Fermentation parameters determination
Fermentation parameters containing the concentrations of residual
glucose, ethanol, glycerol, succinate, biomass, cell viability, and
oxidation reduction potential (ORP) were investigated to assess the CVEF
system.
Triplicate samples were withdrawn and centrifuged at 10,000 rpm at
ambient temperature for 5 min. The cell pellet was collected for biomass
determination, and the supernatant was frozen at −20 °C for HPLC
analysis. An HPLC system with RI-detector (Waters 410, Waters, USA) and
an Aminex HPX 87-H column (300 ×7.8 mm, Bio-Rad Laboratories, Hercules,
CA, USA) was used to analyze glucose, ethanol, glycerol, and succinate
in the effluent according to Wang et al.(2013). Biomass was measured as
dry cell weight (DCW), wash the cell pellet obtained previously twice
with deionized water, overnight at 80°C to constant weight, and weigh
the DCW. Cell viability was evaluated via methylene blue stain
described by Bai et al. (2004). Living cells were calculated from
biomass×viability. ORP value was monitored by an ORP electrode.