|Year : 2013 | Volume
| Issue : 1 | Page : 71-75
Absence of antibiotic activities of Cenchrus setigerus and cenchrus ciliaris seed extracts in different polar solvents
Premlata Singariya1, Padma Kumar1, Krishan K Mourya2
1 Department of Botany, Laboratory of Tissue Culture and Secondary Metabolites, University of Rajasthan, Jaipur, Rajasthan, India
2 Department of Animal Husbandry, Government of Rajasthan Veterinary Hospital, Pahari (Bharatpur), Rajasthan, India
|Date of Web Publication||21-Aug-2013|
Dr. D. S. Kothari Post Doctoral Fellow, Shri Kalyan Nagar, Kartarpura, Jaipur, Rajasthan
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Aim: The aim of present study is to investigate the antibiotic activity of seeds of Cenchrus setigerus and Cenchrus ciliaris extracts, in order to use them as a possible source of new anti-microbial substances against important human pathogens. Materials and Methods: Crude extracts of seeds of both species of Cenchrus grass were evaluated against some important G-ve bacteria. Escherichia coli, Raoultella planticola, Enterobacter aerogenes and one fungus Aspergillus flavus. The dried and powdered seeds were successively extracted with hexane, toluene, isopropyl alcohol, acetone and ethanol using the soxhlet assembly. The antimicrobial activity assay was done by both disc diffusion and serial dilution methods. Results: The results indicate that all the extracts, in different polar solvents did not show any antibacterial activity against R. planticola or any antifungal activity against A. flavus. Conclusion: All extracts in the different polar solvents did not have or had very less antibacterial and antifungal activities.
Keywords: Antibiotic, Aspergillus flavus, Cenchrus, hexane, toluene
|How to cite this article:|
Singariya P, Kumar P, Mourya KK. Absence of antibiotic activities of Cenchrus setigerus and cenchrus ciliaris seed extracts in different polar solvents. J Pharm Negative Results 2013;4:71-5
|How to cite this URL:|
Singariya P, Kumar P, Mourya KK. Absence of antibiotic activities of Cenchrus setigerus and cenchrus ciliaris seed extracts in different polar solvents. J Pharm Negative Results [serial online] 2013 [cited 2020 Jun 6];4:71-5. Available from: http://www.pnrjournal.com/text.asp?2013/4/1/71/116756
| Introduction|| |
Thar desert occupies diverse habitat and landscape that support distinctive plant species. About 682 plant species have been identified including 107 grasses.  Cenchrus setigerus and Cenchrus ciliaris (C 4 grasses) are gaining attention in various field of research, as they are best suited to the present environmental conditions. These grasses are more competitive under the conditions of high temperature, solar radiation and low moisture  and are more efficient at gathering CO 2 and utilizing nitrogen from the atmosphere and recycled N in the soil. , Cenchrus L. (Poaceae) is highly nutritious grass and considered excellent for pasture in hot, dry areas and is valued for its production of palatable forage and intermittent grazing during droughty periods in the tropics. The grass, fed green, turned into silage, or made into hay is said to increase flow of milk in cattle and impart a sleek and glossy appearance. This grass has excellent soil binding capacity which helps to conserve soil in desert areas.  However, Cenchrus is most suitable and highly nutritive grasses for desert environmental conditions, still no antimicrobial work yet have been done on this grass.
E. coli is the culprits for human urinary tract infections.  Raoultella planticola has been determined to cause severe pancreatitis in one case.  E. aerogenes is a nosocomial and pathogenic bacterium that causes opportunistic infections including most types of infections. Aspergillus species are the most common mold causing severe infections. ,
| Materials and Methods|| |
Crude extracts of seeds of Cenchrus ciliaris (CAZRI-358) and Cenchrus setigerus (CAZRI-76) were prepared with a series of non polar to polar solvents by hot extraction method  in soxhlet assembly. Different extracts were then screened for antimicrobial activity by DDA  against a few medically important bacteria and fungi. The fraction showing best activity was then used for determining of Minimum inhibitory concentration (MIC) by tube dilution method  and minimum bactericidal/fungicidal concentration (MBC/MFC).
Collection of plant material
Seeds of C. ciliaris and C. setigerus were collected in the month of August from the CAZRI, Jodhpur, Rajasthan. Selected plant seeds were separately shade dried and powdered with the help of grinder.  Fine powder of each sample was stored in clean container to be used for Soxhlet extraction following the method of Subramanian and Nagarjan  in different polar solvents selected.
Seeds were sequentially extracted with different solvents according to their increasing polarity (hexane <toluene <isopropyl alcohol <acetone <ethanol) by using the Soxhlet apparatus for 18 hours at a temperature not exceeding the boiling point of the respective solvent. The obtained extracts were filtered by using Whatman No. 1 filter paper and then concentrated at 40°C by using an evaporator and stored in amber color bottle for subsequent use in the further antimicrobial and phytochemical analysis. 
Drugs and chemicals used
Gentamycin (for bacteria) and ketoconazole (for fungi)
Hexane, toluene, isopropyl alcohol, acetone and ethanol, Nutrient Agar (NA medium for bacteria), Sabouraud Dextrose Agar (SDA medium for fungi).
The organisms used in this study were namely [Table 1]. Test pathogenic microorganisms were procured from Microbial Type Culture Collection, IMTECH, Chandigarh, India. The reference strains of bacteria were maintained on nutrient agar slants, sub cultured regularly (every 30 days) and stored at 4°C as well as at -80°C by preparing suspensions in 10% glycerol.
Screening for antimicrobial activity
Bacterial colonies were maintained on NA medium and fungi were maintained on SDA medium. DDA  was performed for screening. NA and SDA base plates were seeded with the bacterial and fungal inoculum, respectively (inoculum size 1 × 10 8 CFU/ml for bacteria and 1 × 10 7 cell/ml for fungi). DDA was performed for screening by standard method.  Bacterial growths were seen after a minimum of 18 h and occasionally until 24 h.  Activity index for each extract was calculated.
Serial dilution method
MICs are considered as the "gold standard" for determining the susceptibility of the organisms to antimicrobials.  MIC of antibiotics was evaluated (thrice) using standard micro broth dilution method against inoculum Escherichia More Details coli organisms.  Bacterial and fungal suspensions were used as negative control, while broth containing standard drug was used as positive control.
Determination of minimum bactericidal/fungicidal concentration (MBC/MFC)
Equal volume of the various concentration of each extract and nutrient broth mixed in micro-tubes to make up 0.5 ml of solution. 0.5ml of McFarland standard of the organism suspension was added to each tube. , The tubes were incubated aerobically at 37°C for 24 h for bacteria and 28°C for 48 h for fungi. Two control tubes were maintained for each test batch. These include tube-containing extract without inoculum and the tube containing the growth medium and inoculum. The MBC was determined by sub culturing the test dilution on MHA and further incubated for 24 h. The highest dilution that yielded no single bacterial colony was taken as the MBC.  MBC was calculated for some of the extracts showed high antimicrobial activity against highly sensitive organisms.
Total activity (TA) determination
Total activity is the volume at which the test extract can be diluted with the ability to kill microorganisms. It is calculated by dividing the amount of extract from 1 g plant material by the MIC of the same extract or compound isolated and is expressed in ml/g. 
| Results|| |
Antimicrobial activity [assessed in terms of zone of inhibition (ZOI) in mm* and activity index] of the seeds of C. ciliaris and C. setigerus extracts in different polar solvents, tested against selected microorganisms were recorded [Table 2]. In the present study total ten extracts of selected plant were tested for their bioactivity, among which all these extracts showed insignificant antimicrobial potential against test microbes. However, all these extracts showed no activity or very less activity against E. coli, R. planticola, E. aerogens and antifungal activity against A. flavus at tested concentration. Highest antimicrobial activities were recorded for C. ciliaris in hexane (ZOI-9.17 ± 0.24 and AI-0.459) and for C. setigerus in acetone (ZOI-8.50 ± 0.64 and AI-0.425) against E. aerogens and E. coli, respectively.
|Table 2: Inhibition zone (mm)* and activity index of seeds of Cenchrus grass in different polar solvents against tested pathogens|
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MIC and MBC/MFC
MIC and MBC/MFC values [Table 3] were evaluated for those plant extracts, which were showing activity in disc diffusion assay. The range of MIC and MBC/MFC of extracts recorded was 3.75-15 mg/ml. In the present investigation lowest MIC value 3.75 mg/ml was recorded for C. ciliaris (against E. aerogens) in hexane extract.
|Table 3: Minimum inhibitory concentration and (MBC/MFC) of seeds of Cenchrus in different solvents against tested pathogens|
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Total activity indicates the volume at which extract can be diluted with still having ability to kill microorganism. Seed extracts of C. ciliaris in hexane showed high values of TA 7.73 ml against E. aerogenes [Table 4].
The preliminary phyto-profiling for the seeds of Cenchrus extracts were carried out according to Farnsworth  wherein the consistency was found to be sticky in the hexane and toluene extracts whereas all other extracts were found to be non-sticky. The yield mg/g (w/w) of the extracts was also analyzed wherein the highest yield was found in acetone extract of C. ciliaris (685 ± 11.78 mg/10 g ± S.D.) followed by ethanolic and iso propyl alcohol extract of C. setigerus (346 ± 13.46 mg/10 g ± S.D.) and (346 ± 10.13 mg/10 g ± S.D.) respectively [Table 5].
|Table 5: Preliminary phyto-profile for seeds of Cenchrus grass in different polar solvent|
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| Conclusion|| |
Cenchrus ciliaris and C. setigerus extracts in different polar solvents at a tested concentration were not showing ZOI against the selected pathogens [Table 1] although, water, chloroform and acetic acid extract of seeds of this grass show good activity against these pathogens supported by different researchers. ,, But, in this study these extracts lack of antimicrobial activity well be useful to avoid any study repeated in this direction in the future.
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[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]