Inhibitory effect of chloroform on fermentative hydrogen and methane production from lipid-extracted microalgae

To improve the sustainability of microalgae as a bioenergy feedstock, lipid-extracted microalgae (LEM) are often further treated by anaerobic digestion (AD). However, the residual chloroform used for extracting lipids as a solvent could inhibit this process, an aspect that has not been studied to date. In this study, the inhibitory effect of chloroform on H₂ and CH₄ production was investigated by performing batch tests. To prepare the feedstock, Chlorella vulgaris was ultrasonicated and the supernatant was discarded after centrifugation. In case of H₂ production, it was found that the H₂ yield fell to almost half that of the control (15.6 mL H₂/g COD_added) at 100 mg CHCl₃/L. The reason for the decrease of the H₂ yield with the increase of chloroform level was due to the change of metabolites from acetate and butyrate to lactate via a non-hydrogenic reaction. In comparison with H₂ production, a much more severe inhibitory effect of chloroform on CH₄ production was observed. The inhibitor concentration (IC_{30, 60, and 90}) on H₂ production was 138, 319, and 622 mg CHCl₃/L, respectively, while concentrations of 15, 37, and 86 mg CHCl₃/L were obtained on CH₄ production. When the chloroform concentration was ≥25 mg/L on CH₄ production, more than 2 g COD/L of organic acids remained, resulting in a decrease of CH₄ yield. These findings indicate that the residual chloroform in LEM should be seriously considered to prevent possible microbial inhibition when designing a process for additional energy recovery from microalgae via AD.