General experimental procedures- All the solvents and chemicals used for this research were analytical reagent grade, procured from E. Merck (Germany), BDH (England), AppliChem (Germany) and Sigma Aldrich (Germany). Gas chromatography analyses were performed with a SHIMADZU 2010 Plus gas chromatograph equipped with a flame ionization detector (FID) and a fused silica (5% phenyl/95% polydimethylsiloxane) capillary column (length 30m, inner diameter 0.25 mm, film thickness 0.25 μm) using hydrogen as carrier gas (1.0 ml/min). The injector temperature was 250°C and the column oven was programmed between 50-220 °C at 4 °C/min. The detector (FID) was operated at 260 °C. The absorbance of prepared solutions (extractive or control) of different concentrations for antioxidant activity was performed by using a Parkin Elmer Lambda-25 UV-VIS spectrophotometer (USA). Quartz cells (1 cm × 1 cm) were used as sample holder to record the spectrum. Flowers of Nyctanthes arbor-tristis were collected from BCSIR campus, Dhaka, Bangladesh. The flowers were dried, powdered and extracted successively with n-hexane, dichloromethane, ethyl acetate and methanol at room temperature according to the published procedure (Haque et al., 2019). The resulting extracts were filtered, concentrated, dried and stored in a desiccator for use in subsequent experiments. Identification and quantification of fatty acids-The esterification of fat was carried out by a modified procedure using trifluoride methanol (BF3–MeOH) complex (Griftin, 1960; Metecalfe and Schmitz, 1961; AOAC, 1984). The n-hexane extract (200 mg) was methylated by heating with BF3-CH3OH reagent (5 ml) for 10 min. Methyl esters of fatty acids were isolated by partitioning between water and n-hexane. The esterified fatty acids were taken for GC analysis. Fatty acid methyl esters (Sigma-Aldrich) of capric acid, caprilic acid, lauric acid, myristic acid, palmotelic acid, palmitic acid, linolic acid, oleic acid, stearic acid, arachidic acid, behenic acids and lignoceric acid were used as standard for the identification of sample peaks. The fatty acids were identified by comparison of retention times with the standard fatty acids chromatogram. The peak areas were calculated by software of the instrument. The relative percentages of fatty acids were calculated by using the following formula: Relative % of individual fatty acid = (Individual area/ Total areas for all fatty acids) × 100; Brine shrimp lethality bioassay- Brine shrimp lethality bioassay of crude extracts of n-hexane, dichloromethane, ethyl acetate and methanol were screened for cytotoxicity by the Mayer’s method (Mayer et al., 1982; McLaughlin et al., 1998). Test samples of 4 mg were dissolved in 200 µL of pure dimethylsulfoxide (DMSO) to prepare stock solutions. Then 100 μL of stock sample solution was taken in a test tube each containing 5 ml of simulated seawater and 10 shrimp nauplii. The concentration of prepared sample solution in the first test tube was 400 µg/ml. Then a series of sample solutions of lower concentrations were prepared by consecutive dilution. In each case 100 μL sample was added to each test tube containing 5 ml of brine solution with 10 living nauplii and fresh 100 μL DMSO was added into the mother solution. Finally, the prepared sample concentrations in each test tube were 400, 200, 100, 50, 25, 12.5, 6.25, 3.125, 1.563, 0.781, 0.391 and 0.195 µg/ml. After 24 hours, test tubes were inspected using a magnifying glass and the number of survived nauplii of each test tube was counted visually. The mortality percentages of the nauplii at different concentrations were plotted against the logarithm of particular sample concentration to achieve LC50 value (the concentration when 50% of brine shrimp nauplii died). The LC50 values were calculated by windows Microsoft Excel 2007 software. Standard vincristine sulfate was used as a positive control to compare the results obtained for test samples. Antibacterial study- Antibacterial assay was determined by disc diffusion method (Bauer et al., 1966; Barry, 1980). Four gram-positive bacteria (Bacillus subtilis, Bacillus cereus, Staphylococcus aureus and Enterococcus fecalis) and four gram-negative bacteria (Salmonella typhi, Escherichia coli 12079, Salmonella enteritis and Pseudomonas) were taken for the analysis. Crude extracts of n-hexane, dichloromethane, ethyl acetate and methanol were taken for antimicrobial screening. Standard ciprofloxacin and tetracycline were used as positive control. Each sample and standards were weighed accurately, then dissolved in their required volume of specific solvent (used DMSO for all samples as well as standards). IC50 values were calculated as the concentration of each sample required to give 50% DPPH radical scavenging activity from the graph (linear regression curve) by excel 2007 Office software.