Lyophilizer

To prepare stigmasterol nano-emulsion, 8 g of stigmasterol was added to 80 ml of water and heated at 30°C. To the mixture 6 g of lecithin and 6 ml of olive oil was added and vortexed for 5 min. The mixture was stirred using a magnetic hot plate set at a temperature of 50°C for 10 min and blended using a homogenizer at 6,000 rpm for 2 min and then fine nano emulsions were obtained using a high-pressure homogenizer. The formed emulsion was then freeze dried using a lyophilizer (Yamato, Japan). Lyophilized powder obtained was characterized for particle size analysis and its formulation into biscuit as a functional food ingredient.

The CFS of MSI45 was mixed with an equal volume of six different solvents such as chloroform, ethyl acetate, ethanol, dimethyl ether, dimethyl sulfoxide and methanol in separate extraction flasks and kept for overnight incubation at 4 °C. The solvents were then evaporated in a rotary evaporator (Yamato) and the remaining residues were concentrated in vacuum concentrator (Labconco) and lyophilised in a lyophilizer (Yamato). The lyophilised compounds were dissolved at a concentration of 50 mg L⁻¹ and from this 100 μl of the compound MSI45 was tested for antimicrobial activity using a well diffusion assay.