Statistical optimization for precipitation of bioactive compounds from extracted Centella asiatica using gas anti‐solvent technique

Centella asiatica is a well-known herbal medicine used for antioxidant and antibacterial purposes. Main compounds in the group of triterpene saponins, namely, asiaticoside, asiatic acid, madecassoside, and madecassic acid are considered as the active compounds. These active ingredients were extracted from C. asiatica dried powder by the maceration method. The extracted solution was then used as a starting solution for the gas anti-solvent (GAS) precipitation. The effects of % ethanol in a mixed solvent of ethanol and methanol (0-100%), carbon dioxide flow rate (3-9 ml/min), and temperature (25-45 degrees C) on the amount of precipitates were investigated using a Box-Behnken design of experiments. It was found that % ethanol in the mixed solvent was significant at 5% significance level on madecassoside, asiaticoside, and total triterpene saponins. Higher amounts of total triterpene saponins could be precipitated when using low % ethanol and low temperature. At the optimal conditions, the amount of madecassoside, asiaticoside, madecassic acid, and asiatic acid were found to be 5,733 +/- 537, 2,948 +/- 249, 1,678 +/- 163, and 436 +/- 17 mu g/5 ml of extracted solution, respectively. In the antioxidant studies, the highest Trolox equivalent found in the GAS products using 2,2-diphenyl-1-picrylhydrazyl and 2,2-azion-bis-3-ethylbenzothiazo line-6-sulphonic acid assays was 5,622 +/- 336 and 10,729 +/- 820 mu g/5 ml of extracted solution. Practical applications The novelty of this work covers the feasibility of applying gas anti-solvent (GAS) technique for precipitation of bioactive compounds from extracted Centella asiatica and a statistical optimization of the operating parameters on the highest amount of precipitates. This work is important for extraction and purification applications focused on obtaining high purity of bioactive compounds from crude extracts. The use of dense CO2 as an anti-solvent not only requires mild operating conditions but also provides a solvent-free product. Therefore, the application of the GAS process can then be extended to precipitate active ingredients from other herbal extracts.