Research Journal of Chemical Sciences ______________________________________________ ISSN 2231-606X Vol. 1(8), 80-82, Nov. (2011) Res.J.Chem.Sci. International Science Congress Association 80 Short CommunicationAntibacterial Potential of Achyranthus aspera Linn Procured from Himachal Pradesh, Punjab and Haryana, India Sharma Raj Neeta, Bala Jyoti, Singh Anjuvan and Kaur PrabhjotDepartment of Biotechnology, Lovely School of Bio-Sciences,Lovely Professional University, Phagwara, INDIA Available online at: www.isca.in (Received 09th August 2011, revised 21st August 2011, accepted 27th August 2011) Abstract Achyranthus aspera, Family: Amaranthaceae was procured from Himachal Pradesh, Punjab and Haryana regions in the month of January-2010. Methanol extract of dried whole plants of Achyranthus aspera was evaluated against bacterial species viz., Bacillus cereus, Escherchia coli, Acinetobacter baumanii Staphylococcus aureus, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa and Salmonella typhi. The plants procured from Himachal Pradesh showed the best antimicrobial activity followed by Punjab and Haryana region respectively. Escherchia coli was maximally inhibited (24mm±0.5) by the plants from Himachal Pradesh region followed by Klebsiella pneumoniae (23mm±0.7) and Bacillus cereus (20mm±0.51) at the concentration of 2000µg. The zone of inhibition could not be detected at 500 µg for Proteus mirabilis, Pseudomonas aeruginosa, from all the locations investigated and at 1000 µg also, no inhibition was observed in plant samples collected from Haryana region. All the bacterial species were found susceptible to methanolic extract of whole plants of Achyranthes aspera however at high concentrations. Keywords: Herbal antibiotic, achyranthes aspersa, weed, phytochemicals. IntroductionIndia is a land of rich biodiversity. The plants are potential source of medicines since ancient times. Medicinal plants play an important role in the development of potent therapeutic agents. Achyranthes aspera Linn belongs to the family Amaranthaceae. It is an annual, stiff erect herb, and found commonly as a weed throughout India and is one of the important medicinal plants having many therapeutical uses as odontalgic, Rheumatism, Bronchitis, skin disease and Rabies. Leaf decoction for cardiovascular toxicity has been reported and the methanol crude extract showed high antibacterial activity against Escherchia coli and Bacillus cereus. Root extract of the plant inhibited the growth of Bacillus cereus and Staphylococcus aureus bacterial strains. Seeds are rich in protein, cooked and eaten. Leaf extracts were reported to possess thyroid stimulating and anti peroxidative properties. Roots are also used as astringents to wounds, in abdominal tumor and stomach pain. The benzene extract of the stem bark showed abortifacient activity in the rat. Leaf extracts were reported to possess thyroid stimulating and anti-peroxidative properties. The aqueous and methyl alcohol extracts of the plant also decreased blood glucose levels in normal and alloxan diabetic rabbits . A leaf extract from Achyranthes aspera collected in different areas of the United Arab Emirates was tested against Grampositive bacteria. It showed inhibition against Staphylococcus aureus, Bacillus subtilis, E. coli and Aspergillus terreus, respectively. The root extract was less active. The present investigation was carried out to study the potential of methanolic extract of dried whole plants ofAchyranthes aspera procured from different locations such as Himachal Pradesh, Punjab and Haryana to screen the best strain of Achyranthes aspera in terms of high antimicrobial potential. The extract of dried whole plant was tested against Bacillus cereus, Escherchia coli, Acinetobacter baumanii, Staphylococcus aureus, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa and Salmonella typhi.Material and MethodsFresh plants of Achyranthes aspera Linn. were collected from three locations namely Punjab, Himachal Pradesh and Haryana in the month of January 2010. The plants were identified taxonomically and authenticated in the Department of Biotechnology, Lovely Professional University, Phagwara. Fresh plants were washed thoroughly 3-4 times with running tap water then finally with sterile water followed by shade drying at room temperature for 15-20 days. The dried plant material was made into coarse powder and passed through sieve and then used for crude extraction. Fine powder (20gm) was extracted in 100 ml of methanol at 50-55C for 24 hrs in shaker. The extract was filtered through Whattman filter paper No.1 and then concentrated Research Journal of Chemical Sciences __________________________________________________________ ISSN 2231-606X Vol. 1(8), 80-82, Nov. (2011) Res.J.Chem.SciInternational Science Congress Association 81 by using a rotary evaporator at low temperature (40-50°C) and reduced pressure. The extraction value in methanol was 800 mg. Further, the dried residue was preserved in airtight container and kept at 4-5°C until further use. The extract was then subjected to preliminary qualitative analysis of phytochemicals namely carbohydrate, alkaloid, Terpenoid, Protein (Biuret method), Flavonoids, Tannin and Saponin. The selected microbes were procured from Christian Medical College, Ludhiana, Punjab. The organisms were Escherchia coli, Bacillus cereus, Acinetobactor baumanii, Staphylococcus aureus, Pseudomonas aeruginosa, Kleibsiella pneumoniae, Proteus mirabilis and Salmonella typhi. Nutrient agar media was poured in Petri plates and then inoculum was spread with the help of L- spreader10. A well was made in the plates with sterile borer. The extract was introduced into the well and plates were incubated at 37°C for 24 hrs in B.O.D incubator. The experiment was performed under strict aseptic conditions. The antimicrobial property of the plant was checked against the different strains of bacterial species. Microbial growth was determined by measuring the diameter of zone of inhibition. Results and Discussion chyranthes aspera collected from Himachal Pradesh, Punjab and Haryana region were tested for the presence of carbohydrate, proteins, alkaloids, saponins, terpenoids, tannins and flavonoids and found positive for all the constituents table-1. The same has also already been reported earlier11. Methanol extract of whole plant of Achyranthes aspera was selected to test antibacterial activity against eight different microorganisms using agar well diffusion method. The efficiency of methanolic extract was furnished at 500µg, 1000µg and 2000µg. Plants procured from Himachal Pradesh showed maximum inhibition at 2000µg concentration. Escherichia coli was inhibited maximally (24 ± 0.5), followed by Klebsiella pneumoniae (23 ± 0.7), Bacillus cereus (20 ± 0.51), Acinetobactor baumanii (19±0.15), Pseudomonas aeruginosa (19 ± 0.2), Salmonella typhi (18 ± 0.53), Proteus mirabilis (18 ± 0.62), Staphylococcus aureus(16 ± 0.67) Table-2. Plants collected from Punjab region showed better inhibition of Klebsiella pneumoniae (20 ± 0.65) at 2000µg. Table-1 Phytochemical screening of Achytanthes aspera from Himachal Pradesh, Punjab and Haryana region S.No. Phytochemicals Whole Plant Extract in methanol Himachal Pradesh Punjab Haryana 1. Sugars + + + 2. Proteins + + + 3. Alkaloids + + + 4. Tannins + + + 5. Saponins + + + 6. Terpenoids + + + 7. Flavonoids + + + Table -2 Zone of inhibition of different test organisms against methanol extract of Achyranthes aspera procured from three different locations Test organism Zone of inhibition Himachal pradesh Punjab Haryana Himachal pradesh Punjab Haryana Himachal pradesh Punjab Haryana 2000µg 1000µg 500µg ml - 1 Bacillus cereus 20±0.51 17±0.34 15±0.6 12±0.38 7.3±0.13 8.0±0.56 8.5±0.51 0 0 Escherchia coli 24±0.5 19±.55 19±0.18 12.2±0.41 8.5±0.27 9.0±0.4 10.3±0.41 6.2±0.8 7±0.22 Acinetobactor baumanii 19±0.15 17±0.41 15±0.45 9.5±0.8 10±.34 0 7.7±0.34 7.2±0.67 0 Staphylococcus aureus 16±0.67 19±0.7 17±0.8 9.7±0.23 12±0.27 10±0.56 0 9.4±0.3 8.1±0.9 Klebsiella pneumoniae 23±0.7 20±0.65 20±0.56 13.3±0.7 10±0.55 11.4±0.23 9.5±0.27 8.7±0.52 8.0±0.63 Proteus mirabilis 18±0.62 16±0.5 10±0.25 9.2±0.43 8.4±0.45 0 0 0 0 Pseudomonas aeruginosa 19±0.2 16±0.13 14±0.6 12.4±0.27 10.8±0.7 0 0 0 0 Salmonella typhi 18±0.53 12±0.3 16±0.3 11.5±0.56 7.8±0.15 7.2±0.18 8.3±0.92 0 0 Research Journal of Chemical Sciences __________________________________________________________ ISSN 2231-606X Vol. 1(8), 80-82, Nov. (2011) Res.J.Chem.SciInternational Science Congress Association 82 Plants from Haryana region were found less effective as compared to Himachal Pradesh and Punjab region Table-2. Zone of inhibition was beyond detection limit at 1000µg in case of Acinetobactor baumanii, Proteus mirabilis and Pseudomonas aeruginosa. The results indicate significant impact of environmental conditions and allelopathy on the concentration of secondary metabolites in plants which might be attributed to potentiate antibacterial activity. Antibacterial and antifungal activity has also been tested in leaves of Achyranthus aspera collected from Gulbarga and reported that Achyranthes aspera has non-specific antimicrobial activity12 which is in support of present investigation. At 500µg concentration, the sensitivity was reflected only by Escherichia coli and Klebsiella pneumoniae from all the regions investigated table-2. Available literature indicates that the antibacterial activity is due to different chemical agents present in the extract including essential oils (especially thymol), flavonoids and triterpenoids and other natural phenolic compounds or free hydroxyl groups. These are classified as active antimicrobial compounds13. Flavonoids have a number of nutritional functions and have been described as biological response modifiers; most act as an anti-oxidant and some have anti-inflamatory properties. Flavonoids have been shown to prevent or slows the development of some cancers 14 Several investigators have reported that the methanolic extracts of leaves Achyranthes aspera has significant antimicrobial activity against the Gram-Psitive (S. aureus, Bacillus subtilis), Gram-negative bacterial (K. pneumoniae, E. coli) and fungal species (Aspergillus niger, C.albicans12. Conclusion Although methanolic extract of whole plant is having antibacterial potential yet at higher concentration of constituents. It has also been observed that antimicrobial activity is not specific and dependent on the location of procurement which shows that there is somehow correlation between allelopathic effect and environmental conditions as well on antimicrobial potential of Achyranthus aspera.Further study is needed to identify such factors and the strain of Achyranthes aspera having high antimicrobial potential and its conservation.AcknowledgementAuthors wish to acknowledge the Chancellor, Lovely Professional University, Phagwara Punjab, INDIA for providing financial aid. We also acknowledge Dr. Vandana Berry, Department of Microbiology, Christian Medical College, Ludhiana, Punjab, INDIA for her support. References1.Girach R.D. and Khan A.S.A., Ethnomedicinal uses of Achyranthes aspera leaves in Orissa (India), Int. 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