen, 2nd edition, Alershad library 132 – 133, Sanna, Yemen.
  5. El.Fatatry, H.M. (1975). Isolation and structure assignment of an antimicrobial principle from the volatile oil of Nagilla Sativa L. seeds , Pharmazie 30: 109 – 111
  6. Hanafy, M. S. and Hatem, M. E. (1991). Studies on the antimicrobial activity of Nigella Sativa seed (black cumin), J. Ethnopharmacol. 34:275 – 278
  7. Islam, Sk.N., Begum P., Ahsan, T., Hugue, S. and Ahsan, M.(2004). Immunosuppressive and cytotoxic properties of N.S, Phytother.Res. 18:395 – 398
  8. Khan, M. H., Ashfaq, M.K., Zuberi, H.S., Mahmood, M.S., Gilani A. H. (2003).The in vivo antifungal activity of the aqueous extract from N.S seeds Phytother. Res. 17: 183-186.
  9. Mashhadan, N.V. and Rakhshandeh, H.(2005). Antibacterial and antifungal effects of NS extracts against S.A, Pseudo.aeroginosa. and Candida albicans. Pak. J. Med. Sci., 21 (1): 47 – 52.
  10. Mbarek, L., Ait Mouse, H., Elabbadi, N., Bensalah, M., Gamouth, A., Aboufatima, R., Benharref, A., Chait, A., Kamal, M., Dalal, A., Zyad, A. (2007). Anti-tumor properties of black seed (Nigella Sativa)extract , Brazilian J. of Medical and Biological Research 40: 839 – 847.
  11. Mohammed Labib Salem (2005). Immunomodulatory and therapeutic properties of Nigella Sativa L. seed, International Immunopharmacology 5: 1749 – 1770
  12. Morsi, N.M. (2000). Antimicrobial effect of crude extracts of NS and multiple antibiotics-resistant bacteri, Acta Microbiol. Pol 49: 63-74.
  13. Roy, J., Shaklega, D., Callery, P. and Thomas, J.(2006). Chemical constituents and antimicrobial activity of a traditional herbal medicine containing garlic and black cumin ,Afr J. Tradit Complement Altern Med. 3(2): 1–7
  14. Zohary, D. and Hopf, M. (2000). Domestication of plants in the old world 3rd edition Oxford Univ. Press page:206
  15. Zuridah H. , Fairuz A.r.M., zakri A.h.Z., Rahim M.N.A. (2008). In vitro antibacterial activity of Nigella Sativa against Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumonia, E. coli and B.cereu, Asian Journal of Plant Sciences 7(3): 331 – 333.
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