Harish Gupta^1,2, Kishlay Kumar² and R. C. Agrawal^1*

¹Department of Research, Jawaharlal Nehru Cancer Hospital and Research Centre, Idgah Hills, Bhopal India.

²RKDF College of Pharmacy, Bhopal India.

Abstract

The in vitro antibacterial and antioxidant properties of 50% methanolic extract of fruit from Cassia fistula L. (Indian Laburnum) was investigated. In this study the antibacterial activity of Cassia fistula against different gram positive and gram negative bacteria by disc diffusion method and Antioxidant activity by Fenton reaction was undertaken. Cassia fistula has shown effective against Gram positive bacteria and Gram negative bacteria and the MIC value was measured. In another experiment antioxidant activity was observed and IC50 was shown at 1200 µg. concentration. These results are important because this plant is widely used for medicinal and ornamental purposes.

Keywords

Cassia fistula; antibacterial; antioxidant

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Copy the following to cite this article: Gupta H, Kumar K, Agrawal R. C. Evaluation of Antibacterial and Antioxidant Activity from Fruit Extract of Cassia Fistula L. Biomed Pharmacol J 2010;3(1)

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Introduction

Cassia fistula Linn  is known as Amaltas, in Hindi which is  a  medium  size tree belongs to family Caesalpiniodeae and is widely cultivated throughout India, It is widely used for its medicinal properties. The main property which is reported are  mild laxative¹, cure of  skin disease², wound healing³, Hypoglycemic⁴, antibacterial⁵, antifungal⁶, hypocholestremic⁷ hepatoprotective ^8,9, antitumour ¹⁰, antioxidant^11.12, antifertility ¹³. This plant is widely used in traditional medicine, in the present we are reporting  antibacterial and antioxidant potential  of 50% methanolic extract of the fruit from Cassia fistula.

Material and Method

Plant Material

The fresh fruit of Cassia fistula linn was collected from shops of Bhopal, India and identified by N.K.Pandey of  Central Research Institute , Gwalior, India. A voucher specimen (field book No. 10630) has kept in our laboratory for future reference. The fruit were dried under shed and  dried powdered was  kept in a well closed container for further experiment.

Preparation of extract

Powder of Cassia fistula was treated with petroleum ether for defatiation for 30 minutes then extracted with 50% methanol until the colourless solution was obtained using separating funnel. The upper layer was collected and then extract was kept at water bath for  drying thus the powder obtained which was used for the further experiment.

Test organism

The following gram negative and gram positive bacteria i.e.  Pseudomonas aeroginosae, Staphylococcus aureus, Staphylococcus epidermidis, Shigella flexineri, Bacillus subtilis & Escherichia coli were used for antibacterial activity which were received from stock culture of our laboratory

Methodology

Antibacterial activity

Antibacterial activity of 50% methanolic, extract from fruit of Cassia fistula were investigated using  disk diffusion method ¹⁴. 20 ml of sterile Nutrient agar was added in  the test tube after that Petri plate were prepared and  cultures were swabbed on the top of the solidified media and allow to dry . The test were conducted at four different concentration of the crude extract 25%, 50%, 75% & 100% (100 μl per disk ), the disk which was soaked with extract were placed on the surface of the medium the plate and were incubated for 12-18 h at 37⁰c and the zone  > 8mm were considered not active against microorganism as  reported in the literature¹⁵.

Antioxidant Protocol

The hydroxyl radical attacked  deoxyribose and initiated a series of reaction that eventually resulted  in the formation of Thiobarbituric acid reactive substance (TBARS). The measurement of TBARS thus given an index of free radical scavenging activity. The reaction mixture consisted  of a deoxyribose (3mM, 100μl) Ferric chloride (Fe³⁺ 0.2 mM  50μl) EDTA (0.1mM, 50 μl) ascorbic acid (0.1 mM 100μl) and H₂O₂  (1mM, 100μl) stock solution of Cassia fistula extract  2mg/ml were prepared from which 200-2000 μl were added in Reaction mixture . The final volume was made up to 1 ml by adding adequate quantity of phosphate buffer saline (p^H 7.4) and incubated for 1h at 37⁰c. The Reaction was stopped by adding 0.5ml of  5% TCA  and 0.5ml of 1% TBA the mixture was than incubated  for 20 min in a boiling water bath. The absorbance was measured at 532 nm. Ascorbic acid was used as the positive control.  The results are expressed as the percentage inhibition of TBARS as reported.¹⁶_.

Result and Discussion

Antibacterial activity

50% methanolic extract of Cassia fistula at the different concentration i.e.  25%, 50%, 75%, 100% exhibited antibacterial against Bacillus subtilis (9-13 mm), Staphylococcus epidermidis (10-12mm), Staphylococcus aureus (9-12mm) but the lower activity was observed against Shigella flexineri (9 mm at 100%) E. coli (9 mm at 100%) and Pseudomonas aeroginosae (9 mm at 100%) the Minimum inhibitory concentration (MIC) of Cassia fistula against gram positive bacteria i.e. Bacillus subtilis, Staphylococcus epidermidis, Staphylococcus aureus was 25% but against gram negative bacteria Zone of inhibition was observed only in 100%  extract . Other concentration i.e. 25%,50 % and 75 % has not shown any zone of inhibition (table No.-1.)  Result showed Cassia fistula was effective against gram positive bacteria but it was very less effective against gram negative bacteria. Cassia fistula was also reported significant antibacterial activity against the some bacteria.⁵ Our findings confirm the traditional therapeutic claims of  this herb.

Table 1: Antibacterial activity of Cassia fistula against different bacterial strains

Name of Bacteria                                                        Zone of Inhibition (mm)

Concentration of drug

25%                 50%                 75%                 100%

(+ve)

Bacillus subtilis                                   10                    9                      11                    13

Staphylococcus epidermidis               10                    10                    11                    12

Staphylococcs aureus                          9                      10                    12                    11

(-ve)

Shigella flexineri                                 –                       –                       –                       9

Escherichia Coli                                  –                       –                       –                       9

Pseudomonas aeroginosae                  –                       –

 

Antioxidant activity

The extract of Casia fistula showed   good antioxidant property in Fenton reaction model. The test drug was compared with a low concentration of ascorbic acid. Table-2 showed the % TBARS inhibition of methanolic extract of Cassia fistula and ascorbic acid. Crude 50% methanolic extract of Cassia fistula showed IC₅₀ at 1200 μg and the ascorbic acid showed  IC₅₀ at 200 μg. Aqueous extract of Cassia fistula (Linn.) flowers (ACF) was reported  for its antioxidant effect in alloxan induced diabetic rats ¹² but there are no report about the  antioxidant activity of fruit extract of Cassia fistula using Fenton reaction.

Graph 1:  % inhibition of TBARS by different concentration of Cassia fistulaextract  Click here to View graph

 

Table 2: In vitro antioxidant activity of 50% methanolic fruit extract from Cassia fistula extract

Sr. No.	Concentration of ascorbic acid (μg)	% TBARS inhibition  SEM	Concentration of cassia fistula (μg)	% TBARS inhibition  SEM
1	50	43.420.63	200	28.800.21
2	100	44.172.11	400	30.290.42
3	150	40.880.00	600	35..40.42
4	200	500.84	800	44.160.21
5	25	54.011.26	1000	48.171.48
6	300	58.031.26	1200	5000*
7	350	75.701.83	1400	55.470.84
8	400	54.750.64	1600	32.972.00
9	450	62.782.11	1800	32.110.21
10	500	80.773.16	2000	36.132.11

* denoted IC₅₀ value of cassia extract

 

Conclusion

From the above result it may  be concluded that 50% methanolic fruit extract of Cassia fistula has antibacterial activity against gram positive bacteria but less activity in  gram negative bacteria. The antioxidant activity of this extract was also observed. These results are important because this plant is widely used for medicinal and ornamental purposes.

Acknowledgment

The authors are thankful to Shri Madan Mohan Joshi, Chairmain and K.V.Pandya, Director of the Jawaharlal Nehru Cancer Hospital & Research Center, Bhopal for providing facilities to carry out the present work and also thankful to Ms. Nirmala Gupta ,Ms. Tripti Beohar, Ms. Silky Vishnoi and Ms. Deepika Mathur for help during standardization of the above techniques. Harish Gupta  would like to express his thank to Dr. Neeraj Upmanyu for giving him  permission to do work in JNCH&RC, Bhopal

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