Parametric Studies and Semi-Continuous Harvesting Strategies for Enhancing CO2 Bio-Fixation Rate and High-Density Biomass Production Using Adaptive Laboratory-Evolved Chlorella vulgaris

This study adopts a biochemical approach to sequester CO₂ while producing biomass rich in protein and lipids, using an adapted strain of <i>Chlorella vulgaris</i> (ALE-<i>Cv</i>), which had previously evolved to tolerate a gas mixture containing 10% CO₂ and 90% air. The research studied the operating parameters of the batch photobioreactor for ALE-<i>Cv</i> to evaluate the effects of inoculum size, photoperiod, light intensity, pH of culture, and CO₂ supply rate on biomass productivity and CO₂ bio-fixation rate. The optimal conditions were identified as 16:8 h light-dark cycles, 5000 lux, pH 7, 20 mL of 10 g/L inoculum, and 0.6 VVM; the system achieved a maximum total biomass production of 7.03 ± 0.21 g/L with a specific growth rate of 0.712 day^-1, corresponding to a CO₂ bio-fixation of 13.4 ± 0.45 g/L in batch cultivation. While the pre-adapted strain of <i>Chlorella vulgaris</i> under the same operating conditions, except for the gas supply, which was air, achieved a maximum total biomass production of 0.52 ± 0.008 g/L, and the total CO₂ bio-fixation was 1.036 ± 0.021 g/L during 7-day cultivation. A novel semi-continuous harvesting process, with and without nutrient addition, was also investigated to maximise biomass yield and enable water recycling for culture media. The maximum biomass production in semi-continuous harvesting process with and without nutrition added was 5.29 ± 0.09 and 9.91 ± 0.11 g/L, while the total corresponding CO₂ bio-fixation was 9.70 ± 0.13 and 18.16 ± 0.11 g/L, respectively, during 15-day cultivation. The findings provide critical insights into enhancing CO₂ bio-fixation through adaptive evolution of ALE-<i>Cv</i> and offer optimal operational parameters for future large-scale microalgae cultivation. This research also links microalgae-based CO₂ sequestration to green technologies and the bioeconomy, highlighting its potential contribution to climate change mitigation while supporting environmental sustainability, food security, and ecosystem resilience.