Photo-fermentative bacteria aggregation triggered by L-cysteine during hydrogen production

Abstract

Background: Hydrogen recovered from organic wastes and solar energy by photo-fermentative bacteria (PFB) has been suggested as a promising bioenergy strategy. However, the use of PFB for hydrogen production generally suffers from a serious biomass washout from photobioreactor, due to poor flocculation of PFB. In the continuous operation, PFB cells cannot be efficiently separated from supernatant and rush out with effluent from reactor continuously, which increased the effluent turbidity, meanwhile led to increases in pollutants. Moreover, to replenish the biomass washout, substrate was continuously utilized for cell growth rather than hydrogen production. Consequently, the poor flocculability not only deteriorated the effluent quality, but also decreased the potential yield of hydrogen from substrate. Therefore, enhancing the flocculability of PFB is urgent necessary to further develop photo-fermentative process.

Results: Here, we demonstrated that L-cysteine could improve hydrogen production of Rhodopseudomonas faecalis RLD-53, and more importantly, simultaneously trigger remarkable aggregation of PFB. Experiments showed that L-cysteine greatly promoted the production of extracellular polymeric substances, especially secretion of protein containing more disulfide bonds, and help for enhancement stability of floc of PFB. Through formation of disulfide bonds, L-cysteine not only promoted production of EPS, in particular the secretion of protein, but also stabilized the final confirmation of protein in EPS. In addition, the cell surface elements and functional groups, especially surface charged groups, have also been changed by L-cysteine. Consequently, absolute zeta potential reached a minimum value at 1.0 g/l of L-cysteine, which obviously decreased electrostatic repulsion interaction energy based on DLVO theory. Total interaction energy barrier decreased from 389.77 KT at 0.0 g/l of L-cysteine to 127.21 kT at 1.0 g/l.

Conclusions: Thus, the strain RLD-53 overcame the total energy barrier and flocculated effectively. After a short settlement, the biomass rush out will be significantly reduced and the effluent quality will be greatly improved in the continuous operation. Furthermore, aggregation of PFB could enable high biomass hold-up of photobioreactor, which allows the photobioreactor to operate at low hydraulic retention time and high organic loading rate. Therefore, the described flocculation behaviour during photo-hydrogen production is potentially suitable for practicable application.

Figure 1

Hydrogen production kinetics of R. faecalis RLD-53 at different L-cysteine concentrations.

Figure 2

Cell growth and flocculability of R. faecalis RLD-53 at different L-cysteine concentrations.

Figure 3

Bioflocculation of R. faecalis RLD-53 at different concentration of L-cysteine. (a) photo of bioflocculation; (b) SEM images of bioflocculation.

Figure 4

EPS compositions of R. faecalis RLD-53 at various concentration of L-cysteine. (a), EPS component; (b), thiol group (SH) and disulfide bond (SS) content in EPS.

Figure 5

Relationship between disulfide bonds and components of EPS production. (a), Nucleic acid; (b), Humic substances; (c), Proteins; (d), Polysaccharides.

Figure 6

Spectral decomposition of the amide I bands at different concentration of L-cysteine (g/l). (a), 0.0; (b), 0.5; (c), 1.0; (d), 1.5.

Figure 7

Contribution of EPS protein conformations to flocculability of R. faecalis RLD-53. (a), Aggregated strands; (b), β-Sheet; (c), Random coil; (d), α-Helix; (e), 3-Turn helix; (f), Antiparallel β-sheet/aggregated strands.

Figure 8

Contribution of cell surface functional groups to flocculability of R. faecalis RLD-53. (a), C-(C, H); (b), C-(O, N); (c), C=O, O-C-O; (d), O=C-OH, O=C-OR; (e), C=O; (f), C-OH, C-O-C; (g), C-NH₂; (h), O=C-NH-R.

Figure 9

Effect of charged groups on the zeta potential of R. faecalis RLD-53. (a), COOH, COOR; (b), C-NH₂.

Figure 10

Contact angle and surface thermodynamic properties of R. faecalis RLD-53. (a), Contact angle; (b), surface thermodynamic properties.

Figure 11

Interaction energy profiles as a function of cells distance at various L-cysteine concentrations (g/l). (a), 0.0; (b), 0.5; (c), 1.0; (d), 1.5.