Phytochemical Investigation with Assessment of Cytotoxicity and Antibacterial Activities of the Ethanol Extract of Elaeocarpus serratus

INTRODUCTION

Different types of plant and plant-derived compounds are used in folk medicine for the treatment of different ailments (Pareta et al., 2011). The medicinal plant, Elaeocarpus serratus Linn belonging to the family of Elaeocarpaceae is widely distributed in Chittagong region of Bangladesh. The plant also grows in many other parts of Bangladesh, India, Pakistan, Srilanka, East Africa, Tropical Australia, Subtropical and Tropical Asia (Ghani, 2003). The plant is commonly known as Singhali Jolpai in Bangladesh (Ghani, 1998). A number of glycosides such as myricitrin, mearnsetin 3-O-β-D-glucopyranoside, mearnsitrin, tamarixetin 3-O-α-L-rhamnopyranoside were isolated from the leaves of Elaeocarpus serratus. The plant extract possessed antioxidant properties due to the presence of myricitrin (Jayasinghe et al., 2011). Sriti et al. (2011) reported the presence of alkaloids and anthraquinone glycosides in the chloroform extract of the leaves of the plant. They also detected flavonoids in petroleum ether extract of the same plant. Rheumatism and poisoning are treated by the extract of the leaves and on the other hand, fruits are used to treat diarrhea, dysentery and to increase appetite in patients by the stimulation of taste buds (Sharker and Shahid, 2010).

From literature survey, it was found that few works had been done on the cytotoxicity and antibacterial activities of the crude extracts of leaves and fruits of the plant. This stimulated interest to further investigate this plant with a view to determine the antibacterial and cytotoxic properties of the ethanol extract of Elaeocarpus serratus. The study was also designed to detect the phytochemical composition of the crude extract.

MATERIALS AND METHODS

Collection and preparation of plant extract: The leaves of Elaeocarpus serratus were collected from Sitakunda of Bangladesh in the month of October, 2010 at day time. After collection, they were washed by running tap water and were kept under shadow and air-dried for 21 days. Then, the dried leaves were cut into small pieces and ground into a course powder and stored in an air-tight container until the analysis started. About 338 g of the powdered materials were soaked in 1400 mL of 70% ethanol in a glass container and kept for a period of 21 days with continuous shaking. The whole mixture was then filtered by a piece of clean and white cotton materials. Finally, the filtrate was evaporated until dried to render to a reddish black color of the crude ethanol extract of the plant and the yield value of the ethanol extract of leaves of Elaeocarpus serratus was 3.57%.

Chemicals and standard drug: Ethanol was collected from LOBA Chemicals Pvt. Ltd., India and levofloxacin was obtained from Incepta Pharmaceuticals Ltd., Bangladesh. All the chemicals used in the whole study were of analytical grade.

Preliminary phytochemical screening: Preliminary phytochemical analysis of the ethanol extract was carried out to identify the presence of several types of phytochemical compounds such as alkaloid, cardiac glycosides, steroid, flavonoid, tannin, reducing sugar, saponin and gum according to the standard procedures (Trease and Evans, 1989).

Test microorganisms: The test microorganisms used in this study were Bacillus subtilis, Bacillus megaterium, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Shigella dysenteriae and Vibrio cholerae.

Cytotoxic activity: The cytotoxic activity of the ethanol extract was tested on brine shrimp nauplii according to brine shrimp lethality bioassay (Meyer et al., 1982). From this study, LC₅₀ (μg mL^-1) and LC₉₀ (μg mL^-1) of the ethanol extract were determined.

In vitro antimicrobial assay: The antimicrobial activity of the ethanol extract was investigated using agar disc diffusion method (Bauer et al., 1966) where the concentrations of the ethanol extract and the standard drug, levofloxacin were 500 and 10 μg disc^-1, respectively.

Statistical analysis: Three replicates of each sample were used for statistical analysis and the results of the experiment were expressed as Means±Standard Deviation (SD).

RESULTS

Preliminary phytochemical analysis: The phytochemical screening studies showed the presence of alkaloid, glycoside, tannin, saponin, flavonoid and carbohydrate. The results of phytochemical analysis are shown in Table 1.

Cytotoxic activity: The brine shrimp lethality bioassay showed the cytotoxic properties of the ethanol extract of the plant. The LC₅₀ (μg mL^-1) and LC₉₀ (μg mL^-1) were deduced from the best-fit line slope where the values of LC₅₀ (μg mL^-1) and LC₉₀ (μg mL^-1) were 141.25 and 870.96 μg mL^-1, respectively which are shown in Table 2.

Antibacterial activity: In agar disc diffusion method, three gm (+) and five gm (-) bacterial strains were used to observe the antibacterial actions of the ethanol extract of the plant at the concentration of 500 μg disc^-1. Table 3 showed the results of antibacterial activity of the extract with significant zone of inhibition. It was observed that the extract was effective against B. subtilis, B. megaterium, P. aeruginosa, S. typhi and V. cholerae with zone of inhibition between 16.93±0.14 to 32.95±0.39 mm.

Table 1:	Results of preliminary phytochemical investigation of ethanol extract of the leaves of Elaeocarpus serratus
+: Positive; -: Negetive

Table 2:	Brine shrimp lethality bioassays of ethanol extract of the leaves of Elaeocarpus serratus

Table 3:	Antimicrobial activity of ethanol extract of the leaves of Elaeocarpus serratus
Data were represented as Mean±SD of triplicate determination; (-): No inhibition

The results also showed that the extract did not elicit antibacterial activity against S. aureus, E. coli and S. dysenteriae. The maximum zone of inhibition was recorded against S. typhi (32.95±0.39 mm) which was almost similar to that of the standard drug, levofloxacin.