*Article* **Production of Omega-3 Fatty Acids from the Microalga** *Crypthecodinium cohnii* **by Utilizing Both Pentose and Hexose Sugars from Agricultural Residues**

**Georgia Asimakopoulou <sup>1</sup> , Anthi Karnaouri <sup>1</sup> , Savvas Staikos <sup>1</sup> , Stylianos D. Stefanidis <sup>2</sup> , Konstantinos G. Kalogiannis <sup>2</sup> , Angelos A. Lappas <sup>2</sup> and Evangelos Topakas 1,\***


**Abstract:** The core objective of this work was to take advantage of the unexploited wheat straw biomass, currently considered as a broadly available waste stream from the Greek agricultural sector, towards the integrated valorization of sugar streams for the microbial production of polyunsaturated omega-3 fatty acids (PUFAs). The OxiOrganosolv pretreatment process was applied using acetone and ethanol as organic solvents without any additional catalyst. The results proved that both celluloserich solid pulp and hemicellulosic oligosaccharides-rich aqueous liquid fraction after pretreatment can be efficiently hydrolyzed enzymatically, thus resulting in high yields of fermentable monosaccharides. The latter were supplied as carbon sources to the heterotrophic microalga *Crypthecodinium cohnii* for the production of PUFAs, more specifically docosahexaenoic acid (DHA). The solid fractions consisted mainly of hexose sugars and led to higher DHA productivity than their pentose-rich liquid counterparts, which can be attributed to the different carbon source and C/N ratio in the two streams. The best performance was obtained with the solid pulp pretreated with ethanol at 160 ◦C for 120 min and an O<sup>2</sup> pressure of 16 bar. The total fatty acids content reached 70.3 wt% of dried cell biomass, of which 32.2% was DHA. The total DHA produced was 7.1 mg per g of untreated wheat straw biomass.

**Keywords:** microalgae; DHA; lignocellulosic biomass; organosolv fractionation; liquid fraction; solid pulp; omega-3 fatty acids
