Optimization of green extractions for the recovery of docosahexaenoic acid (DHA) from Crypthecodinium cohnii

Crypthecodinium cohnii represents a heterotrophic microalga rich in omega-3 polyunsaturated fatty acids (PUFAs) with significant health benefits, and especially docosahexaenoic acid (DHA). In the present study, freeze-dried C. cohnii biomass, cultivated in fed-batch bioreactors with a feed of acetic acid 33% (v/v), was used. In the first set of experiments, ultrasound assisted extraction (UAE) and microwave assisted extraction (MAE) were employed with the aim of discovering the most efficient technique regarding the recovery of DHA. Organic and food grade solvents were used, namely n-hexane, 2-2-butanol and n-hexane: isopropanol (2:3), under specific conditions. Extraction yields (EYs) were calculated and PUFAs content was evaluated through Gas Chromatography (GC). UAE using n-hexane: isopropanol (2:3) and 2-butanol presented the highest yields and were further examined in the second set of experiments in order to optimize the recovery of DHA, while changing the extraction time and the ultrasound power. Enzymatic Hydrolysis (EH) was also applied using four enzyme preparations, three concentrations of C. cohnii biomass and n-hexane: isopropanol (2:3) as solvent. In addition, homogenisation assisted extraction in microemulsion (HAE) was studied while changing the solvent, time and rotation speed. Simple mathematical models were applied in order to correlate the EY with the applied conditions for every extraction method. Parameters screening during UAE demonstrated that the best extraction time was 15 min, reaching the yield of 26.17% and total fatty acids (TFAs) content of 261.75 mg/g dry weight (DW). Among all used extraction techniques, UAE recovered maximum quantities of TFAs, while HAE was proven the most efficient regarding DHA content (131.20 mg/g DW). In conclusion, the proposed green extraction methods were found to be attractive for the recovery of PUFAs from C. cohnii.