«In Vitro Antioxidant and Antimicrobial Activities of Aerva lanata L. Keywords: Antioxidant, Antimicrobial, Aerva lanata, phytochemicals ABSTRACT ...»
May 2016 Vol.:6, Issue:2
© All rights are reserved by Prasanna. G et al.
In Vitro Antioxidant and Antimicrobial Activities of Aerva lanata L.
Keywords: Antioxidant, Antimicrobial, Aerva lanata,
Manohari R and Prasanna. G The present investigation has been carried out to evaluate the phytochemical, in vitro antioxidant and antimicrobial activities P.G and Research Department of Biochemistry, of aqueous and ethanolic leaves extracts of Aerva lanata L.
Sengamala Thayaar Educational Trust Phytochemicals were analyzed by qualitative method and the Women’s College, Mannargudi, Thiruvarur Dt, Tamil results revealed the presence of alkaloids, phytosterols, terpenoids, flavonoids, tannins, saponins and steroids etc., The Nadu, India.
in vitro antioxidant activity was evaluated using total Submission: 5 May 2016 antioxidant capacity, nitric oxide scavenging activity, reducing Accepted: 10 May 2016 power assay and hydrogen peroxide scavenging activity.
Published: 25 May 2016 Ethanolic extract showed the highest antioxidant activity than aqueous when compared with standard vitamin C.
Antimicrobial activity was also performed by using agar disc diffusion method against E.coli, B. subtilis, A.niger, T. viride.
Ethanol and aqueous extracts were active against both bacterial and fungal species when compared to antibiotic ciprofloxacin.
www.ijppr.humanjournals.com As a result, ethnologic leaves extract of Aerva lanata L. showed high efficiency of antioxidant and antimicrobial activity due to the presence of various phytochemicals.
INTRODUCTIONOxygen is essential for the survival of all on this earth. Free radical is a chemical compound which contains an unpaired electron spinning on the peripheral layer around the nucleus. Cell damage caused by free radicals appears to be a major contributor to aging and degenerative diseases such as cancer, cardiovascular disease, cataracts, immune system decline, liver disease, diabetes mellitus, inflammation, renal failure, brain dysfunction and stress among others1. Free radicals include Superoxide radical (SOR), Hydroxyl radical (OH), Hydroperoxyl radical (HPR), Alkoxyl radical (AR), Peroxyl radical (PR), Nitric oxide radical (NOR). Non-free radical includes Singlet O2, Hydrogen peroxide (H2O2), Hydro chlorous acid (HOCL), Peroxynitrite ONOO-. In addition, there is another class of free radicals that are nitrogen derived called reactive nitrogen species (RNS) 2. All these capable of reacting with membrane lipids, nucleic acids, proteins and enzymes and other small molecules, resulting in cellular damage. In living organisms, various ROSs can be formed in different ways including normal aerobic respiration stimulated polymorphonuclear leukocytes, macrophages and peroxisomes. Antioxidant may be defined as radical scavenger which protects the human body against free radicals that may cause pathological condition such as ischemia, anemia, asthma, arthritis, inflammation and Parkinson’s disease. Antioxidant means “against oxidation” Antioxidant work to protect lipids from peroxidation by radicals. The human body has an elaborate antioxidant defense system.
Microorganisms are in large part responsible for determining the course and human history. They are carried by air current from the earth’s surface to the upper atmosphere. The condition that favours the survival for the growth of many microorganisms in those under which people normally like it is inevitable that we among a multitude of microbes3, 4. Infectious diseases also known as contagious diseases or transmissible diseases and include communicable diseases comprise clinically evident illness (i.e., characteristic medical signs and/or symptoms of disease) resulting from the infection, presence and growth of pathogenic biological agents in an individual host organism. In certain cases, infectious diseases may be asymptomatic for many or their entire course. Infectious pathogens include some viruses, bacteria, fungi, protozoa, multicellular parasites and aberrant proteins known as prions. Today, there are a large number of antimicrobial agents used in medicinal practice are aimed at eliminating infecting microorganism are at preventing the establishment of an infection. In India, herbal medicines have been the basis
of treatment and care for various diseases. Physiological conditions in traditional methods practiced such as Ayurveda, Unani and Siddha5. Keeping this view in our study, we evaluated the phytochemicals in vitro antioxidant and antimicrobial activity of Aerva lanata leaves.
MATERIALS AND METHODSCollection of plant material The plant was collected from in and around Athamangalam, Nagai District, Tamil Nadu. The collected samples were carefully kept in polythene bags. These plant samples were authenticated by Dr.S.Johnbritto, The Director, The Rabinet Herbarium Centre for Molecular Systematic, St.
Joseph’s College, Tiruchirappalli and a voucher specimen was deposited in the Department of Biochemistry, S.T.E.T Women’s College, Mannargudi, Thiruvarur, Tamil Nadu. The leaves were dried in the shade and stored in airtight containers until further studies.
Extraction of plant material
Aqueous and ethanol extracts were prepared according to the methodology of Indian pharmacopoeia13. The shady dried plant materials were subjected to pulverization to get coarse powder. The coarse powder material was subjected to soxhlet extraction separately and successively with ethanol. The extract was concentrated to dryness in flash evaporator under reduced pressure and controlled temperature (40-50°C). The aqueous and ethanolic extract put in airtight container and stored in refrigerator.
Phytochemicals analysis Aqueous and ethanolic extracts of Aerva lanata Linn., were subjected to qualitative test for the identification of various plant constituents14.
In vitro antioxidant activity The antioxidant activity of plant extracts was determined by different in vitro models such as total antioxidant15, reducing power assay16, H2O2 scavenging activity17 and nitric oxide scavenging activity18.
In vitro antimicrobial activity Selection of microorganism Totally four pathogenic microorganisms such as E. coli, Bacillus subtilis, Aspergillus niger and Trichoderma viride were selected for this investigation.
Inoculum preparation The young microbial inoculum was prepared and used in the entire research period. The nutrient broth for bacteria and potato dextrose agar (PDA) for fungi were prepared and poured into tubes and sterilized. The pure microbial cultures were collected from Microbiology Department, S.T.E.T Women’s College, Mannargudi and inoculated in the tube using inoculation needles or loops. The tubes were incubated at 37°C for 24 hours.
Disc diffusion method19
The antimicrobial activity of the leaves extracts were tested against selected bacterial and fungal strains. The 20ml of sterilized nutrient agar medium were poured into each sterile Petri plate and allowed to solidify. Test microbial cultures were evenly spread over the appropriate media by using sterile cotton swab. Then a well of 0.5cm was made in the medium using a sterile cork borer 150µl of each ethanol and aqueous. Plant extracts were transferred into separate well after these plants were incubated at 37oC for 24-48 hours. After incubation period, the results were
observed and measured the diameter of inhibition zone around each well. The standard antibiotic (positive) like ciprofloxacin was also performed.
RESULTS In the present study, the antioxidant and antimicrobial activity of aqueous and ethanolic extract of Aerva lanata were examined. Table 1 represents the qualitative analysis of phytochemical constituents of Aerva lanata L. and the results revealed that the presence of alkaloids, carbohydrate, phytosterols, terpenoids, flavonoids, protein, amino acids, volatile oils, tannins, saponins and steroids were present in aqueous and ethanol extracts. Phlobatannins and phenolic compounds were absent in both aqueous and ethanolic plant extracts.
The in vitro antioxidant activity of aqueous and ethanolic extracts of Aerva lanata L. were assayed by different in vitro models, including total antioxidant capacity, reducing power assay, hydrogen peroxide scavenging activity and nitric acid scavenging activity.
Table 2 showed the total antioxidant capacity of ethanolic and aqueous extract of Aerva lanata L.
The total antioxidant capacity of ethanolic extract was 84.4% and aqueous extract showed 25.5%. Among the two extracts, ethanolic extracts showed the highest antioxidant activity which was nearer to that of standard antioxidant vitamin C (92%).
Nitric oxide scavenging activity of ethanolic and aqueous extract of Aerva lanata L. represented in Table 2. Ethanolic extract showed 66.7% of scavenging activity and aqueous extract having 33.5% and vitamin C showed 70.33%. Ethanolic extract exhibited moderate scavenging activity compared to vitamin C.
Reducing power assay of aqueous and ethanolic extract of Aerva lanata L were also represented in Table 2. Reducing power of ethanolic extract was found to be 70% and in aqueous extract, it was 44.2%. Reference compound vitamin C showed 72.23%. Ethanolic extract exhibited highest scavenging activity than aqueous extract. When compared to vitamin C ethanol extract showed moderate activity.
Table 2 also showed hydrogen peroxide scavenging activity of ethanol and aqueous extract of Aerva lanata L. Ethanolic extract showed 60% of hydrogen peroxide scavenging capability,
which is nearer to the value showed by reference compound vitamin C (65.3%). But aqueous extract having least potentiality (25.1%) when compared to ethanolic extract.
Antimicrobial activity In the present study, we also analysed the antimicrobial activity of Aerva lanata leaves extracts.
Bacteria like E.coli, B.subtilis, and fungi like A.niger, and T.viride were selected for antibacterial and antifungal activities of plant extracts respectively. Zone of inhibition of both extracts were represented in Table 3.
When tested by disc diffusion method, ethanolic leaf extracts of Aerva lanata L. showed significant inhibitory activity against E.coli, (6mm) and B.subtilis (5mm) than aqueous leaf extract. In aqueous extract, zone of inhibition for E.coli and B.subtilis were 4mm and 3mm respectively. Antibiotic ciprofloxacin was also performed and their zone of inhibition for E.coli and B.subtilis were 2mm and 2mm respectively. Both ethanolic and aqueous extracts exhibit higher antibacterial activity than the standard drug.
Ethanolic extract of Aerva lanata L. showed significant antifungal activity against A.niger (8mm) and T.viride (4mm) and aqueous extract exhibited 6mm for A.niger and 5mm for T.viride.
The ethanolic extract exhibited highest antifungal activity against A.niger whereas the aqueous extract showed maximum activity against T.viride. All the results were compared with standard antibiotic ciprofloxacin (3mm for A.niger and 3mm for T.viride) and both extracts showed better results than ciprofloxacin.
DISCUSSION Free radical is a molecule with an unpaired electron and is involved in bacterial and parasitic infections, lung damage, inflammation, reperfusion injury, cardiovascular disorders, atherosclerosis, aging and neoplastic diseases20. Flavonoids, which are well known antioxidants and free radical scavengers such as kaempferol 3 - rhamnoside and kaempferol 3 rhamnogalactoside have been reported to be present in Aerva lanata L.21. The significant hepatoprotective, nephroprotective and antioxidant effect of Aerva lanata L. may be due to presence of alkaloids and its inhibitory effect on microsomal Cyt P450 enzyme or on lipid peroxidation11. Reactive oxygen species are generated in the human body cause oxidative damage and responsible for many degenerative diseases such as coronary heart disease, atherosclerosis, diabetes, aging and cancer22. Many antioxidant compounds occurring naturally in plants sources have been identified and proved as free radical scavenger both in vitro as well as in vivo. Aerva lanata L. has been used as a traditional medicine for various ailments which are
closely associated with free radical formation. Hence in the present investigation, we have evaluated the free radical scavenging activity of Aerva Lanata L.
Total antioxidant capacity of the ethanolic extracts of Aerva lanata L. was more when compared to aqueous extract. The total antioxidant capacity of the extract was calculated based on the formation of phosphorus molybdenum complex which was measured at spectrometrically at 695nm15. Nitric oxide (NO) is an important chemical mediator generated by endothelial cells, macrophages, neurons, etc. and involved in regulation of various physiological processes23.
Excess concentration of NO is associated with several diseases24. Oxygen reacts with the excess nitric oxide to generate nitrite and peroxynitrite anions, which act as free radicals 25, 26. In the present study, the extract competes with oxygen to react with nitric oxide and thus inhibits the generation of the anions.
For measurements of ethanolic and aqueous extract of Aerva lanata L. the reducing ability, the Fe3+ to Fe 2+ transformation was investigated in the presence of Aerva lanata L. The reducing capacity of a compound may serve as a significant indicator of its potential antioxidant activity.