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A Comparative Phytochemical and Biological Study Between Different Solvent Extracts of Bombax Ceiba Roots Available in Bangladesh

Nazia Hoque*
Department of Pharmacy, East West University, Dhaka, Bangladesh

Sabera Rahman
Department of Pharmacy, East West University, Dhaka, Bangladesh

Ishrat Jahan
Department of Pharmacy, East West University, Dhaka, Bangladesh

Meena Afroze Shanta
Department of Pharmacy, East West University, Dhaka, Bangladesh

Nigar Sultana Tithi
Department of Pharmacy, East West University, Dhaka, Bangladesh

Nishat Nasrin
Department of Pharmacy, East West University, Dhaka, Bangladesh

Abstract/ Executive Summary:

Use of different solvent systems for extraction of plant materials may cause variation in their bioactivities. The present study was conducted to evaluate the presence of different phytoconstituents and to compare in vitro bioactivities of petroleum ether, dichloromethane (DCM) and methanol extracts of Bombax ceiba (B. ceiba) roots available in Bangladesh. Preliminary phytochemical screening was conducted using specific standard procedure. Antioxidant activity of the extracts was evaluated using DPPH radical scavenging assay. Determination of total phenolic and flavonoid content was also carried out. Antibacterial and cytotoxic activities were investigated using disc diffusion method and brine shrimp lethality bioassay, respectively. All the experiments were carried out from February 2016 to September 2016. Phytochemical evaluation revealed the presence of alkaloids, terpenoids, carbohydrates, tannins, flavonoids, saponins and steroids. The methanol extract showed the highest DPPH radical scavenging activity and had the highest phenolic (187.42 ± 3.77 mg/g, GAE) and flavonoid content (74.67 ± 4 mg/g, QE) followed by the DCM and petroleum ether extracts. The extracts showed positive correlation between DPPH radical scavenging activity with the phenolic and flavonoid content. All the extracts showed mild to moderate in vitro antibacterial activity with zone of inhibition ranging from 7 mm to 13 mm. In brine shrimp lethality bioassay, the observed LC50 values for petroleum ether, DCM and methanol extracts were 70.72 μg/ml, 37.72 μg/ml and 22.58 μg/ml, respectively which revealed strong cytotoxic potential of the extracts compared to the positive control. The results indicated that B. ceiba roots could be a very potent source of natural radical scavenger and cytotoxic agent.

Keywords: Bombax ceiba, Antioxidant Activity, DPPH, Antibacterial Activity, Cytotoxic Effect

Location: Department of Pharmacy, East West University, Dhaka, Bangladesh

Start Date:

End Date:

Program Area: Development of Host and Medicinal Plants

Commodity: Medicinal Plants

Objectives:

An attempt to evaluate the variation in pharmacological activities due to different solvent system, we have reported here a study on the antioxidant, antimicrobial and cytotoxic activities of the different solvent extracts of B. ceiba roots.

Methodology:

2.1. Chemicals and Solvents Chemicals and reagents were obtained from Sigma-Aldrich co. (USA), Merck (Germany) and Fine Chemicals (India). All the solvents used throughout the experiments were of analytical grades. 2.2. Plant Collection and Extraction of the Plant Material The plant material was collected in February, 2016 from Rampura, Dhaka, Bangladesh and was identified and authenticated by Mr. Abdur Rahim, Technical Officer, Department of Botany, Jahangirnagar University. The roots were dried and pulverized into coarse powder. Two hundred grams of powdered plant materials were taken into sealed container and macerated for seven days at room temperature using 1.0 liter of petroleum ether with occasional shaking and stirring. Same procedure was repeated using dichloromethane (DCM) and methanol as solvents. The extracts were then filtered using sterilized cotton filter followed by whatman no 1 filter papers. The solvents were totally evaporated using rotary evaporator under reduced pressure at 40⁰C - 50⁰C temperature. The obtained crude extracts were collected, weighed and then stored at 4⁰C until further use. Table 1 shows the yield percentages (w/w) of B. ceiba roots in different solvents used for extraction. 2.3. Phytochemical Screening Chemical tests were carried out on the freshly prepared extracts using standard procedures to assess the existence of active phytochemical constituents. The following reagents and chemicals were used: alkaloids with Wagner’s and Hager’s reagent, carbohydrates with Molisch’s test, tannins with 0.1% ferric chloride, terpenoids with modified Salkowski test, flavonoids with the use of concentrated hydrochloric acid, saponins with distilled water and glycosides by using alcohol with few drops of H2SO4, followed by neutralization with NaOH solution and boiling with Fehling’s solution. These were identified by characteristic color changes using standard procedures. 2.4.1. Determination of Total Phenolic Content To determine the total phenolic content, 1 ml of each extract (250 μg/ml) and the standard gallic acid (50 - 250 μg/ml) were oxidized with 10% Folin-Ciocalteu reagent and then neutralized with 700 mM sodium carbonate solution. The obtained mixtures were allowed to stand for 60 minutes at room temperature and absorbance of the resulting solutions was taken at 765 nm against reagent blank. A standard curve was prepared for gallic acid and used to determine the total phenolic content of the extracts. The results were expressed as gallic acid equivalents (mg/g, GAE) and as mean ± SD. 2.4.2. Determination of Total Flavonoid Content Aluminium chloride colorimetric method [18] was used to determine the flavonoid content. One milliliter of each extract (250 μg/ml) in methanol was mixed with 200 μl of 10% w/v aluminium chloride and 1 M potassium acetate solution. Then 5.6 ml distilled water was added to the mixture and was kept at room temperature for 40 minutes. The absorbance of the resulting solutions was measured at 415 nm against reagent blank. Total flavonoid content of the extracts was determined from a standard curve prepared for quercetin (50 - 250 μg/ml) solution (Figure 2). The estimated results were expressed as quercetin equivalents (mg/g, QE).

DPPH Radical Scavenging Activity Methanol was used to prepare a stock solution (400 µg/ml) of 2, 2-Diphenyl-1-picrylhydrazyl (DPPH) and 100 µl of this stock solution was added to 5 ml solution of the extracts and the standard (ascorbic acid) of different concentrations (12.5 - 200 µg/ml). After proper mixing the solutions were kept in dark for 20 minutes and by using a spectrophotometer (Shimadzu UV PC-1800), the absorbance of the solutions were measured at 517 nm against methanol. Scavenging activity was measured by reduction in the absorbance of methanol solution of DPPH and expressed as the percentage inhibition using the formula: ( 010 - AAA − ×100 where A0 is the absorbance of the control and A1 is the absorbance of the extract/standard. IC50 values were calculated from the graph constructed as a plot of % inhibition vs. concentration. 2.6. Antibacterial Assay Five Gram-positive and six Gram-negative bacterial strains were used in the antibacterial assay following the disc diffusion method. Suspension of each microorganism (100 µl, containing approximately 100 - 150 CFU/ml) was spread over nutrient agar media and dried, sterile filter paper discs of 6 mm diameter were placed gently on those agar plates after impregnated with 300 µg and 600 µg of different extracts prepared by dissolving them in methanol followed by evaporation. After 24 hours of incubation at 37⁰C the diameter of zone of inhibition was measured in mm. Kanamycin (30 µg/disc) was used as positive control while blank disc (impregnated with solvents) was used as negative control.

Source of Information: Pharmacology & Pharmacy, 2018, 9, 53-66; ISSN Online: 2157-9431 ISSN Print: 2157-9423

Article Link (Online Journal): http://www.scirp.org/journal/pp

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

The results obtained from the study showed that different extracts of B. ceiba roots possess potential antioxidant, antibacterial and cytotoxic properties which demonstrate clearly the scientific basis of traditional uses of the plant. So, the present study suggests that B. ceiba roots can be a useful source to lead the development of new drugs. Further studies are required to establish its antioxidant activity by other in vitro and in vivo methods, cytotoxic activity against different cancer cell lines and to identify the chemical compounds responsible for such activities.

Knowledge Management: Journal