Pulsed Electric Field (PEF) treatment was used as pre-treatment on the microalgae strain Auxenochlorella protothecoides (A.p.) prior to organic solvent extraction of lipids. Experiments were performed on fresh biomass from mixotrophic or autotrophic culture which both had an evaluated lipid content of 30-35 % of cell dry weight. Lipid yield was determined gravimetrically and compared to the reference lipid content assessed by bead-milling and subsequent Soxhlet extraction. The biomass was concentrated at 10 % w/w solids prior to PEFtreatment and further dewatered afterwards to approximately 25 % w/w before extraction. PEFtreatment with an energy input of 1.5 MJ per kilogram of dry matter induced electropermeabilisation of the microalgae cells detected by the increase of the conductivity of the microalgae supernatant. This greatly increased the lipid yield upon subsequent monophasic solvent extraction. A mixture of Water/Ethanol/Hexane 1:18:7.3 vol/vol/vol enabled to recover 92 %, and 72 %, of the evaluated lipid content of mixotrophically, and autotrophically respectively, grown A.p., after two hours of extraction. Recovery increased to 97 %, and 90 % respectively, after 20 hours of extraction. The same extraction system on untreated biomass yielded maximum 10 % of lipid content. The highest yields were obtained with 80 mL of solvent for 1 g dry biomass but solvent volume could be reduced by a factor two in case of mixotrophically grown microalgae. However, the solvent:biomass ratio still remains high, and includes a water-miscible solvent, ethanol. Total lipid extraction was confirmed by nile red staining of residual biomass combined with fluorescence microscopy imaging and flow cytometry. Gas chromatography analyses of extracted lipids after transesterification revealed that PEF-treatment did not alter their fatty acid composition. Overall PEF-treatment shows promising features for upscaling especially in a biorefinery concept since it avoids potentially harmfull temperature increase and small debris problematic for further processing.

No abstract

Dichloromethane (DCM) is a solvent commonly used in laboratories for microalgae hydrothermal liquefaction (HTL) product separation. The addition of DCM would lead to an "overestimation effect" of biocrude yield and diminish biocrude quality. However, it is currently not clear to what extent this overestimation effect will impact a continuous HTL process. In this study, Chlorella vulgaris microalgae was processed in a continuous stirred tank reactor at different temperatures (300, 325, 350, 375, and 400 °C) at 24 MPa for 15 min holding time. Two separation methods were applied to investigate the effect of using DCM in a cHTL product separation procedure in terms of product yield, biocrude elemental content, and aqueous product (AP) composition. Subsequently, the feasibility of reusing AP for algae cultivation has been evaluated. Results suggest that 350 °C is the optimal temperature for cHTL operation, leading to the highest biocrude yield, and an average increase in biocrude yield of 9 wt % was achieved when using DCM in cHTL product separation. Within the temperature range investigated, an average biocrude yield estimation can be proposed by yield non-DCM ≈ 0.818 × yield DCM . The AP has been characterized by total organic carbon and total nitrogen, high-performance liquid chromatography, and inductively coupled plasma optical emission spectroscopy. Results show that at 350−375 °C more nitrogen and other ions were directed into the AP, which could be advantageous in nutrient recovery. With the help of optical density testing, algae was shown to exhibit a better growth using AP with activated carbon absorption purification treatment as compared to the standard medium. The recovery of water and nutrients from the HTL-AP could improve the economics of a microalgae biorefinery process.