Fungal and Algal Biomass Extraction

Fungal biomass from MTPS and 1.5 L cultivations were filtered through a Whatman No. I filter paper in a vacuum setup (GE Whatman, Maidstone, UK), while in case of the 25 L cultivations, a 75-μm aperture test sieve was used (Endecotts, London, UK) for biomass separation. After filtration, the fungal biomass was washed thoroughly with distilled water. In case of microalga C. cohnii, the biomass was separated from the medium by centrifugation at 3000 rpm and it was washed once with distilled water. In the next step, the fungal and algal biomass was frozen at − 20 °C and then was lyophilized overnight in an Alpha 1-2 LDPlus freeze-dryer (Martin Christ, Osterode am Harz, Germany) at − 55 °C and 0.01 mbar pressure. The freeze-dried biomass was also used to calculate cell dry weight (CDW, g/L). Approximately 10 mg of freeze-dried biomass was transferred into 2-mL screw-cap tubes containing 500 μL distilled water and 250 ± 30 mg acid-washed glass beads (800 μm, OPS Diagnostics, Lebanon, USA). Biomass was then homogenized for 1–2 min in a FastPrep-24 high-speed benchtop homogenizer (MP Biomedicals, USA) at 6.5 m s⁻¹. This homogenized fungal suspension was used for HTS-FTIR analysis. Lipid extraction protocol was performed according to previously described protocol (Kosa et al. 2017 (link)).