Nanocurcumin Preparation Protocol

Nanocurcumin was prepared according to methods described in our previous report (24 (link)). Briefly, 10 g of curcumin (Sigma, USA) was dissolved in 2.5 liters of distilled ethanol at room temperature and filtered to obtain a clear solution. This solution was then stirred in a high-speed homogenizer (T 25 digital Ultra-Turrax) at 12,000 to 15,000 rpm, and the required volume of Milli Q water containing 0.1% citric acid (Merck, India) was added to it slowly over a period of 1 h until the ethanol concentration reached 40% (vol/vol) and curcumin particles started to precipitate from the solution. The entire suspension was then homogenized over ice in a high-pressure homogenizer (Avestin C5 high-pressure homogenizer) at 30,000 lb/in² for 30 cycles. The aqueous suspension was then made to 0.1% polysorbate 80 (Sigma, USA), homogenized at 12,000 to 15,000 rpm (T 25 digital Ultra-Turrax; IKA, USA) again for 1 h, and filtered. The filtered slurry was dried at 80°C in an oven to obtain curcumin powder. The particle size was determined by using a high-resolution transmission electron microscope (JEM 2100F; JEOL, USA).