Water reuse and growth inhibition mechanisms for cultivation of microalga Euglena gracilis

Background Microalgae can contribute to more than 40% of global primary biomass production and are suitable candidates for various biotechnology applications such as food, feed products, drugs, fuels, and wastewater treatment. However, the primary limitation for large-scale algae production is the fact that algae requires large amounts of fresh water for cultivation. To address this issue, scientists around the world are working on ways to reuse the water to grow microalgae so that it can be grown in successive cycles without the need for fresh water. Results In this study, we present the results when we cultivate microalgae with cultivation water that is purified and reused. Specifically, we purify the cultivation water using an ultrafiltration membrane (UFM) treatment and investigate how this treatment affects: the biomass and biochemical components of the microalgae; characteristics of microalgae growth inhibitors; the mechanism whereby potential growth inhibitors are secreted (followed using metabolomics analysis); the effect of activated carbon (AC) treatment and advanced oxidation processes (AOPs) on the removal of growth inhibitors of Euglena gracilis. Firstly, the results show that E. gracilis can be only cultivated through two growth cycles with water that has been filtered and reused, and the growth of E. gracilis is significantly inhibited when the water is used a third time. Secondly, as the number of reused water cycles increases, the Cl- concentration gradually increases in the cultivation water. When the Cl- concentration accumulates to a level of fivefold higher than that of the control, growth of E. gracilis is inhibited as the osmolality tolerance range is exceeded. Interestingly, the osmolality of the reused water can be reduced by replacing NH4Cl with urea as the source of nitrogen in the cultivation water. Thirdly, E. gracilis secretes humic acid (HA)-which is produced by the metabolic pathways for valine, leucine, and isoleucine biosynthesis and by linoleic acid metabolism-into the cultivation water. Because HA contains large fluorescent functional groups, specifically extended pi(pi)-systems containing C=C and C=O groups and aromatic rings, we were able to observe a positive correlation between HA concentration and the rate of inhibition of E. gracilis growth using fluorescence spectroscopy. Moreover, photosynthetic efficiency is adversely interfered by HA, thereby reductions in the synthetic efficiency of paramylon and lipid in E. gracilis. In this way, we are able to confirm that HA is the main growth inhibitor of E. gracilis. Finally, we verify that all the HA is removed or converted into nutrients efficiently by AC or UV/H2O2/O-3 treatments, respectively. As a result of these treatments, growth of E. gracilis is restored (AC treatment) and the amount of biomass is promoted (UV/H2O2/O-3 treatment). Conclusions These studies have important practical and theoretical significance for the cyclic cultivation of E. gracilis and for saving water resources. Our work may also provide a useful reference for other microalgae cultivation.

Automated summary

The study found that microalgae can only be cultivated through two growth cycles with purified and reused water, with significant growth inhibition observed in the third cycle. As the number of water reuse cycles increases, the Cl- concentration gradually rises in the cultivation water, leading to inhibition of microalgae growth. Additionally, the study identified humic acid (HA) as the main inhibitor of microalgae growth, and demonstrated that HA can be efficiently removed by treatments such as activated carbon and advanced oxidation processes.