Bio Activity Study On Methanol And Aqueous Extracts Biology Essay

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The study proposed on the plant Terninalia arjuna belongs to the family combretaceae. T. arjuna traditionally known to be used to cure many health related issues such as heart ailments, diabetics and also has wound healing property. Earlier reports on this tree show that it can be used as Anti-HIV, Anti-Tumor, Hepatotoxicity drug etc. The tree is widely distributed throughout India and mostly seen in Western Ghats. In this study the T. arjuna bark and core wood were collected from Tirunelveli District., Tamil Nadu. After collection, the tree parts were subjected to successive extraction using methanol and water successively. Following, different concentrations of both extracts will be used to explore the Hemolytic, larvicidal, effect of pH, presence of toxic heavy metals and antimicrobial assay on selected microbial species. (Include the Results - in short)

Keywords: T. arjuna, Hemolytic Assay, Larvicidal Activity,Antimicrobial assay,Core Wood and Bark

Guide: Dr. Padma Thiagarajan

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Professor, SBST

VIT University, Vellore 632014

Tamil Nadu, India.

INTRODUCTION

In developing countries, mortality due to malarial infection among children is becoming a major issue. In India the etiology is due to un-hygiene practice which leads to high breeding of Anopheles, resulting in severe sickness [1, 2]. The antibiotic resistant pattern of these mosquito larvae have been changing frequently. There is a need for establishing a particular type of drug which is harmful to larvae without effecting human system. Hemolytic assay is also performed to check the effect of extract on human. As Red Blood Cells resemble the mechanism of other cells, the study was designed to check the effect of extracts for hemolysis.

Many plants in India are used in folk lore medicine for treatment of larval infections [3]. The potential benefits of herbal medicines would lie in their efficacy, safety, little or no side effects and relatively low cost. In India, there are several hundred plants belong to the family combretaceae such as T.arjuna have been reported to contain a wide range of secondary metabolites that have a cytotoxic effects [4]. These cytotoxic effects have been demonstrated in experiments performed. However not much work has been done on the antibacterial activity of roots, leaves and fruits of T.arjuna.in the present work we describe antibacterial activity of aqueous and methanol extracts of bark and corewood.

The medicinal value of plants lies in some chemical substances that produce a definite physiological action on the human body. Screening of medicinal plants for elementological activities are important for finding potential new compounds for therapeutic use. Screening of medicinal plants for elementological activities are important for finding potential new compounds for therapeutic use. (16) Total three elements (zinc,cadmium and lead)are analysed in these plant extracts.

MATERIALS AND METHODOLOGY

Raw materials: The bark and core wood of T. arjuna was powdered and extracted using soxhlet apparatus with methanol and water as solvents according to the standard protocols. [5]

Blood sample: Blood sample collected from the healthy donor

Larvae : Mosquito Larvae collected from water source

Chemicals: Phosphate buffer saline and 1% saline, concentrated sulphuric acid, Muller Hinton Agar, lead nitrate, cadmium chloride, zinc sulphate.

Instrumentation: centrifuge micro pipette, spectrophotometer, ph meter, atomic absorption spectrophotometer.

Clinical samples: Staphylococcu sp,E.coli, Pseudomonas, Candida sp, Bacillus sp.

2.1 Haemolytic assay:

Hemolytic assay was performed according to the method described by Bulmus et al. [6] 5ml of blood is collected from healthy donor and centrifuged at 2000 rpm for 15 minutes. To the volume of plasma aspirated equal volume of PBS is added. The entire plasma is washed using PBS. The extra PBS is removed and after the cells settle they are diluted by a factor of 10%. This mixture (RBC +PBS) is made up to a volume of 20ml. Different concentrations (50,250,500) of the aqueous and methanolic extract is prepared using PBS. To 1.5ml of each of these concentrations containing RBC and PBS, 1.5ml of sample is added. This is incubated in a water bath at 370 C for 60 minutes. The mixture is then centrifuged and supernatant is used to check for the absorbance at 541 nm with methanol as blank and triton x as a negative control.

2.2 Larvicidal assay

The assay was carried out according to the method described by WHO, 2005 [7]. Ten larvae are dispersed into separate beakers marked for various concentrations. 500μg and 1000μg of sample is prepared using methanol and 1 ml of each concentration is added to the corresponding beaker containing larvae and left for incubation at room temperature for 24 to 48 hours. 1 % methanol was used as control.

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2.3. Estimation of Ph:

Different concentration of methanol bark and core wood extract was prepared and pH was checked using a pH meter.

2.4. Atomic absorption spectroscopy:

Standards of lead, zinc and cadmium were prepare at different concentration. The extract was dissolved in concentrated sulphuric acid and filtered. The filterate was subjected to AAS along with standards.

2.5 Antimicrobial activity:

Bacterial and fungal species were screened for different concentrations of bark and corewood methanol and aqueous extracts.

3. RESULTS AND DISCUSSION

Plants are known to have beneficial therapeutic effects documented in Traditional Indian System of Medicine [8, 9]. Though bioactivity of T.arjuna has been used in treatment of various aliments since time immemorial, their larvicidal and hemolytic activities were yet to be explored [10]. The present study revealed that at 500mg concentration of methonolic extract was able to show significant larvicidal property. This was more apparent when the concentrations of extract were increased and the same is tabulated (Table 1). The results obtained showed a time and concentration/dose dependent in larvicidal activity which reveals aqueous extract of both bark and core wood were found to be 100% lethal at 1000 mg concentration. A similar observation was reported in the earlier study carried out on the same family [11, 12].

For hemolytic activity, even at the concentration of 1000mg both the plant extract methanol and aqueous were not able to lyse the RBCs. This result is also in accordance with the previous reports. [13, 14].

(WHERE are th Discussion for the below results)???????????????? Send me back with discussion. Call me as well!...

In case of atomic absorption spectroscopy the plant showed absence of cadmium.lead was present only in corewood aqueous extract. Zinc was present in mild concentration. Preferably corewood extract showed maximum amount of zinc.

The results of pH estimation showed no drastic changes in methanol and aqueous extracts of corewood and bark. (Tabulate the pH values)

The antimicrobial activity of both,extract at various concentrated did not show any antimicrobial property. and results are tabulated(Table 2).

T. arjuna

Extract

Conc (μg)

Death % at various Time durations

12 hrs

24hrs

48hrs

Bark

Methonol

500

10

30

70

1000

20

40

90

Aqueous

500

10

30

90

1000

10

100

-

Core-wood

Methonol

500

20

50

90

1000

40

60

80

Aqueous

500

30

60

80

1000

40

60

100

Table 1: Larvicidal Activity of T. arjuna Bark and Core-wood

Microbial strains

Bark wood

Core wood

Bacillus species

nil

nil

Pseudomonas

nil

nil

Stap.aureus

nil

nil

Candida1

nil

nil

Candida 2

nil

nil

Table 2:Antimicrobial Activity of T.arjuna bark and core-wood

CONCLUSION

The study concludes, methonolic extracts of T. arjuna at minimum concentration of 500mg, was lethal for larvae and in-case of hemolytic property both the extracts were non-hemolytic making them therapeutically safe for humans. These assays have been carried out for other plants but not touched upon the core wood and hence these were reperformed using corewood.